WorldWideScience

Sample records for rna-silencing suppressor activity

  1. Suppressors of RNA silencing encoded by tomato leaf curl ...

    Indian Academy of Sciences (India)

    2013-01-06

    Jan 6, 2013 ... Virus encoded RNA-silencing suppressors (RSSs) are the key components evolved by the viruses to ... severe disease symptom in the host (Briddon et al. ..... Voinnet O 2001 RNA silencing as a plant immune system against.

  2. Supervised learning classification models for prediction of plant virus encoded RNA silencing suppressors.

    Directory of Open Access Journals (Sweden)

    Zeenia Jagga

    Full Text Available Viral encoded RNA silencing suppressor proteins interfere with the host RNA silencing machinery, facilitating viral infection by evading host immunity. In plant hosts, the viral proteins have several basic science implications and biotechnology applications. However in silico identification of these proteins is limited by their high sequence diversity. In this study we developed supervised learning based classification models for plant viral RNA silencing suppressor proteins in plant viruses. We developed four classifiers based on supervised learning algorithms: J48, Random Forest, LibSVM and Naïve Bayes algorithms, with enriched model learning by correlation based feature selection. Structural and physicochemical features calculated for experimentally verified primary protein sequences were used to train the classifiers. The training features include amino acid composition; auto correlation coefficients; composition, transition, and distribution of various physicochemical properties; and pseudo amino acid composition. Performance analysis of predictive models based on 10 fold cross-validation and independent data testing revealed that the Random Forest based model was the best and achieved 86.11% overall accuracy and 86.22% balanced accuracy with a remarkably high area under the Receivers Operating Characteristic curve of 0.95 to predict viral RNA silencing suppressor proteins. The prediction models for plant viral RNA silencing suppressors can potentially aid identification of novel viral RNA silencing suppressors, which will provide valuable insights into the mechanism of RNA silencing and could be further explored as potential targets for designing novel antiviral therapeutics. Also, the key subset of identified optimal features may help in determining compositional patterns in the viral proteins which are important determinants for RNA silencing suppressor activities. The best prediction model developed in the study is available as a

  3. A viral suppressor protein inhibits host RNA silencing by hooking up with Argonautes

    KAUST Repository

    Jin, Hailing; Zhu, Jian-Kang

    2010-01-01

    RNA viruses are particularly vulnerable to RNAi-based defenses in the host, and thus have evolved specific proteins, known as viral suppressors of RNA silencing (VSRs), as a counterdefense. In this issue of Genes & Development, Azevedo and colleagues (pp. 904-915) discovered that P38, the VSR of Turnip crinkle virus, uses its glycine/tryptophane (GW) motifs as an ARGONAUTE (AGO) hook to attract and disarm the host's essential effector of RNA silencing. Several GW motif-containing cellular proteins are known to be important partners of AGOs in RNA silencing effector complexes in yeast, plants, and animals. The GW motif appears to be a versatile and effective tool for regulating the activities of RNA silencing pathways, and the use of GW mimicry to compete for and inhibit host AGOs may be a strategy used by many pathogens to counteract host RNAi-based defenses. © 2010 by Cold Spring Harbor Laboratory Press.

  4. A viral suppressor protein inhibits host RNA silencing by hooking up with Argonautes

    KAUST Repository

    Jin, Hailing

    2010-05-01

    RNA viruses are particularly vulnerable to RNAi-based defenses in the host, and thus have evolved specific proteins, known as viral suppressors of RNA silencing (VSRs), as a counterdefense. In this issue of Genes & Development, Azevedo and colleagues (pp. 904-915) discovered that P38, the VSR of Turnip crinkle virus, uses its glycine/tryptophane (GW) motifs as an ARGONAUTE (AGO) hook to attract and disarm the host\\'s essential effector of RNA silencing. Several GW motif-containing cellular proteins are known to be important partners of AGOs in RNA silencing effector complexes in yeast, plants, and animals. The GW motif appears to be a versatile and effective tool for regulating the activities of RNA silencing pathways, and the use of GW mimicry to compete for and inhibit host AGOs may be a strategy used by many pathogens to counteract host RNAi-based defenses. © 2010 by Cold Spring Harbor Laboratory Press.

  5. ABCE1 is a highly conserved RNA silencing suppressor.

    Directory of Open Access Journals (Sweden)

    Kairi Kärblane

    Full Text Available ATP-binding cassette sub-family E member 1 (ABCE1 is a highly conserved protein among eukaryotes and archaea. Recent studies have identified ABCE1 as a ribosome-recycling factor important for translation termination in mammalian cells, yeast and also archaea. Here we report another conserved function of ABCE1. We have previously described AtRLI2, the homolog of ABCE1 in the plant Arabidopsis thaliana, as an endogenous suppressor of RNA silencing. In this study we show that this function is conserved: human ABCE1 is able to suppress RNA silencing in Nicotiana benthamiana plants, in mammalian HEK293 cells and in the worm Caenorhabditis elegans. Using co-immunoprecipitation and mass spectrometry, we found a number of potential ABCE1-interacting proteins that might support its function as an endogenous suppressor of RNA interference. The interactor candidates are associated with epigenetic regulation, transcription, RNA processing and mRNA surveillance. In addition, one of the identified proteins is translin, which together with its binding partner TRAX supports RNA interference.

  6. The Ebola virus VP35 protein is a suppressor of RNA silencing.

    Directory of Open Access Journals (Sweden)

    Joost Haasnoot

    2007-06-01

    Full Text Available RNA silencing or interference (RNAi is a gene regulation mechanism in eukaryotes that controls cell differentiation and developmental processes via expression of microRNAs. RNAi also serves as an innate antiviral defence response in plants, nematodes, and insects. This antiviral response is triggered by virus-specific double-stranded RNA molecules (dsRNAs that are produced during infection. To overcome antiviral RNAi responses, many plant and insect viruses encode RNA silencing suppressors (RSSs that enable them to replicate at higher titers. Recently, several human viruses were shown to encode RSSs, suggesting that RNAi also serves as an innate defence response in mammals. Here, we demonstrate that the Ebola virus VP35 protein is a suppressor of RNAi in mammalian cells and that its RSS activity is functionally equivalent to that of the HIV-1 Tat protein. We show that VP35 can replace HIV-1 Tat and thereby support the replication of a Tat-minus HIV-1 variant. The VP35 dsRNA-binding domain is required for this RSS activity. Vaccinia virus E3L protein and influenza A virus NS1 protein are also capable of replacing the HIV-1 Tat RSS function. These findings support the hypothesis that RNAi is part of the innate antiviral response in mammalian cells. Moreover, the results indicate that RSSs play a critical role in mammalian virus replication.

  7. The Enamovirus P0 protein is a silencing suppressor which inhibits local and systemic RNA silencing through AGO1 degradation

    International Nuclear Information System (INIS)

    Fusaro, Adriana F.; Correa, Regis L.; Nakasugi, Kenlee; Jackson, Craig; Kawchuk, Lawrence; Vaslin, Maite F.S.; Waterhouse, Peter M.

    2012-01-01

    The P0 protein of poleroviruses and P1 protein of sobemoviruses suppress the plant's RNA silencing machinery. Here we identified a silencing suppressor protein (SSP), P0 PE , in the Enamovirus Pea enation mosaic virus-1 (PEMV-1) and showed that it and the P0s of poleroviruses Potato leaf roll virus and Cereal yellow dwarf virus have strong local and systemic SSP activity, while the P1 of Sobemovirus Southern bean mosaic virus supresses systemic silencing. The nuclear localized P0 PE has no discernable sequence conservation with known SSPs, but proved to be a strong suppressor of local silencing and a moderate suppressor of systemic silencing. Like the P0s from poleroviruses, P0 PE destabilizes AGO1 and this action is mediated by an F-box-like domain. Therefore, despite the lack of any sequence similarity, the poleroviral and enamoviral SSPs have a conserved mode of action upon the RNA silencing machinery.

  8. The Luteovirus P4 Movement Protein Is a Suppressor of Systemic RNA Silencing.

    Science.gov (United States)

    Fusaro, Adriana F; Barton, Deborah A; Nakasugi, Kenlee; Jackson, Craig; Kalischuk, Melanie L; Kawchuk, Lawrence M; Vaslin, Maite F S; Correa, Regis L; Waterhouse, Peter M

    2017-10-10

    The plant viral family Luteoviridae is divided into three genera: Luteovirus , Polerovirus and Enamovirus . Without assistance from another virus, members of the family are confined to the cells of the host plant's vascular system. The first open reading frame (ORF) of poleroviruses and enamoviruses encodes P0 proteins which act as silencing suppressor proteins (VSRs) against the plant's viral defense-mediating RNA silencing machinery. Luteoviruses, such as barley yellow dwarf virus-PAV (BYDV-PAV), however, have no P0 to carry out the VSR role, so we investigated whether other proteins or RNAs encoded by BYDV-PAV confer protection against the plant's silencing machinery. Deep-sequencing of small RNAs from plants infected with BYDV-PAV revealed that the virus is subjected to RNA silencing in the phloem tissues and there was no evidence of protection afforded by a possible decoy effect of the highly abundant subgenomic RNA3. However, analysis of VSR activity among the BYDV-PAV ORFs revealed systemic silencing suppression by the P4 movement protein, and a similar, but weaker, activity by P6. The closely related BYDV-PAS P4, but not the polerovirus potato leafroll virus P4, also displayed systemic VSR activity. Both luteovirus and the polerovirus P4 proteins also showed transient, weak local silencing suppression. This suggests that systemic silencing suppression is the principal mechanism by which the luteoviruses BYDV-PAV and BYDV-PAS minimize the effects of the plant's anti-viral defense.

  9. Yellow fever virus capsid protein is a potent suppressor of RNA silencing that binds double-stranded RNA.

    Science.gov (United States)

    Samuel, Glady Hazitha; Wiley, Michael R; Badawi, Atif; Adelman, Zach N; Myles, Kevin M

    2016-11-29

    Mosquito-borne flaviviruses, including yellow fever virus (YFV), Zika virus (ZIKV), and West Nile virus (WNV), profoundly affect human health. The successful transmission of these viruses to a human host depends on the pathogen's ability to overcome a potentially sterilizing immune response in the vector mosquito. Similar to other invertebrate animals and plants, the mosquito's RNA silencing pathway comprises its primary antiviral defense. Although a diverse range of plant and insect viruses has been found to encode suppressors of RNA silencing, the mechanisms by which flaviviruses antagonize antiviral small RNA pathways in disease vectors are unknown. Here we describe a viral suppressor of RNA silencing (VSR) encoded by the prototype flavivirus, YFV. We show that the YFV capsid (YFC) protein inhibits RNA silencing in the mosquito Aedes aegypti by interfering with Dicer. This VSR activity appears to be broadly conserved in the C proteins of other medically important flaviviruses, including that of ZIKV. These results suggest that a molecular "arms race" between vector and pathogen underlies the continued existence of flaviviruses in nature.

  10. Suppressors of RNA silencing encoded by tomato leaf curl

    Indian Academy of Sciences (India)

    Whitefly-transmitted begomoviruses infecting tomato crop code for five different proteins, ORF AC4, ORF AC2 and ORF AV2 in DNA-A component, ORF BV1 in DNA-B ... In the present study suppressor function of ORF C1 of three betasatellites Tomato leaf curl Bangalore betasatellite ToLCBB-[IN:Hess:08], Cotton leaf curl ...

  11. The Luteovirus P4 Movement Protein Is a Suppressor of Systemic RNA Silencing

    Directory of Open Access Journals (Sweden)

    Adriana F. Fusaro

    2017-10-01

    Full Text Available The plant viral family Luteoviridae is divided into three genera: Luteovirus, Polerovirus and Enamovirus. Without assistance from another virus, members of the family are confined to the cells of the host plant’s vascular system. The first open reading frame (ORF of poleroviruses and enamoviruses encodes P0 proteins which act as silencing suppressor proteins (VSRs against the plant’s viral defense-mediating RNA silencing machinery. Luteoviruses, such as barley yellow dwarf virus-PAV (BYDV-PAV, however, have no P0 to carry out the VSR role, so we investigated whether other proteins or RNAs encoded by BYDV-PAV confer protection against the plant’s silencing machinery. Deep-sequencing of small RNAs from plants infected with BYDV-PAV revealed that the virus is subjected to RNA silencing in the phloem tissues and there was no evidence of protection afforded by a possible decoy effect of the highly abundant subgenomic RNA3. However, analysis of VSR activity among the BYDV-PAV ORFs revealed systemic silencing suppression by the P4 movement protein, and a similar, but weaker, activity by P6. The closely related BYDV-PAS P4, but not the polerovirus potato leafroll virus P4, also displayed systemic VSR activity. Both luteovirus and the polerovirus P4 proteins also showed transient, weak local silencing suppression. This suggests that systemic silencing suppression is the principal mechanism by which the luteoviruses BYDV-PAV and BYDV-PAS minimize the effects of the plant’s anti-viral defense.

  12. Alfalfa dwarf cytorhabdovirus P protein is a local and systemic RNA silencing supressor which inhibits programmed RISC activity and prevents transitive amplification of RNA silencing.

    Science.gov (United States)

    Bejerman, Nicolás; Mann, Krin S; Dietzgen, Ralf G

    2016-09-15

    Plants employ RNA silencing as an innate defense mechanism against viruses. As a counter-defense, plant viruses have evolved to express RNA silencing suppressor proteins (RSS), which target one or more steps of the silencing pathway. In this study, we show that the phosphoprotein (P) encoded by the negative-sense RNA virus alfalfa dwarf virus (ADV), a species of the genus Cytorhabdovirus, family Rhabdoviridae, is a suppressor of RNA silencing. ADV P has a relatively weak local RSS activity, and does not prevent siRNA accumulation. On the other hand, ADV P strongly suppresses systemic RNA silencing, but does not interfere with the short-distance spread of silencing, which is consistent with its lack of inhibition of siRNA accumulation. The mechanism of suppression appears to involve ADV P binding to RNA-induced silencing complex proteins AGO1 and AGO4 as shown in protein-protein interaction assays when ectopically expressed. In planta, we demonstrate that ADV P likely functions by inhibiting miRNA-guided AGO1 cleavage and prevents transitive amplification by repressing the production of secondary siRNAs. As recently described for lettuce necrotic yellows cytorhabdovirus P, but in contrast to other viral RSS known to disrupt AGO activity, ADV P sequence does not contain any recognizable GW/WG or F-box motifs, which suggests that cytorhabdovirus P proteins may use alternative motifs to bind to AGO proteins. Crown Copyright © 2016. Published by Elsevier B.V. All rights reserved.

  13. The bifunctional abiotic stress signalling regulator and endogenous RNA silencing suppressor FIERY1 is required for lateral root formation

    KAUST Repository

    Chen, Hao

    2010-09-28

    The Arabidopsis FIERY1 (FRY1) locus was originally identified as a negative regulator of stress-responsive gene expression and later shown to be required for suppression of RNA silencing. In this study we discovered that the FRY1 locus also regulates lateral root formation. Compared with the wild type, fry1 mutant seedlings generated significantly fewer lateral roots under normal growth conditions and also exhibited a dramatically reduced sensitivity to auxin in inducing lateral root initiation. Using transgenic plants that overexpress a yeast homolog of FRY1 that possesses only the 3\\', 5\\'-bisphosphate nucleotidase activity but not the inositol 1-phosphatase activity, we demonstrated that the lateral root phenotypes in fry1 result from loss of the nucleotidase activity. Furthermore, a T-DNA insertion mutant of another RNA silencing suppressor, XRN4 (but not XRN2 or XRN3), which is an exoribonuclease that is inhibited by the substrate of the FRY1 3\\', 5\\'-bisphosphate nucleotidase, exhibits similar lateral root defects. Although fry1 and xrn4 exhibited reduced sensitivity to ethylene, our experiments demonstrated that restoration of ethylene sensitivity in the fry1 mutant is not sufficient to rescue the lateral root phenotypes of fry1. Our results indicate that RNA silencing modulated by FRY1 and XRN4 plays an important role in shaping root architecture. © 2010 Blackwell Publishing Ltd.

  14. A viral suppressor of RNA silencing inhibits ARGONAUTE 1 function by precluding target RNA binding to pre-assembled RISC.

    Science.gov (United States)

    Kenesi, Erzsébet; Carbonell, Alberto; Lózsa, Rita; Vértessy, Beáta; Lakatos, Lóránt

    2017-07-27

    In most eukaryotes, RNA silencing is an adaptive immune system regulating key biological processes including antiviral defense. To evade this response, viruses of plants, worms and insects have evolved viral suppressors of RNA silencing proteins (VSRs). Various VSRs, such as P1 from Sweet potato mild mottle virus (SPMMV), inhibit the activity of RNA-induced silencing complexes (RISCs) including an ARGONAUTE (AGO) protein loaded with a small RNA. However, the specific mechanisms explaining this class of inhibition are unknown. Here, we show that SPMMV P1 interacts with AGO1 and AGO2 from Arabidopsis thaliana, but solely interferes with AGO1 function. Moreover, a mutational analysis of a newly identified zinc finger domain in P1 revealed that this domain could represent an effector domain as it is required for P1 suppressor activity but not for AGO1 binding. Finally, a comparative analysis of the target RNA binding capacity of AGO1 in the presence of wild-type or suppressor-defective P1 forms revealed that P1 blocks target RNA binding to AGO1. Our results describe the negative regulation of RISC, the small RNA containing molecular machine. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  15. The Enamovirus P0 protein is a silencing suppressor which inhibits local and systemic RNA silencing through AGO1 degradation

    Energy Technology Data Exchange (ETDEWEB)

    Fusaro, Adriana F. [University of Sydney, NSW 2006 (Australia); CSIRO Plant Industry, Canberra, P.O. Box 1600, ACT 2601 (Australia); Correa, Regis L. [CSIRO Plant Industry, Canberra, P.O. Box 1600, ACT 2601 (Australia); Depto. de Virologia, IMPPG, UFRJ, 21941-902 (Brazil); Nakasugi, Kenlee; Jackson, Craig [University of Sydney, NSW 2006 (Australia); Kawchuk, Lawrence [Research Centre, Agriculture and Agri-Food Canada, Lethbridge, AB T1J4B1 (Canada); Vaslin, Maite F.S. [Depto. de Virologia, IMPPG, UFRJ, 21941-902 (Brazil); Waterhouse, Peter M., E-mail: peter.waterhouse@sydney.edu.au [University of Sydney, NSW 2006 (Australia); CSIRO Plant Industry, Canberra, P.O. Box 1600, ACT 2601 (Australia)

    2012-05-10

    The P0 protein of poleroviruses and P1 protein of sobemoviruses suppress the plant's RNA silencing machinery. Here we identified a silencing suppressor protein (SSP), P0{sup PE}, in the Enamovirus Pea enation mosaic virus-1 (PEMV-1) and showed that it and the P0s of poleroviruses Potato leaf roll virus and Cereal yellow dwarf virus have strong local and systemic SSP activity, while the P1 of Sobemovirus Southern bean mosaic virus supresses systemic silencing. The nuclear localized P0{sup PE} has no discernable sequence conservation with known SSPs, but proved to be a strong suppressor of local silencing and a moderate suppressor of systemic silencing. Like the P0s from poleroviruses, P0{sup PE} destabilizes AGO1 and this action is mediated by an F-box-like domain. Therefore, despite the lack of any sequence similarity, the poleroviral and enamoviral SSPs have a conserved mode of action upon the RNA silencing machinery.

  16. Elicitation of hypersensitive responses in Nicotiana glutinosa by the suppressor of RNA silencing protein P0 from poleroviruses.

    Science.gov (United States)

    Wang, Ken-Der; Empleo, Roman; Nguyen, Tan Tri V; Moffett, Peter; Sacco, Melanie Ann

    2015-06-01

    Plant disease resistance (R) proteins that confer resistance to viruses recognize viral gene products with diverse functions, including viral suppressors of RNA silencing (VSRs). The P0 protein from poleroviruses is a VSR that targets the ARGONAUTE1 (AGO1) protein for degradation, thereby disrupting RNA silencing and antiviral defences. Here, we report resistance against poleroviruses in Nicotiana glutinosa directed against Turnip yellows virus (TuYV) and Potato leafroll virus (PLRV). The P0 proteins from TuYV (P0(T) (u) ), PLRV (P0(PL) ) and Cucurbit aphid-borne yellows virus (P0(CA) ) were found to elicit a hypersensitive response (HR) in N. glutinosa accession TW59, whereas other accessions recognized P0(PL) only. Genetic analysis showed that recognition of P0(T) (u) by a resistance gene designated RPO1 (Resistance to POleroviruses 1) is inherited as a dominant allele. Expression of P0 from a Potato virus X (PVX) expression vector transferred recognition to the recombinant virus on plants expressing RPO1, supporting P0 as the unique Polerovirus factor eliciting resistance. The induction of HR required a functional P0 protein, as P0(T) (u) mutants with substitutions in the F-box motif that abolished VSR activity were unable to elicit HR. We surmised that the broad P0 recognition seen in TW59 and the requirement for the F-box protein motif could indicate detection of P0-induced AGO1 degradation and disruption of RNA silencing; however, other viral silencing suppressors, including the PVX P25 that also causes AGO1 degradation, failed to elicit HR in N. glutinosa. Investigation of P0 elicitation of RPO1 could provide insight into P0 activities within the cell that trigger resistance. © 2014 BSPP AND JOHN WILEY & SONS LTD.

  17. The potyviral suppressor of RNA silencing confers enhanced resistance to multiple pathogens

    International Nuclear Information System (INIS)

    Pruss, Gail J.; Lawrence, Christopher B.; Bass, Troy; Li Qingshun Q.; Bowman, Lewis H.; Vance, Vicki

    2004-01-01

    Helper component-protease (HC-Pro) is a plant viral suppressor of RNA silencing, and transgenic tobacco expressing HC-Pro has increased susceptibility to a broad range of viral pathogens. Here we report that these plants also exhibit enhanced resistance to unrelated heterologous pathogens. Tobacco mosaic virus (TMV) infection of HC-Pro-expressing plants carrying the N resistance gene results in fewer and smaller lesions compared to controls without HC-Pro. The resistance to TMV is compromised but not eliminated by expression of nahG, which prevents accumulation of salicylic acid (SA), an important defense signaling molecule. HC-Pro-expressing plants are also more resistant to tomato black ring nepovirus (TBRV) and to the oomycete Peronospora tabacina. Enhanced TBRV resistance is SA-independent, whereas the response to P. tabacina is associated with early induction of markers characteristic of SA-dependent defense. Thus, a plant viral suppressor of RNA silencing enhances resistance to multiple pathogens via both SA-dependent and SA-independent mechanisms

  18. The potyviral suppressor of RNA silencing confers enhanced resistance to multiple pathogens.

    Science.gov (United States)

    Pruss, Gail J; Lawrence, Christopher B; Bass, Troy; Li, Qingshun Q; Bowman, Lewis H; Vance, Vicki

    2004-03-01

    Helper component-protease (HC-Pro) is a plant viral suppressor of RNA silencing, and transgenic tobacco expressing HC-Pro has increased susceptibility to a broad range of viral pathogens. Here we report that these plants also exhibit enhanced resistance to unrelated heterologous pathogens. Tobacco mosaic virus (TMV) infection of HC-Pro-expressing plants carrying the N resistance gene results in fewer and smaller lesions compared to controls without HC-Pro. The resistance to TMV is compromised but not eliminated by expression of nahG, which prevents accumulation of salicylic acid (SA), an important defense signaling molecule. HC-Pro-expressing plants are also more resistant to tomato black ring nepovirus (TBRV) and to the oomycete Peronospora tabacina. Enhanced TBRV resistance is SA-independent, whereas the response to P. tabacina is associated with early induction of markers characteristic of SA-dependent defense. Thus, a plant viral suppressor of RNA silencing enhances resistance to multiple pathogens via both SA-dependent and SA-independent mechanisms.

  19. BIOLOGICAL FUNCTION OF TOMBUSVIRUS-ENCODED SUPPRESSOR OF RNA SILENCING IN PLANTS

    Directory of Open Access Journals (Sweden)

    Omarov R.T.

    2012-08-01

    Full Text Available RNA interference (RNAi plays multiple biological roles in eukaryotic organisms to regulate gene expression. RNAi also operates as a conserved adaptive molecular immune mechanism against invading viruses. The antiviral RNAi pathway is initiated with the generation of virus-derived short-interfering RNAs (siRNAs that are used for subsequent sequence-specific recognition and degradation of the cognate viral RNA molecules. As an efficient counter-defensive strategy, most plant viruses evolved the ability to encode specific proteins capable of interfering with RNAi, and this process is commonly known as RNA silencing suppression. Virus-encoded suppressors of RNAi (VSRs operate at different steps in the RNAi pathway and display distinct biochemical properties that enable these proteins to efficiently interfere with the host-defense system. Tombusvirus-encoded P19 is an important pathogenicity factor, required for symptom development and elicitation of a hypersensitive response in a host-dependent manner. Protein plays a crucial role of TBSV P19 in protecting viral RNA during systemic infection on Nicotiana benthamiana. The X-ray crystallographic studies conducted by two independent groups revealed the existence of a P19-siRNA complex; a conformation whereby caliper tryptophan residues on two subunits of P19 dimers measure and bind 21-nt siRNA duplexes. These structural studies provided the first details on the possible molecular mechanism of any viral suppressor to block RNAi. The association between P19 and siRNAs was also shown to occur in infected plants These and related studies revealed that in general the ability of P19 to efficiently sequester siRNAs influences symptom severity, however this is not a strict correlation in all hosts.The current working model is that during TBSV infection of plants, P19 appropriates abundantly circulating Tombusvirus-derived siRNAs thereby rendering these unavailable to program RISC, to prevent degradation of

  20. Antiviral RNA silencing suppression activity of Tomato spotted wilt virus NSs protein.

    Science.gov (United States)

    Ocampo Ocampo, T; Gabriel Peralta, S M; Bacheller, N; Uiterwaal, S; Knapp, A; Hennen, A; Ochoa-Martinez, D L; Garcia-Ruiz, H

    2016-06-17

    In addition to regulating gene expression, RNA silencing is an essential antiviral defense system in plants. Triggered by double-stranded RNA, silencing results in degradation or translational repression of target transcripts. Viruses are inducers and targets of RNA silencing. To condition susceptibility, most plant viruses encode silencing suppressors that interfere with this process, such as the Tomato spotted wilt virus (TSWV) NSs protein. The mechanism by which NSs suppresses RNA silencing and its role in viral infection and movement remain to be determined. We cloned NSs from the Hawaii isolate of TSWV and using two independent assays show for the first time that this protein restored pathogenicity and supported the formation of local infection foci by suppressor-deficient Turnip mosaic virus and Turnip crinkle virus. Demonstrating the suppression of RNA silencing directed against heterologous viruses establishes the foundation to determine the means used by NSs to block this antiviral process.

  1. The Heterologous Expression of the p22 RNA Silencing Suppressor of the Crinivirus Tomato Chlorosis Virus from Tobacco Rattle Virus and Potato Virus X Enhances Disease Severity but Does Not Complement Suppressor-Defective Mutant Viruses.

    Science.gov (United States)

    Landeo-Ríos, Yazmín; Navas-Castillo, Jesús; Moriones, Enrique; Cañizares, M. Carmen

    2017-11-24

    To counteract host antiviral RNA silencing, plant viruses express suppressor proteins that function as pathogenicity enhancers. The genome of the Tomato chlorosis virus (ToCV) (genus Crinivirus , family Closteroviridae ) encodes an RNA silencing suppressor, the protein p22, that has been described as having one of the longest lasting local suppressor activities when assayed in Nicotiana benthamiana . Since suppression of RNA silencing and the ability to enhance disease severity are closely associated, we analyzed the effect of expressing p22 in heterologous viral contexts. Thus, we studied the effect of the expression of ToCV p22 from viral vectors Tobacco rattle virus (TRV) and Potato virus X (PVX), and from attenuated suppressor mutants in N. benthamiana plants. Our results show that although an exacerbation of disease symptoms leading to plant death was observed in the heterologous expression of ToCV p22 from both viruses, only in the case of TRV did increased viral accumulation occur. The heterologous expression of ToCV p22 could not complement suppressor-defective mutant viruses.

  2. Heterologous RNA-silencing suppressors from both plant- and animal-infecting viruses support plum pox virus infection.

    Science.gov (United States)

    Maliogka, Varvara I; Calvo, María; Carbonell, Alberto; García, Juan Antonio; Valli, Adrian

    2012-07-01

    HCPro, the RNA-silencing suppressor (RSS) of viruses belonging to the genus Potyvirus in the family Potyviridae, is a multifunctional protein presumably involved in all essential steps of the viral infection cycle. Recent studies have shown that plum pox potyvirus (PPV) HCPro can be replaced successfully by cucumber vein yellowing ipomovirus P1b, a sequence-unrelated RSS from a virus of the same family. In order to gain insight into the requirement of a particular RSS to establish a successful potyviral infection, we tested the ability of different heterologous RSSs from both plant- and animal-infecting viruses to substitute for HCPro. Making use of engineered PPV chimeras, we show that PPV HCPro can be replaced functionally by some, but not all, unrelated RSSs, including the NS1 protein of the mammal-infecting influenza A virus. Interestingly, the capacity of a particular RSS to replace HCPro does not correlate strictly with its RNA silencing-suppression strength. Altogether, our results suggest that not all suppression strategies are equally suitable for efficient escape of PPV from the RNA-silencing machinery. The approach followed here, based on using PPV chimeras in which an under-consideration RSS substitutes for HCPro, could further help to study the function of diverse RSSs in a 'highly sensitive' RNA-silencing context, such as that taking place in plant cells during the process of a viral infection.

  3. Phytophthora suppressor of RNA silencing 2 is a conserved RxLR effector that promotes infection in soybean and Arabidopsis thaliana.

    Science.gov (United States)

    Xiong, Qin; Ye, Wenwu; Choi, Duseok; Wong, James; Qiao, Yongli; Tao, Kai; Wang, Yuanchao; Ma, Wenbo

    2014-12-01

    The genus Phytophthora consists of notorious and emerging pathogens of economically important crops. Each Phytophthora genome encodes several hundreds of cytoplasmic effectors, which are believed to manipulate plant immune response inside the host cells. However, the majority of Phytophthora effectors remain functionally uncharacterized. We recently discovered two effectors from the soybean stem and root rot pathogen Phytophthora sojae with the activity to suppress RNA silencing in plants. These effectors are designated Phytophthora suppressor of RNA silencing (PSRs). Here, we report that the P. sojae PSR2 (PsPSR2) belongs to a conserved and widespread effector family in Phytophthora. A PsPSR2-like effector produced by P. infestans (PiPSR2) can also suppress RNA silencing in plants and promote Phytophthora infection, suggesting that the PSR2 family effectors have conserved functions in plant hosts. Using Agrobacterium rhizogenes-mediated hairy roots induction, we demonstrated that the expression of PsPSR2 rendered hypersusceptibility of soybean to P. sojae. Enhanced susceptibility was also observed in PsPSR2-expressing Arabidopsis thaliana plants during Phytophthora but not bacterial infection. These experiments provide strong evidence that PSR2 is a conserved Phytophthora effector family that performs important virulence functions specifically during Phytophthora infection of various plant hosts.

  4. Identification and characterization of two RNA silencing suppressors encoded by ophioviruses

    NARCIS (Netherlands)

    Robles Luna, Gabriel; Reyes, Carina A.; Peña, Eduardo J.; Ocolotobiche, Eliana; Baeza, Cecilia; Borniego, Maria Belén; Kormelink, Richard; García, María Laura

    2017-01-01

    Citrus psorosis virus and Mirafiori lettuce big-vein virus are two members of the genus Ophiovirus, family Ophioviridae. So far, how these viruses can interfere in the antiviral RNA silencing pathway is not known. In this study, using a local GFP silencing assay on Nicotiana benthamiana, the

  5. Diverging affinity of tospovirus RNA silencing suppressor proteins, NSs, for various RNA duplex molecules

    NARCIS (Netherlands)

    Schnettler, E.; Hemmes, J.C.; Huisman, R.; Goldbach, R.W.; Prins, M.W.; Kormelink, R.J.M.

    2010-01-01

    The tospovirus NSs protein was previously shown to suppress the antiviral RNA silencing mechanism in plants. Here the biochemical analysis of NSs proteins from different tospoviruses, using purified NSs or NSs containing cell extracts, is described. The results showed that all tospoviral NSs

  6. The Ebola virus VP35 protein is a suppressor of RNA silencing

    NARCIS (Netherlands)

    Haasnoot, J.; Vries, de W.; Geutjes, E.J.; Prins, M.W.; Haan, de P.; Berkhout, B.

    2007-01-01

    RNA silencing or interference (RNAi) is a gene regulation mechanism in eukaryotes that controls cell differentiation and developmental processes via expression of microRNAs. RNAi also serves as an innate antiviral defence response in plants, nematodes, and insects. This antiviral response is

  7. Diverging affinity of tospovirus RNA silencing suppressor proteins, NSs, for various RNA duplex molecules.

    Science.gov (United States)

    Schnettler, Esther; Hemmes, Hans; Huismann, Rik; Goldbach, Rob; Prins, Marcel; Kormelink, Richard

    2010-11-01

    The tospovirus NSs protein was previously shown to suppress the antiviral RNA silencing mechanism in plants. Here the biochemical analysis of NSs proteins from different tospoviruses, using purified NSs or NSs containing cell extracts, is described. The results showed that all tospoviral NSs proteins analyzed exhibited affinity to small double-stranded RNA molecules, i.e., small interfering RNAs (siRNAs) and micro-RNA (miRNA)/miRNA* duplexes. Interestingly, the NSs proteins from tomato spotted wilt virus (TSWV), impatiens necrotic spot virus (INSV), and groundnut ringspot virus (GRSV) also showed affinity to long double-stranded RNA (dsRNA), whereas tomato yellow ring virus (TYRV) NSs did not. The TSWV NSs protein was shown to be capable of inhibiting Dicer-mediated cleavage of long dsRNA in vitro. In addition, it suppressed the accumulation of green fluorescent protein (GFP)-specific siRNAs during coinfiltration with an inverted-repeat-GFP RNA construct in Nicotiana benthamiana. In vivo interference of TSWV NSs in the miRNA pathway was shown by suppression of an enhanced GFP (eGFP) miRNA sensor construct. The ability to stabilize miRNA/miRNA* by different tospovirus NSs proteins in vivo was demonstrated by increased accumulation and detection of both miRNA171c and miRNA171c* in tospovirus-infected N. benthamiana. All together, these data suggest that tospoviruses interfere in the RNA silencing pathway by sequestering siRNA and miRNA/miRNA* molecules before they are uploaded into their respective RNA-induced silencing complexes. The observed affinity to long dsRNA for only a subset of the tospoviruses studied is discussed in light of evolutional divergence and their ancestral relation to the animal-infecting members of the Bunyaviridae.

  8. Characterization of the RNA silencing suppression activity of the Ebola virus VP35 protein in plants and mammalian cells.

    Science.gov (United States)

    Zhu, Yali; Cherukuri, Nil Celebi; Jackel, Jamie N; Wu, Zetang; Crary, Monica; Buckley, Kenneth J; Bisaro, David M; Parris, Deborah S

    2012-03-01

    Ebola virus (EBOV) causes a lethal hemorrhagic fever for which there is no approved effective treatment or prevention strategy. EBOV VP35 is a virulence factor that blocks innate antiviral host responses, including the induction of and response to alpha/beta interferon. VP35 is also an RNA silencing suppressor (RSS). By inhibiting microRNA-directed silencing, mammalian virus RSSs have the capacity to alter the cellular environment to benefit replication. A reporter gene containing specific microRNA target sequences was used to demonstrate that prior expression of wild-type VP35 was able to block establishment of microRNA silencing in mammalian cells. In addition, wild-type VP35 C-terminal domain (CTD) protein fusions were shown to bind small interfering RNA (siRNA). Analysis of mutant proteins demonstrated that reporter activity in RSS assays did not correlate with their ability to antagonize double-stranded RNA (dsRNA)-activated protein kinase R (PKR) or bind siRNA. The results suggest that enhanced reporter activity in the presence of VP35 is a composite of nonspecific translational enhancement and silencing suppression. Moreover, most of the specific RSS activity in mammalian cells is RNA binding independent, consistent with VP35's proposed role in sequestering one or more silencing complex proteins. To examine RSS activity in a system without interferon, VP35 was tested in well-characterized plant silencing suppression assays. VP35 was shown to possess potent plant RSS activity, and the activities of mutant proteins correlated strongly, but not exclusively, with RNA binding ability. The results suggest the importance of VP35-protein interactions in blocking silencing in a system (mammalian) that cannot amplify dsRNA.

  9. Biochemical and single-molecule analyses of the RNA silencing suppressing activity of CrPV-1A.

    Science.gov (United States)

    Watanabe, Mariko; Iwakawa, Hiro-Oki; Tadakuma, Hisashi; Tomari, Yukihide

    2017-10-13

    Viruses often encode viral silencing suppressors (VSSs) to counteract the hosts' RNA silencing activity. The cricket paralysis virus 1A protein (CrPV-1A) is a unique VSS that binds to a specific Argonaute protein (Ago)-the core of the RNA-induced silencing complex (RISC)-in insects to suppress its target cleavage reaction. However, the precise molecular mechanism of CrPV-1A action remains unclear. Here we utilized biochemical and single-molecule imaging approaches to analyze the effect of CrPV-1A during target recognition and cleavage by Drosophila Ago2-RISC. Our results suggest that CrPV-1A obstructs the initial target searching by Ago2-RISC via base pairing in the seed region. The combination of biochemistry and single-molecule imaging may help to pave the way for mechanistic understanding of VSSs with diverse functions. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  10. Genetic variability and evolutionary implications of RNA silencing suppressor genes in RNA1 of sweet potato chlorotic stunt virus isolates infecting sweetpotato and related wild species.

    Directory of Open Access Journals (Sweden)

    Arthur K Tugume

    sweetpotato. A second virus encoding an RNase3-like RNA silencing suppressor was detected. Overall, results provided many novel and important insights into evolutionary biology of SPCSV.

  11. Mimic Phosphorylation of a βC1 Protein Encoded by TYLCCNB Impairs Its Functions as a Viral Suppressor of RNA Silencing and a Symptom Determinant.

    Science.gov (United States)

    Zhong, Xueting; Wang, Zhan Qi; Xiao, Ruyuan; Cao, Linge; Wang, Yaqin; Xie, Yan; Zhou, Xueping

    2017-08-15

    Phosphorylation of the βC1 protein encoded by the betasatellite of tomato yellow leaf curl China virus (TYLCCNB-βC1) by SNF1-related protein kinase 1 (SnRK1) plays a critical role in defense of host plants against geminivirus infection in Nicotiana benthamiana However, how phosphorylation of TYLCCNB-βC1 impacts its pathogenic functions during viral infection remains elusive. In this study, we identified two additional tyrosine residues in TYLCCNB-βC1 that are phosphorylated by SnRK1. The effects of TYLCCNB-βC1 phosphorylation on its functions as a viral suppressor of RNA silencing (VSR) and a symptom determinant were investigated via phosphorylation mimic mutants in N. benthamiana plants. Mutations that mimic phosphorylation of TYLCCNB-βC1 at tyrosine 5 and tyrosine 110 attenuated disease symptoms during viral infection. The phosphorylation mimics weakened the ability of TYLCCNB-βC1 to reverse transcriptional gene silencing and to suppress posttranscriptional gene silencing and abolished its interaction with N. benthamiana ASYMMETRIC LEAVES 1 in N. benthamiana leaves. The mimic phosphorylation of TYLCCNB-βC1 had no impact on its protein stability, subcellular localization, or self-association. Our data establish an inhibitory effect of phosphorylation of TYLCCNB-βC1 on its pathogenic functions as a VSR and a symptom determinant and provide a mechanistic explanation of how SnRK1 functions as a host defense factor. IMPORTANCE Tomato yellow leaf curl China virus (TYLCCNV), which causes a severe yellow leaf curl disease in China, is a monopartite geminivirus associated with the betasatellite (TYLCCNB). TYLCCNB encodes a single pathogenicity protein, βC1 (TYLCCNB-βC1), which functions as both a viral suppressor of RNA silencing (VSR) and a symptom determinant. Here, we show that mimicking phosphorylation of TYLCCNB-βC1 weakens its ability to reverse transcriptional gene silencing, to suppress posttranscriptional gene silencing, and to interact with N

  12. Phylogenetic Studies of the Three RNA Silencing Suppressor Genes of South American CTV Isolates Reveal the Circulation of a Novel Genetic Lineage

    Directory of Open Access Journals (Sweden)

    María José Benítez-Galeano

    2015-07-01

    Full Text Available Citrus Tristeza Virus (CTV is the most economically important virus of citrus worldwide. Genetic diversity and population structure of CTV isolates from all citrus growing areas from Uruguay were analyzed by RT-PCR and cloning of the three RNA silencing suppressor genes (p25, p20 and p23. Bayesian phylogenetic analysis revealed the circulation of three known genotypes (VT, T3, T36 in the country, and the presence of a new genetic lineage composed by isolates from around the world, mainly from South America. Nucleotide and amino acid identity values for this new genetic lineage were both higher than 97% for the three analyzed regions. Due to incongruent phylogenetic relationships, recombination analysis was performed using Genetic Algorithms for Recombination Detection (GARD and SimPlot software. Recombination events between previously described CTV isolates were detected. High intra-sample variation was found, confirming the co-existence of different genotypes into the same plant. This is the first report describing: (1 the genetic diversity of Uruguayan CTV isolates circulating in the country and (2 the circulation of a novel CTV genetic lineage, highly present in the South American region. This information may provide assistance to develop an effective cross-protection program.

  13. Viral RNA Silencing Suppression: The Enigma of Bunyavirus NSs Proteins

    Directory of Open Access Journals (Sweden)

    Marcio Hedil

    2016-07-01

    Full Text Available The Bunyaviridae is a family of arboviruses including both plant- and vertebrate-infecting representatives. The Tospovirus genus accommodates plant-infecting bunyaviruses, which not only replicate in their plant host, but also in their insect thrips vector during persistent propagative transmission. For this reason, they are generally assumed to encounter antiviral RNA silencing in plants and insects. Here we present an overview on how tospovirus nonstructural NSs protein counteracts antiviral RNA silencing in plants and what is known so far in insects. Like tospoviruses, members of the related vertebrate-infecting bunyaviruses classified in the genera Orthobunyavirus, Hantavirus and Phlebovirus also code for a NSs protein. However, for none of them RNA silencing suppressor activity has been unambiguously demonstrated in neither vertebrate host nor arthropod vector. The second part of this review will briefly describe the role of these NSs proteins in modulation of innate immune responses in mammals and elaborate on a hypothetical scenario to explain if and how NSs proteins from vertebrate-infecting bunyaviruses affect RNA silencing. If so, why this discovery has been hampered so far.

  14. Viral RNA Silencing Suppression: The Enigma of Bunyavirus NSs Proteins.

    Science.gov (United States)

    Hedil, Marcio; Kormelink, Richard

    2016-07-23

    The Bunyaviridae is a family of arboviruses including both plant- and vertebrate-infecting representatives. The Tospovirus genus accommodates plant-infecting bunyaviruses, which not only replicate in their plant host, but also in their insect thrips vector during persistent propagative transmission. For this reason, they are generally assumed to encounter antiviral RNA silencing in plants and insects. Here we present an overview on how tospovirus nonstructural NSs protein counteracts antiviral RNA silencing in plants and what is known so far in insects. Like tospoviruses, members of the related vertebrate-infecting bunyaviruses classified in the genera Orthobunyavirus, Hantavirus and Phlebovirus also code for a NSs protein. However, for none of them RNA silencing suppressor activity has been unambiguously demonstrated in neither vertebrate host nor arthropod vector. The second part of this review will briefly describe the role of these NSs proteins in modulation of innate immune responses in mammals and elaborate on a hypothetical scenario to explain if and how NSs proteins from vertebrate-infecting bunyaviruses affect RNA silencing. If so, why this discovery has been hampered so far.

  15. Antiviral RNA silencing viral counter defense in plants

    NARCIS (Netherlands)

    Bucher, E.C.

    2006-01-01

    The research described in this thesis centres around the mechanism of RNA silencing in relation to virus-host interaction, an area of increasing importance. It shows how this recently disclosed mechanism can be used to produce virus-resistant plants. Based on the activity of the RNA silencing

  16. The VP3 factor from viruses of Birnaviridae family suppresses RNA silencing by binding both long and small RNA duplexes.

    Directory of Open Access Journals (Sweden)

    Adrian Valli

    Full Text Available RNA silencing is directly involved in antiviral defense in a wide variety of eukaryotic organisms, including plants, fungi, invertebrates, and presumably vertebrate animals. The study of RNA silencing-mediated antiviral defences in vertebrates is hampered by the overlap with other antiviral mechanisms; thus, heterologous systems are often used to study the interplay between RNA silencing and vertebrate-infecting viruses. In this report we show that the VP3 protein of the avian birnavirus Infectious bursal disease virus (IBDV displays, in addition to its capacity to bind long double-stranded RNA, the ability to interact with double-stranded small RNA molecules. We also demonstrate that IBDV VP3 prevents the silencing mediated degradation of a reporter mRNA, and that this silencing suppression activity depends on its RNA binding ability. Furthermore, we find that the anti-silencing activity of IBDV VP3 is shared with the homologous proteins expressed by both insect- and fish-infecting birnaviruses. Finally, we show that IBDV VP3 can functionally replace the well-characterized HCPro silencing suppressor of Plum pox virus, a potyvirus that is unable to infect plants in the absence of an active silencing suppressor. Altogether, our results support the idea that VP3 protects the viral genome from host sentinels, including those of the RNA silencing machinery.

  17. The VP3 factor from viruses of Birnaviridae family suppresses RNA silencing by binding both long and small RNA duplexes.

    Science.gov (United States)

    Valli, Adrian; Busnadiego, Idoia; Maliogka, Varvara; Ferrero, Diego; Castón, José R; Rodríguez, José Francisco; García, Juan Antonio

    2012-01-01

    RNA silencing is directly involved in antiviral defense in a wide variety of eukaryotic organisms, including plants, fungi, invertebrates, and presumably vertebrate animals. The study of RNA silencing-mediated antiviral defences in vertebrates is hampered by the overlap with other antiviral mechanisms; thus, heterologous systems are often used to study the interplay between RNA silencing and vertebrate-infecting viruses. In this report we show that the VP3 protein of the avian birnavirus Infectious bursal disease virus (IBDV) displays, in addition to its capacity to bind long double-stranded RNA, the ability to interact with double-stranded small RNA molecules. We also demonstrate that IBDV VP3 prevents the silencing mediated degradation of a reporter mRNA, and that this silencing suppression activity depends on its RNA binding ability. Furthermore, we find that the anti-silencing activity of IBDV VP3 is shared with the homologous proteins expressed by both insect- and fish-infecting birnaviruses. Finally, we show that IBDV VP3 can functionally replace the well-characterized HCPro silencing suppressor of Plum pox virus, a potyvirus that is unable to infect plants in the absence of an active silencing suppressor. Altogether, our results support the idea that VP3 protects the viral genome from host sentinels, including those of the RNA silencing machinery.

  18. Induction of cell death by tospoviral protein NSs and the motif critical for cell death does not control RNA silencing suppression activity.

    Science.gov (United States)

    Singh, Ajeet; Permar, Vipin; Jain, R K; Goswami, Suneha; Kumar, Ranjeet Ranjan; Canto, Tomas; Palukaitis, Peter; Praveen, Shelly

    2017-08-01

    Groundnut bud necrosis virus induces necrotic symptoms in different hosts. Previous studies showed reactive oxygen species-mediated programmed cell death (PCD) resulted in necrotic symptoms. Transgenic expression of viral protein NSs mimics viral symptoms. Here, we showed a role for NSs in influencing oxidative burst in the cell, by analyzing H 2 O 2 accumulation, activities of antioxidant enzymes and expression levels of vacuolar processing enzymes, H 2 O 2 -responsive microRNA 319a.2 plus its possible target metacaspase-8. The role of NSs in PCD, was shown using two NSs mutants: one in the Trp/GH3 motif (a homologue of pro-apototic domain) (NSs S189R ) and the other in a non-Trp/GH3 motif (NSs L172R ). Tobacco rattle virus (TRV) expressing NSs S189R enhanced the PCD response, but not TRV-NSs L172R , while RNA silencing suppression activity was lost in TRV-NSs L172R , but not in TRV-NSs S189R . Therefore, we propose dual roles of NSs in RNA silencing suppression and induction of cell death, controlled by different motifs. Copyright © 2017 Elsevier Inc. All rights reserved.

  19. Soilborne wheat mosaic virus (SBWMV 19K protein belongs to a class of cysteine rich proteins that suppress RNA silencing

    Directory of Open Access Journals (Sweden)

    Howard Amanda

    2005-03-01

    Full Text Available Abstract Amino acid sequence analyses indicate that the Soilborne wheat mosaic virus (SBWMV 19K protein is a cysteine-rich protein (CRP and shares sequence homology with CRPs derived from furo-, hordei-, peclu- and tobraviruses. Since the hordei- and pecluvirus CRPs were shown to be pathogenesis factors and/or suppressors of RNA silencing, experiments were conducted to determine if the SBWMV 19K CRP has similar activities. The SBWMV 19K CRP was introduced into the Potato virus X (PVX viral vector and inoculated to tobacco plants. The SBWMV 19K CRP aggravated PVX-induced symptoms and restored green fluorescent protein (GFP expression to GFP silenced tissues. These observations indicate that the SBWMV 19K CRP is a pathogenicity determinant and a suppressor of RNA silencing.

  20. AGO/RISC-mediated antiviral RNA silencing in a plant in vitro system.

    Science.gov (United States)

    Schuck, Jana; Gursinsky, Torsten; Pantaleo, Vitantonio; Burgyán, Jozsef; Behrens, Sven-Erik

    2013-05-01

    AGO/RISC-mediated antiviral RNA silencing, an important component of the plant's immune response against RNA virus infections, was recapitulated in vitro. Cytoplasmic extracts of tobacco protoplasts were applied that supported Tombusvirus RNA replication, as well as the formation of RNA-induced silencing complexes (RISC) that could be functionally reconstituted with various plant ARGONAUTE (AGO) proteins. For example, when RISC containing AGO1, 2, 3 or 5 were programmed with exogenous siRNAs that specifically targeted the viral RNA, endonucleolytic cleavages occurred and viral replication was inhibited. Antiviral RNA silencing was disabled by the viral silencing suppressor p19 when this was present early during RISC formation. Notably, with replicating viral RNA, only (+)RNA molecules were accessible to RISC, whereas (-)RNA replication intermediates were not. The vulnerability of viral RNAs to RISC activity also depended on the RNA structure of the target sequence. This was most evident when we characterized viral siRNAs (vsiRNAs) that were particularly effective in silencing with AGO1- or AGO2/RISC. These vsiRNAs targeted similar sites, suggesting that accessible parts of the viral (+)RNA may be collectively attacked by different AGO/RISC. The in vitro system was, hence, established as a valuable tool to define and characterize individual molecular determinants of antiviral RNA silencing.

  1. Crop-associated virus reduces the rooting depth of non-crop perennial native grass more than non-crop-associated virus with known viral suppressor of RNA silencing (VSR).

    Science.gov (United States)

    Malmstrom, Carolyn M; Bigelow, Patrick; Trębicki, Piotr; Busch, Anna K; Friel, Colleen; Cole, Ellen; Abdel-Azim, Heba; Phillippo, Colin; Alexander, Helen M

    2017-09-15

    As agricultural acreage expanded and came to dominate landscapes across the world, viruses gained opportunities to move between crop and wild native plants. In the Midwestern USA, virus exchange currently occurs between widespread annual Poaceae crops and remnant native perennial prairie grasses now under consideration as bioenergy feedstocks. In this region, the common aphid species Rhopalosiphum padi L. (the bird cherry-oat aphid) transmits several virus species in the family Luteoviridae, including Barley yellow dwarf virus (BYDV-PAV, genus Luteovirus) and Cereal yellow dwarf virus (CYDV-RPV and -RPS, genus Polerovirus). The yellow dwarf virus (YDV) species in these two genera share genetic similarities in their 3'-ends, but diverge in the 5'-regions. Most notably, CYDVs encode a P0 viral suppressor of RNA silencing (VSR) absent in BYDV-PAV. Because BYDV-PAV has been reported more frequently in annual cereals and CYDVs in perennial non-crop grasses, we examine the hypothesis that the viruses' genetic differences reflect different affinities for crop and non-crop hosts. Specifically, we ask (i) whether CYDVs might persist within and affect a native non-crop grass more strongly than BYDV-PAV, on the grounds that the polerovirus VSR could better moderate the defenses of a well-defended perennial, and (ii) whether the opposite pattern of effects might occur in a less defended annual crop. Because previous work found that the VSR of CYDV-RPS possessed greater silencing suppressor efficiency than that of CYDV-RPV, we further explored (iii) whether a novel grass-associated CYDV-RPS isolate would influence a native non-crop grass more strongly than a comparable CYDV-RPV isolate. In growth chamber studies, we found support for this hypothesis: only grass-associated CYDV-RPS stunted the shoots and crowns of Panicum virgatum L. (switchgrass), a perennial native North American prairie grass, whereas crop-associated BYDV-PAV (and coinfection with BYDV-PAV and CYDV-RPS) most

  2. Tospovirus : induction and suppression of RNA silencing

    NARCIS (Netherlands)

    Hedil, Marcio

    2016-01-01

    While infecting their hosts, viruses must deal with host immunity. In plants the antiviral RNA silencing pathway is an important part of plant innate immunity. Tospoviruses are segmented negative-stranded RNA viruses of plants. To counteract the antiviral RNA silencing response in plants,

  3. The P0 protein encoded by cotton leafroll dwarf virus (CLRDV) inhibits local but not systemic RNA silencing.

    Science.gov (United States)

    Delfosse, Verónica C; Agrofoglio, Yamila C; Casse, María F; Kresic, Iván Bonacic; Hopp, H Esteban; Ziegler-Graff, Véronique; Distéfano, Ana J

    2014-02-13

    Plants employ RNA silencing as a natural defense mechanism against viruses. As a counter-defense, viruses encode silencing suppressor proteins (SSPs) that suppress RNA silencing. Most, but not all, the P0 proteins encoded by poleroviruses have been identified as SSP. In this study, we demonstrated that cotton leafroll dwarf virus (CLRDV, genus Polerovirus) P0 protein suppressed local silencing that was induced by sense or inverted repeat transgenes in Agrobacterium co-infiltration assay in Nicotiana benthamiana plants. A CLRDV full-length infectious cDNA clone that is able to infect N. benthamiana through Agrobacterium-mediated inoculation also inhibited local silencing in co-infiltration assays, suggesting that the P0 protein exhibits similar RNA silencing suppression activity when expressed from the full-length viral genome. On the other hand, the P0 protein did not efficiently inhibit the spread of systemic silencing signals. Moreover, Northern blotting indicated that the P0 protein inhibits the generation of secondary but not primary small interfering RNAs. The study of CLRDV P0 suppression activity may contribute to understanding the molecular mechanisms involved in the induction of cotton blue disease by CLRDV infection. Copyright © 2013 Elsevier B.V. All rights reserved.

  4. Visualization of plant viral suppressor silencing activity in intact leaf lamina by quantitative fluorescent imaging

    Directory of Open Access Journals (Sweden)

    Francis Kevin P

    2011-08-01

    Full Text Available Abstract Background Transient expression of proteins in plants has become a favoured method over the production of stably transformed plants because, in addition to enabling high protein yields, it is both fast and easy to apply. An enhancement of transient protein expression can be achieved by plant virus-encoded RNA silencing suppressor proteins. Since viral suppressor proteins differ in their efficiency to enhance transient protein expression in plants, we developed a whole-leaf green fluorescent protein (GFP-based imaging assay to quantitatively assess suppressor protein activity. Results In a transient GFP-expression assay using wild-type and GFP-transgenic N. benthamiana, addition of the plant viral suppressors Beet mild yellowing virus (BMYV-IPP P0 or Plum pox virus (PPV HC-Pro was shown to increase fluorescent protein expression 3-4-fold, 7 days post inoculation (dpi when compared to control plants. In contrast, in agroinfiltrated patches without suppressor activity, near complete silencing of the GFP transgene was observed in the transgenic N. benthamiana at 21 dpi. Both co-infiltrated suppressors significantly enhanced GFP expression over time, with HC-Pro co-infiltrations leading to higher short term GFP fluorescence (at 7 dpi and P0 giving higher long term GFP fluorescence (at 21 dpi. Additionally, in contrast to HC-Pro co-infiltrations, an area of complete GFP silencing was observed at the edge of P0 co-infiltrated areas. Conclusions Fluorescence imaging of whole intact leaves proved to be an easy and effective method for spatially and quantitatively observing viral suppressor efficiency in plants. This suppressor assay demonstrates that plant viral suppressors greatly enhanced transient GFP expression, with P0 showing a more prolonged suppressor activity over time than HC-Pro. Both suppressors could prove to be ideal candidates for enhancing target protein expression in plants.

  5. Amino acid sequence motifs essential for P0-mediated suppression of RNA silencing in an isolate of potato leafroll virus from Inner Mongolia.

    Science.gov (United States)

    Zhuo, Tao; Li, Yuan-Yuan; Xiang, Hai-Ying; Wu, Zhan-Yu; Wang, Xian-Bin; Wang, Ying; Zhang, Yong-Liang; Li, Da-Wei; Yu, Jia-Lin; Han, Cheng-Gui

    2014-06-01

    Polerovirus P0 suppressors of host gene silencing contain a consensus F-box-like motif with Leu/Pro (L/P) requirements for suppressor activity. The Inner Mongolian Potato leafroll virus (PLRV) P0 protein (P0(PL-IM)) has an unusual F-box-like motif that contains a Trp/Gly (W/G) sequence and an additional GW/WG-like motif (G139/W140/G141) that is lacking in other P0 proteins. We used Agrobacterium infiltration-mediated RNA silencing assays to establish that P0(PL-IM) has a strong suppressor activity. Mutagenesis experiments demonstrated that the P0(PL-IM) F-box-like motif encompasses amino acids 76-LPRHLHYECLEWGLLCG THP-95, and that the suppressor activity is abolished by L76A, W87A, or G88A substitution. The suppressor activity is also weakened substantially by mutations within the G139/W140/G141 region and is eliminated by a mutation (F220R) in a C-terminal conserved sequence of P0(PL-IM). As has been observed with other P0 proteins, P0(PL-IM) suppression is correlated with reduced accumulation of the host AGO1-silencing complex protein. However, P0(PL-IM) fails to bind SKP1, which functions in a proteasome pathway that may be involved in AGO1 degradation. These results suggest that P0(PL-IM) may suppress RNA silencing by using an alternative pathway to target AGO1 for degradation. Our results help improve our understanding of the molecular mechanisms involved in PLRV infection.

  6. Conifers have a unique small RNA silencing signature

    OpenAIRE

    Dolgosheina, Elena V.; Morin, Ryan D.; Aksay, Gozde; Sahinalp, S. Cenk; Magrini, Vincent; Mardis, Elaine R.; Mattsson, Jim; Unrau, Peter J.

    2008-01-01

    Plants produce small RNAs to negatively regulate genes, viral nucleic acids, and repetitive elements at either the transcriptional or post-transcriptional level in a process that is referred to as RNA silencing. While RNA silencing has been extensively studied across the different phyla of the animal kingdom (e.g., mouse, fly, worm), similar studies in the plant kingdom have focused primarily on angiosperms, thus limiting evolutionary studies of RNA silencing in plants. Here we report on an u...

  7. Strategies underlying RNA silencing suppression by negative strand RNA viruses

    NARCIS (Netherlands)

    Hemmes, J.C.

    2007-01-01

    The research described in this thesis focused on the strategies of negative strand RNA viruses to counteract antiviral RNA silencing. In plants and insects, RNA silencing has been shown to act as a sequence specific antiviral defence mechanism that is characterised by the processing of double

  8. Interplays between soil-borne plant viruses and RNA silencing-mediated antiviral defense in roots

    Directory of Open Access Journals (Sweden)

    Ida Bagus Andika

    2016-09-01

    Full Text Available Although the majority of plant viruses are transmitted by arthropod vectors and invade the host plants through the aerial parts, there is a considerable number of plant viruses that infect roots via soil-inhabiting vectors such as plasmodiophorids, chytrids, and nematodes. These soil-borne viruses belong to diverse families, and many of them cause serious diseases in major crop plants. Thus, roots are important organs for the life cycle of many viruses. Compared to shoots, roots have a distinct metabolism and particular physiological characteristics due to the differences in development, cell composition, gene expression patterns, and surrounding environmental conditions. RNA silencing is an important innate defense mechanism to combat virus infection in plants, but the specific information on the activities and molecular mechanism of RNA silencing-mediated viral defense in root tissue is still limited. In this review, we summarize and discuss the current knowledge regarding RNA silencing aspects of the interactions between soil-borne viruses and host plants. Overall, research evidence suggests that soil-borne viruses have evolved to adapt to the distinct mechanism of antiviral RNA silencing in roots.

  9. Analysis of Tomato spotted wilt virus NSs protein indicates the importance of the N-terminal for avirulence and RNA silencing suppression

    NARCIS (Netherlands)

    Ronde, de D.; Pasquier, A.; Ying, S.; Butterbach, P.B.E.; Lohuis, D.; Kormelink, R.J.M.

    2014-01-01

    Recently, Tomato spotted wilt virus (TSWV) nonstructural protein NSs has been identified unambiguously as an avirulence (Avr) determinant for Tomato spotted wilt (Tsw)-based resistance. The observation that NSs from two natural resistance-breaking isolates had lost RNA silencing suppressor (RSS)

  10. Anti-viral RNA silencing: do we look like plants ?

    Directory of Open Access Journals (Sweden)

    Lecellier Charles-Henri

    2006-01-01

    Full Text Available Abstract The anti-viral function of RNA silencing was first discovered in plants as a natural manifestation of the artificial 'co-suppression', which refers to the extinction of endogenous gene induced by homologous transgene. Because silencing components are conserved among most, if not all, eukaryotes, the question rapidly arose as to determine whether this process fulfils anti-viral functions in animals, such as insects and mammals. It appears that, whereas the anti-viral process seems to be similarly conserved from plants to insects, even in worms, RNA silencing does influence the replication of mammalian viruses but in a particular mode: micro(miRNAs, endogenous small RNAs naturally implicated in translational control, rather than virus-derived small interfering (siRNAs like in other organisms, are involved. In fact, these recent studies even suggest that RNA silencing may be beneficial for viral replication. Accordingly, several large DNA mammalian viruses have been shown to encode their own miRNAs. Here, we summarize the seminal studies that have implicated RNA silencing in viral infection and compare the different eukaryotic responses.

  11. Conifers have a unique small RNA silencing signature.

    Science.gov (United States)

    Dolgosheina, Elena V; Morin, Ryan D; Aksay, Gozde; Sahinalp, S Cenk; Magrini, Vincent; Mardis, Elaine R; Mattsson, Jim; Unrau, Peter J

    2008-08-01

    Plants produce small RNAs to negatively regulate genes, viral nucleic acids, and repetitive elements at either the transcriptional or post-transcriptional level in a process that is referred to as RNA silencing. While RNA silencing has been extensively studied across the different phyla of the animal kingdom (e.g., mouse, fly, worm), similar studies in the plant kingdom have focused primarily on angiosperms, thus limiting evolutionary studies of RNA silencing in plants. Here we report on an unexpected phylogenetic difference in the size distribution of small RNAs among the vascular plants. By extracting total RNA from freshly growing shoot tissue, we conducted a survey of small RNAs in 24 vascular plant species. We find that conifers, which radiated from the other seed-bearing plants approximately 260 million years ago, fail to produce significant amounts of 24-nucleotide (nt) RNAs that are known to guide DNA methylation and heterochromatin formation in angiosperms. Instead, they synthesize a diverse population of small RNAs that are exactly 21-nt long. This finding was confirmed by high-throughput sequencing of the small RNA sequences from a conifer, Pinus contorta. A conifer EST search revealed the presence of a novel Dicer-like (DCL) family, which may be responsible for the observed change in small RNA expression. No evidence for DCL3, an enzyme that matures 24-nt RNAs in angiosperms, was found. We hypothesize that the diverse class of 21-nt RNAs found in conifers may help to maintain organization of their unusually large genomes.

  12. Salicylic acid-mediated and RNA-silencing defense mechanisms cooperate in the restriction of systemic spread of plum pox virus in tobacco.

    Science.gov (United States)

    Alamillo, Josefa M; Saénz, Pilar; García, Juan Antonio

    2006-10-01

    Plum pox virus (PPV) is able to replicate in inoculated leaves of Nicotiana tabacum, but is defective in systemic movement in this host. However, PPV produces a systemic infection in transgenic tobacco expressing the silencing suppressor P1/HC-Pro from tobacco etch virus (TEV). In this work we show that PPV is able to move to upper non-inoculated leaves of tobacco plants expressing bacterial salicylate hydroxylase (NahG) that degrades salicylic acid (SA). Replication and accumulation of PPV is higher in the locally infected leaves of plants deficient in SA or expressing TEV P1/HC-Pro silencing suppressor. Accumulation of viral derived small RNAs was reduced in the NahG transgenic plants, suggesting that SA might act as an enhancer of the RNA-silencing antiviral defense in tobacco. Besides, expression of SA-mediated defense transcripts, such as those of pathogenesis-related (PR) proteins PR-1 and PR-2 or alternative oxidase-1, as well as that of the putative RNA-dependent RNA polymerase NtRDR1, is induced in response to PPV infection, and the expression patterns of these defense transcripts are altered in the TEV P1/HC-Pro transgenic plants. Long-distance movement of PPV is highly enhanced in NahG x P1/HC-Pro double-transgenic plants and systemic symptoms in these plants reveal that the expression of an RNA-silencing suppressor and the lack of SA produce additive but distinct effects. Our results suggest that SA might act as an enhancer of the RNA-silencing antiviral defense in tobacco, and that silencing suppressors, such as P1/HC-Pro, also alter the SA-mediated defense. Both an RNA-silencing and an SA-mediated defense mechanism could act together to limit PPV infection.

  13. Two Novel Motifs of Watermelon Silver Mottle Virus NSs Protein Are Responsible for RNA Silencing Suppression and Pathogenicity.

    Science.gov (United States)

    Huang, Chung-Hao; Hsiao, Weng-Rong; Huang, Ching-Wen; Chen, Kuan-Chun; Lin, Shih-Shun; Chen, Tsung-Chi; Raja, Joseph A J; Wu, Hui-Wen; Yeh, Shyi-Dong

    2015-01-01

    The NSs protein of Watermelon silver mottle virus (WSMoV) is the RNA silencing suppressor and pathogenicity determinant. In this study, serial deletion and point-mutation mutagenesis of conserved regions (CR) of NSs protein were performed, and the silencing suppression function was analyzed through agroinfiltration in Nicotiana benthamiana plants. We found two amino acid (aa) residues, H113 and Y398, are novel functional residues for RNA silencing suppression. Our further analyses demonstrated that H113 at the common epitope (CE) ((109)KFTMHNQ(117)), which is highly conserved in Asia type tospoviruses, and the benzene ring of Y398 at the C-terminal β-sheet motif ((397)IYFL(400)) affect NSs mRNA stability and protein stability, respectively, and are thus critical for NSs RNA silencing suppression. Additionally, protein expression of other six deleted (ΔCR1-ΔCR6) and five point-mutated (Y15A, Y27A, G180A, R181A and R212A) mutants were hampered and their silencing suppression ability was abolished. The accumulation of the mutant mRNAs and proteins, except Y398A, could be rescued or enhanced by co-infiltration with potyviral suppressor HC-Pro. When assayed with the attenuated Zucchini yellow mosaic virus vector in squash plants, the recombinants carrying individual seven point-mutated NSs proteins displayed symptoms much milder than the recombinant carrying the wild type NSs protein, suggesting that these aa residues also affect viral pathogenicity by suppressing the host silencing mechanism.

  14. The RNA silencing pathway: the bits and pieces that matter.

    Directory of Open Access Journals (Sweden)

    2005-07-01

    Full Text Available Cellular pathways are generally proposed on the basis of available experimental knowledge. The proposed pathways, however, may be inadequate to describe the phenomena they are supposed to explain. For instance, by means of concise mathematical models we are able to reveal shortcomings in the current description of the pathway of RNA silencing. The silencing pathway operates by cleaving siRNAs from dsRNA. siRNAs can associate with RISC, leading to the degradation of the target mRNA. We propose and analyze a few small extensions to the pathway: a siRNA degrading RNase, primed amplification of aberrant RNA pieces, and cooperation between aberrant RNA to trigger amplification. These extensions allow for a consistent explanation for various types of silencing phenomena, such as virus induced silencing, transgene and transposon induced silencing, and avoidance of self-reactivity, as well as for differences found between species groups.

  15. RNA silencing is required for Arabidopsis defence against Verticillium wilt disease

    NARCIS (Netherlands)

    Ellendorff, U.; Fradin, E.F.; Jonge, de R.; Thomma, B.P.H.J.

    2009-01-01

    RNA silencing is a conserved mechanism in eukaryotes that plays an important role in various biological processes including regulation of gene expression. RNA silencing also plays a role in genome stability and protects plants against invading nucleic acids such as transgenes and viruses. Recently,

  16. RNAi suppressors encoded by pathogenic human viruses

    NARCIS (Netherlands)

    de Vries, Walter; Berkhout, Ben

    2008-01-01

    RNA silencing or RNAi interference (RNAi) serves as an innate antiviral mechanism in plants, fungi and animals. Human viruses, like plant viruses, encode suppressor proteins or RNAs that block or modulate the RNAi pathway. This review summarizes the mechanisms by which pathogenic human viruses

  17. RNA Silencing in Plants: Mechanisms, Technologies and Applications in Horticultural Crops.

    Science.gov (United States)

    Guo, Qigao; Liu, Qing; Smith, Neil A; Liang, Guolu; Wang, Ming-Bo

    2016-12-01

    Understanding the fundamental nature of a molecular process or a biological pathway is often a catalyst for the development of new technologies in biology. Indeed, studies from late 1990s to early 2000s have uncovered multiple overlapping but functionally distinct RNA silencing pathways in plants, including the posttranscriptional microRNA and small interfering RNA pathways and the transcriptional RNA-directed DNA methylation pathway. These findings have in turn been exploited for developing artificial RNA silencing technologies such as hairpin RNA, artificial microRNA, intrinsic direct repeat, 3' UTR inverted repeat, artificial trans-acting siRNA, and virus-induced gene silencing technologies. Some of these RNA silencing technologies, such as the hairpin RNA technology, have already been widely used for genetic improvement of crop plants in agriculture. For horticultural plants, RNA silencing technologies have been used to increase disease and pest resistance, alter plant architecture and flowering time, improve commercial traits of fruits and flowers, enhance nutritional values, remove toxic compounds and allergens, and develop high-value industrial products. In this article we aim to provide an overview of the RNA silencing pathways in plants, summarize the existing RNA silencing technologies, and review the current progress in applying these technologies for the improvement of agricultural crops particularly horticultural crops.

  18. Analysis of Tomato spotted wilt virus NSs protein indicates the importance of the N-terminal domain for avirulence and RNA silencing suppression.

    Science.gov (United States)

    de Ronde, Dryas; Pasquier, Adrien; Ying, Su; Butterbach, Patrick; Lohuis, Dick; Kormelink, Richard

    2014-02-01

    Recently, Tomato spotted wilt virus (TSWV) nonstructural protein NSs has been identified unambiguously as an avirulence (Avr) determinant for Tomato spotted wilt (Tsw)-based resistance. The observation that NSs from two natural resistance-breaking isolates had lost RNA silencing suppressor (RSS) activity and Avr suggested a link between the two functions. To test this, a large set of NSs mutants was generated by alanine substitutions in NSs from resistance-inducing wild-type strains (NSs(RI) ), amino acid reversions in NSs from resistance-breaking strains (NSs(RB)), domain deletions and swapping. Testing these mutants for their ability to suppress green fluorescent protein (GFP) silencing and to trigger a Tsw-mediated hypersensitive response (HR) revealed that the two functions can be separated. Changes in the N-terminal domain were found to be detrimental for both activities and indicated the importance of this domain, additionally supported by domain swapping between NSs(RI) and NSs(RB). Swapping domains between the closely related Tospovirus Groundnut ringspot virus (GRSV) NSs and TSWV NSs(RI) showed that Avr functionality could not simply be transferred between species. Although deletion of the C-terminal domain rendered NSs completely dysfunctional, only a few single-amino-acid mutations in the C-terminus affected both functions. Mutation of a GW/WG motif (position 17/18) rendered NSs completely dysfunctional for RSS and Avr activity, and indicated a putative interaction between NSs and Argonaute 1 (AGO1), and its importance in TSWV virulence and viral counter defence against RNA interference. © 2013 BSPP AND JOHN WILEY & SONS LTD.

  19. The dynamics and efficacy of antiviral RNA silencing: A model study

    Directory of Open Access Journals (Sweden)

    Hogeweg Paulien

    2008-03-01

    Full Text Available Abstract Background Mathematical modeling is important to provide insight in the complicated pathway of RNA silencing. RNA silencing is an RNA based mechanism that is widely used by eukaryotes to fight viruses, and to control gene expression. Results We here present the first mathematical model that combines viral growth with RNA silencing. The model involves a plus-strand RNA virus that replicates through a double-strand RNA intermediate. The model of the RNA silencing pathway consists of cleavage of viral RNA into siRNA by Dicer, target cleavage of viral RNA via the RISC complex, and a secondary response. We found that, depending on the strength of the silencing response, different viral growth patterns can occur. Silencing can decrease viral growth, cause oscillations, or clear the virus completely. Our model can explain various observed phenomena, even when they seem contradictory at first: the diverse responses to the removal of RNA dependent RNA polymerase; different viral growth curves; and the great diversity in observed siRNA ratios. Conclusion The model presented here is an important step in the understanding of the natural functioning of RNA silencing in viral infections.

  20. Stability of RNA silencing-based traits after virus infection

    DEFF Research Database (Denmark)

    Jørgensen, Bodil; Albrechtsen, Merete

    2007-01-01

    with constructs based on virus coat protein (CP) genes or other viral genes has been successfully used to engineer PTGS-mediated virus resistance into a large number of crop plants and some transgenic lines have been commercially exploited. However the discovery that plant viruses encode suppressors of gene...... silencing has raised concerns that virus infection of crop plants might reverse the new silencing-based traits. Most studies of virus suppression of silencing have used model systems based on silencing of reporter genes. A few studies have analysed the effects of virus infections on plants with genetically...... engineered virus resistance based on either a simple sense or an inverted repeat construct. We decided to use genetically engineered virus resistance in potato as a model system for further studies of the effect of virus infection on genetically engineered traits. We present for the first time a comparison...

  1. Recovery of Nicotiana benthamiana plants from a necrotic response induced by a nepovirus is associated with RNA silencing but not with reduced virus titer.

    Science.gov (United States)

    Jovel, Juan; Walker, Melanie; Sanfaçon, Hélène

    2007-11-01

    Recovery of plants from virus-induced symptoms is often described as a consequence of RNA silencing, an antiviral defense mechanism. For example, recovery of Nicotiana clevelandii from a nepovirus (tomato black ring virus) is associated with a decreased viral RNA concentration and sequence-specific resistance to further virus infection. In this study, we have characterized the interaction of another nepovirus, tomato ringspot virus (ToRSV), with host defense responses during symptom induction and subsequent recovery. Early in infection, ToRSV induced a necrotic phenotype in Nicotiana benthamiana that showed characteristics typical of a hypersensitive response. RNA silencing was also activated during ToRSV infection, as evidenced by the presence of ToRSV-derived small interfering RNAs (siRNAs) that could direct degradation of ToRSV sequences introduced into sensor constructs. Surprisingly, disappearance of symptoms was not accompanied by a commensurate reduction in viral RNA levels. The stability of ToRSV RNA after recovery was also observed in N. clevelandii and Cucumis sativus and in N. benthamiana plants carrying a functional RNA-dependent RNA polymerase 1 ortholog from Medicago truncatula. In experiments with a reporter transgene (green fluorescent protein), ToRSV did not suppress the initiation or maintenance of transgene silencing, although the movement of the silencing signal was partially hindered. Our results demonstrate that although RNA silencing is active during recovery, reduction of virus titer is not required for the initiation of this phenotype. This scenario adds an unforeseen layer of complexity to the interaction of nepoviruses with the host RNA silencing machinery. The possibility that viral proteins, viral RNAs, and/or virus-derived siRNAs inactivate host defense responses is discussed.

  2. High-throughput sequencing of RNA silencing-associated small RNAs in olive (Olea europaea L..

    Directory of Open Access Journals (Sweden)

    Livia Donaire

    Full Text Available Small RNAs (sRNAs of 20 to 25 nucleotides (nt in length maintain genome integrity and control gene expression in a multitude of developmental and physiological processes. Despite RNA silencing has been primarily studied in model plants, the advent of high-throughput sequencing technologies has enabled profiling of the sRNA component of more than 40 plant species. Here, we used deep sequencing and molecular methods to report the first inventory of sRNAs in olive (Olea europaea L.. sRNA libraries prepared from juvenile and adult shoots revealed that the 24-nt class dominates the sRNA transcriptome and atypically accumulates to levels never seen in other plant species, suggesting an active role of heterochromatin silencing in the maintenance and integrity of its large genome. A total of 18 known miRNA families were identified in the libraries. Also, 5 other sRNAs derived from potential hairpin-like precursors remain as plausible miRNA candidates. RNA blots confirmed miRNA expression and suggested tissue- and/or developmental-specific expression patterns. Target mRNAs of conserved miRNAs were computationally predicted among the olive cDNA collection and experimentally validated through endonucleolytic cleavage assays. Finally, we use expression data to uncover genetic components of the miR156, miR172 and miR390/TAS3-derived trans-acting small interfering RNA (tasiRNA regulatory nodes, suggesting that these interactive networks controlling developmental transitions are fully operational in olive.

  3. An RNA-seq transcriptome analysis of histone modifiers and RNA silencing genes in soybean during floral initiation process.

    Directory of Open Access Journals (Sweden)

    Lim Chee Liew

    Full Text Available Epigenetics has been recognised to play vital roles in many plant developmental processes, including floral initiation through the epigenetic regulation of gene expression. The histone modifying proteins that mediate these modifications involve the SET domain-containing histone methyltransferases, JmjC domain-containing demethylase, acetylases and deacetylases. In addition, RNA interference (RNAi-associated genes are also involved in epigenetic regulation via RNA-directed DNA methylation and post-transcriptional gene silencing. Soybean, a major crop legume, requires a short day to induce flowering. How histone modifications regulate the plant response to external cues that initiate flowering is still largely unknown. Here, we used RNA-seq to address the dynamics of transcripts that are potentially involved in the epigenetic programming and RNAi mediated gene silencing during the floral initiation of soybean. Soybean is a paleopolyploid that has been subjected to at least two rounds of whole genome duplication events. We report that the expanded genomic repertoire of histone modifiers and RNA silencing genes in soybean includes 14 histone acetyltransferases, 24 histone deacetylases, 47 histone methyltransferases, 15 protein arginine methyltransferases, 24 JmjC domain-containing demethylases and 47 RNAi-associated genes. To investigate the role of these histone modifiers and RNA silencing genes during floral initiation, we compared the transcriptional dynamics of the leaf and shoot apical meristem at different time points after a short-day treatment. Our data reveal that the extensive activation of genes that are usually involved in the epigenetic programming and RNAi gene silencing in the soybean shoot apical meristem are reprogrammed for floral development following an exposure to inductive conditions.

  4. RNA Silencing in Plants: Mechanisms, Technologies and Applications in Horticultural Crops

    OpenAIRE

    Guo, Qigao; Liu, Qing; Smith, Neil A.; Liang, Guolu; Wang, Ming-Bo

    2016-01-01

    Understanding the fundamental nature of a molecular process or a biological pathway is often a catalyst for the development of new technologies in biology. Indeed, studies from late 1990s to early 2000s have uncovered multiple overlapping but functionally distinct RNA silencing pathways in plants, including the posttranscriptional microRNA and small interfering RNA pathways and the transcriptional RNA-directed DNA methylation pathway. These findings have in turn been exploited for developing ...

  5. Telomeric trans-silencing: an epigenetic repression combining RNA silencing and heterochromatin formation.

    Directory of Open Access Journals (Sweden)

    Thibaut Josse

    2007-09-01

    Full Text Available The study of P-element repression in Drosophila melanogaster led to the discovery of the telomeric Trans-Silencing Effect (TSE, a repression mechanism by which a transposon or a transgene inserted in subtelomeric heterochromatin (Telomeric Associated Sequence or TAS has the capacity to repress in trans in the female germline, a homologous transposon, or transgene located in euchromatin. TSE shows variegation among egg chambers in ovaries when silencing is incomplete. Here, we report that TSE displays an epigenetic transmission through meiosis, which involves an extrachromosomal maternally transmitted factor. We show that this silencing is highly sensitive to mutations affecting both heterochromatin formation (Su(var205 encoding Heterochromatin Protein 1 and Su(var3-7 and the repeat-associated small interfering RNA (or rasiRNA silencing pathway (aubergine, homeless, armitage, and piwi. In contrast, TSE is not sensitive to mutations affecting r2d2, which is involved in the small interfering RNA (or siRNA silencing pathway, nor is it sensitive to a mutation in loquacious, which is involved in the micro RNA (or miRNA silencing pathway. These results, taken together with the recent discovery of TAS homologous small RNAs associated to PIWI proteins, support the proposition that TSE involves a repeat-associated small interfering RNA pathway linked to heterochromatin formation, which was co-opted by the P element to establish repression of its own transposition after its recent invasion of the D. melanogaster genome. Therefore, the study of TSE provides insight into the genetic properties of a germline-specific small RNA silencing pathway.

  6. Mild and severe cereal yellow dwarf viruses differ in silencing suppressor efficiency of the P0 protein.

    Science.gov (United States)

    Almasi, Reza; Miller, W Allen; Ziegler-Graff, Véronique

    2015-10-02

    Viral pathogenicity has often been correlated to the expression of the viral encoded-RNA silencing suppressor protein (SSP). The silencing suppressor activity of the P0 protein encoded by cereal yellow dwarf virus-RPV (CYDV-RPV) and -RPS (CYDV-RPS), two poleroviruses differing in their symptomatology was investigated. CYDV-RPV displays milder symptoms in oat and wheat whereas CYDV-RPS is responsible for more severe disease. We showed that both P0 proteins (P0(CY-RPV) and P0(CY-RPS)) were able to suppress local RNA silencing induced by either sense or inverted repeat transgenes in an Agrobacterium tumefaciens-mediated expression assay in Nicotiana benthamiana. P0(CY-RPS) displayed slightly higher activity. Systemic spread of the silencing signal was not impaired. Analysis of short-interfering RNA (siRNA) abundance revealed that accumulation of primary siRNA was not affected, but secondary siRNA levels were reduced by both CYDV P0 proteins, suggesting that they act downstream of siRNA production. Correlated with this finding we showed that both P0 proteins partially destabilized ARGONAUTE1. Finally both P0(CY-RPV) and P0(CY-RPS) interacted in yeast cells with ASK2, a component of an E3-ubiquitin ligase, with distinct affinities. Copyright © 2015 Elsevier B.V. All rights reserved.

  7. Drosophila PAF1 Modulates PIWI/piRNA Silencing Capacity.

    Science.gov (United States)

    Clark, Josef P; Rahman, Reazur; Yang, Nachen; Yang, Linda H; Lau, Nelson C

    2017-09-11

    To test the directness of factors in initiating PIWI-directed gene silencing, we employed a Piwi-interacting RNA (piRNA)-targeted reporter assay in Drosophila ovary somatic sheet (OSS) cells [1]. This assay confirmed direct silencing roles for piRNA biogenesis factors and PIWI-associated factors [2-12] but suggested that chromatin-modifying proteins may act downstream of the initial silencing event. Our data also revealed that RNA-polymerase-II-associated proteins like PAF1 and RTF1 antagonize PIWI-directed silencing. PAF1 knockdown enhances PIWI silencing of reporters when piRNAs target the transcript region proximal to the promoter. Loss of PAF1 suppresses endogenous transposable element (TE) transcript maturation, whereas a subset of gene transcripts and long-non-coding RNAs adjacent to TE insertions are affected by PAF1 knockdown in a similar fashion to piRNA-targeted reporters. Additionally, transcription activation at specific TEs and TE-adjacent loci during PIWI knockdown is suppressed when PIWI and PAF1 levels are both reduced. Our study suggests a mechanistic conservation between fission yeast PAF1 repressing AGO1/small interfering RNA (siRNA)-directed silencing [13, 14] and Drosophila PAF1 opposing PIWI/piRNA-directed silencing. Copyright © 2017 Elsevier Ltd. All rights reserved.

  8. The interaction between endogenous 30S ribosomal subunit protein S11 and Cucumber mosaic virus LS2b protein affects viral replication, infection and gene silencing suppressor activity.

    Directory of Open Access Journals (Sweden)

    Ruilin Wang

    Full Text Available Cucumber mosaic virus (CMV is a model virus for plant-virus protein interaction and mechanism research because of its wide distribution, high-level of replication and simple genome structure. The 2b protein is a multifunctional protein encoded by CMV that suppresses RNA silencing-based antiviral defense and contributes to CMV virulence in host plants. In this report, 12 host proteins were identified as CMV LS2b binding partners using the yeast two-hybrid screen system from the Arabidopsis thaliana cDNA library. Among the host proteins, 30S ribosomal subunit protein S11 (RPS11 was selected for further studies. The interaction between LS2b and full-length RPS11 was confirmed using the yeast two-hybrid system. Bimolecular fluorescence complementation (BIFC assays observed by confocal laser microscopy and Glutathione S-transferase (GST pull-down assays were used to verify the interaction between endogenous NbRPS11 and viral CMVLS2b both in vivo and in vitro. TRV-based gene silencing vector was used to knockdown NbRPS11 transcription, and immunoblot analysis revealed a decline in infectious viral RNA replication and a decrease in CMV infection in RPS11 down-regulated Nicotiana benthamiana plants. Thus, the knockdown of RPS11 likely inhibited CMV replication and accumulation. The gene silencing suppressor activity of CMV2b protein was reduced by the RPS11 knockdown. This study demonstrated that the function of viral LS2b protein was remarkably affected by the interaction with host RPS11 protein.

  9. Tumor-suppressor activity of RRIG1 in breast cancer

    International Nuclear Information System (INIS)

    Zhang, Guihong; Brewster, Abenaa; Guan, Baoxiang; Fan, Zhen; Brown, Powel H; Xu, Xiao-Chun

    2011-01-01

    Retinoid receptor-induced gene-1 (RRIG1) is a novel gene that has been lost in several types of human cancers. The aim of this study was to determine whether RRIG1 plays a role in breast cancer, such as in the suppression of breast cancer cell growth and invasion. Immunohistochemistry was used to detect RRIG1 expression in breast tissue specimens. Gene transfection was used to restore or knock down RRIG1 expression in breast cancer cell lines for analysis of cell viability, colony formation, and migration/invasion potential. Reverse-transcription polymerase chain reaction and western blot assays were used to detect the changes in gene expression. The RhoA activation assay was used to assess RRIG1-induced inhibition of RhoA activity. The immunohistochemical data showed that RRIG1 expression was reduced in breast cancer tissues compared with normal and atypical hyperplastic breast tissues. RRIG1 expression was inversely correlated with lymph node metastasis of breast cancer but was not associated with the status of hormone receptors, such as estrogen receptor, progesterone receptor, or HER2. Furthermore, restoration of RRIG1 expression inhibited proliferation, colony formation, migration, and invasion of breast cancer cells. Expression of RRIG1 also reduced phosphorylated Erk1/2 and Akt levels; c-Jun, MMP9, and Akt expressions; and RhoA activity. In contrast, knockdown of RRIG1 expression promoted breast cancer cell proliferation, colony formation, migration, and invasion potential. The data from the current study indicated that RRIG1 expression was reduced or lost in breast cancer and that restoration of RRIG1 expression suppressed breast cancer cell growth and invasion capacity. Future studies will determine the underlying molecular mechanisms and define RRIG1 as a tumor-suppressor gene in breast cancer

  10. MicroRNA silencing in primates: towards development of novel therapeutics

    DEFF Research Database (Denmark)

    Petri, Andreas; Lindow, Morten; Kauppinen, Sakari

    2009-01-01

    MicroRNAs (miRNA) comprise an abundant class of small noncoding RNAs that act as important posttranscriptional regulators of gene expression. Accumulating evidence showing that aberrantly expressed miRNAs play important roles in human cancers underscores them as potential targets for therapeutic ...... intervention. Recent reports on efficient miRNA silencing in rodents and nonhuman primates using high-affinity targeting by chemically modified antisense oligonucleotides highlight the utility of such compounds in the development of miRNA-based cancer therapeutics....

  11. Nanosystems based on siRNA silencing HuR expression counteract diabetic retinopathy in rat.

    Science.gov (United States)

    Amadio, Marialaura; Pascale, Alessia; Cupri, Sarha; Pignatello, Rosario; Osera, Cecilia; D Agata, Velia; D Amico, Agata Grazia; Leggio, Gian Marco; Ruozi, Barbara; Govoni, Stefano; Drago, Filippo; Bucolo, Claudio

    2016-09-01

    We evaluated whether specifically and directly targeting human antigen R (HuR), a member of embryonic lethal abnormal vision (ELAV) proteins family, may represent a new potential therapeutic strategy to manage diabetic retinopathy. Nanosystems loaded with siRNA silencing HuR expression (lipoplexes), consisting of solid lipid nanoparticles (SLN) and liposomes (SUV) were prepared. Photon correlation spectroscopy analysis, Zeta potential measurement and atomic force microscopy (AFM) studies were carried out to characterize the complexation of siRNA with the lipid nanocarriers. Nanosystems were evaluated by using AFM and scanning electron microscopy. The lipoplexes were injected into the eye of streptozotocin (STZ)-induced diabetic rats. Retinal HuR and VEGF levels were detected by Western blot and ELISA, respectively. Retinal histology was also carried out. The results demonstrated that retinal HuR and VEGF are significantly increased in STZ-rats and are blunted by HuR siRNA treatment. Lipoplexes with a weak positive surface charge and with a 4:1 N/P (cationic lipid nitrogen to siRNA phosphate) ratio exert a better transfection efficiency, significantly dumping retinal HuR and VEGF levels. In conclusion, we demonstrated that siRNA can be efficiently delivered into the rat retina using lipid-based nanocarriers, and some of the lipoplexes loaded with siRNA silencing HuR expression are potential candidates to manage retinal diseases. Copyright © 2016 Elsevier Ltd. All rights reserved.

  12. [Lentivirus-mediated shRNA silencing of LAMP2A inhibits the proliferation of multiple myeloma cells].

    Science.gov (United States)

    Li, Lixuan; Li, Jia

    2015-05-01

    To study the effects of lentivirus-mediated short hairpin RNA (shRNA) silencing of lysosome-associated membrane protein type 2A (LAMP2A) expression on the proliferation of multiple myeloma cells. The constructed shRNA lentiviral vector was applied to infect human multiple myeloma cell line MM.1S, and stable expression cell line was obtained by puromycin screening. Western blotting was used to verify the inhibitory effect on LAMP2A protein expression. MTT assay was conducted to detect the effect of knocked-down LAMP2A on MM.1S cell proliferation, and the anti-tumor potency of suberoylanilide hydroxamic acid (SAHA) against the obtained MM.1S LAMP2A(shRNA) stable cell line. Lactate assay was performed to observe the impact of low LAMP2A expression on cell glycolysis. The stable cell line with low LAMP2A expression were obtained with the constructed human LAMP2A-shRNA lentiviral vector. Down-regulation of LAMP2A expression significantly inhibited MM.1S cell proliferation and enhanced the anti-tumor activity of SAHA. Interestingly, decreased LAMP2A expression also inhibited MM.1S cell lactic acid secretion. Down-regulation of LAMP2A expression could inhibit cell proliferation in multiple myeloma cells.

  13. The bifunctional abiotic stress signalling regulator and endogenous RNA silencing suppressor FIERY1 is required for lateral root formation

    KAUST Repository

    Chen, Hao; Xiong, Liming

    2010-01-01

    root defects. Although fry1 and xrn4 exhibited reduced sensitivity to ethylene, our experiments demonstrated that restoration of ethylene sensitivity in the fry1 mutant is not sufficient to rescue the lateral root phenotypes of fry1. Our results

  14. Tsw gene-based resistance is triggered by a functional RNA silencing suppressor protein of the Tomato spotted wilt virus

    NARCIS (Netherlands)

    Ronde, de D.; Butterbach, P.B.E.; Lohuis, H.; Hedil, M.; Lent, van J.W.M.; Kormelink, R.J.M.

    2013-01-01

    As a result of contradictory reports, the avirulence (Avr) determinant that triggers Tsw gene-based resistance in Capsicum annuum against the Tomato spotted wilt virus (TSWV) is still unresolved. Here, the N and NSs genes of resistance-inducing (RI) and resistance-breaking (RB) isolates were cloned

  15. The human ARF tumor suppressor senses blastema activity and suppresses epimorphic tissue regeneration

    Science.gov (United States)

    Hesse, Robert G; Kouklis, Gayle K; Ahituv, Nadav; Pomerantz, Jason H

    2015-01-01

    The control of proliferation and differentiation by tumor suppressor genes suggests that evolution of divergent tumor suppressor repertoires could influence species’ regenerative capacity. To directly test that premise, we humanized the zebrafish p53 pathway by introducing regulatory and coding sequences of the human tumor suppressor ARF into the zebrafish genome. ARF was dormant during development, in uninjured adult fins, and during wound healing, but was highly expressed in the blastema during epimorphic fin regeneration after amputation. Regenerative, but not developmental signals resulted in binding of zebrafish E2f to the human ARF promoter and activated conserved ARF-dependent Tp53 functions. The context-dependent activation of ARF did not affect growth and development but inhibited regeneration, an unexpected distinct tumor suppressor response to regenerative versus developmental environments. The antagonistic pleiotropic characteristics of ARF as both tumor and regeneration suppressor imply that inducing epimorphic regeneration clinically would require modulation of ARF –p53 axis activation. DOI: http://dx.doi.org/10.7554/eLife.07702.001 PMID:26575287

  16. Expression profiling of c-kit and its impact after esiRNA silencing during gonadal development in catfish.

    Science.gov (United States)

    Laldinsangi, C; Senthilkumaran, B

    2018-04-03

    C-kit receptor is a member of a family of growth factor receptors that have tyrosine kinase activity, and are involved in the transduction of growth regulatory signals across plasma membrane by activation of its ligand, kitl/scf. The present study analysed mRNA and protein expression profiles of c-kit in the gonads of catfish, Clarias gariepinus, using real time PCR, in situ hybridization and immunohistochemistry. Tissue distribution analysis revealed higher expression mainly in the catfish gonads. Ontogeny studies showed minimal expression during early developmental stages and highest during 50-75 days post hatch, and the dimorphic expression in gonads decreased gradually till adulthood, which might suggest an important role for this gene around later stages of sex differentiation and gonadal development. Expression of C-kit was analysed at various phases of gonadal cycle in both male and female, which showed minimal expression during the resting phase, and higher expression in male compared to females during the pre-spawning phase. In vitro and in vivo induction using human chorionic gonadotropin elevated the expression of c-kit indicating the regulatory influence of hypothalamo-hypophyseal axis. In vivo transient gene silencing using c-kit-esiRNA in adult catfish during gonadal recrudescence showed a decrease in c-kit expression, which affected the expression level of germ cell meiotic marker sycp3, as well as several factors and steroidogenic enzyme genes involved in germ cell development. Decrease in the levels of serum 11-KT and T were also observed after esiRNA silencing. The findings of this study suggest that c-kit has an important role in the process of germ cell proliferation, development and maturation during gonadal development and recrudescence in catfish. Copyright © 2018. Published by Elsevier Inc.

  17. Identification of Novel Fibrosis Modifiers by In Vivo siRNA Silencing

    Directory of Open Access Journals (Sweden)

    Elisabeth H. Vollmann

    2017-06-01

    Full Text Available Fibrotic diseases contribute to 45% of deaths in the industrialized world, and therefore a better understanding of the pathophysiological mechanisms underlying tissue fibrosis is sorely needed. We aimed to identify novel modifiers of tissue fibrosis expressed by myofibroblasts and their progenitors in their disease microenvironment through RNA silencing in vivo. We leveraged novel biology, targeting genes upregulated during liver and kidney fibrosis in this cell lineage, and employed small interfering RNA (siRNA-formulated lipid nanoparticles technology to silence these genes in carbon-tetrachloride-induced liver fibrosis in mice. We identified five genes, Egr2, Atp1a2, Fkbp10, Fstl1, and Has2, which modified fibrogenesis based on their silencing, resulting in reduced Col1a1 mRNA levels and collagen accumulation in the liver. These genes fell into different groups based on the effects of their silencing on a transcriptional mini-array and histological outcomes. Silencing of Egr2 had the broadest effects in vivo and also reduced fibrogenic gene expression in a human fibroblast cell line. Prior to our study, Egr2, Atp1a2, and Fkbp10 had not been functionally validated in fibrosis in vivo. Thus, our results provide a major advance over the existing knowledge of fibrogenic pathways. Our study is the first example of a targeted siRNA assay to identify novel fibrosis modifiers in vivo.

  18. Soluble suppressor supernatants elaborated by concanavalin A-activated human mononuclear cells. Characterization of a soluble suppressor of B cell immunoglobulin production

    International Nuclear Information System (INIS)

    Fleisher, T.A.; Greene, W.C.; Blaese, R.M.; Waldmann, T.A.

    1981-01-01

    Human peripheral blood mononuclear cells (PBMC) activated with the mitogenic lectin concanavalin A (Con A) elaborate a soluble immune suppressor supernatant (SISS) that contains at least 2 distinct suppressor factors. One of these, SISS-B, inhibits polyclonal B cell immunoglobulin production, whereas the other, SISS-T, suppresses T cell proliferation to both mitogens and antigens. The latter mediator is discussed in the companion paper. Characteristics of the human soluble suppressor of B cell immunoglobulin production (SISS-B) include: 1) inhibition by a noncytotoxic mechanism, 2) loss of activity in the presence of the monosaccharide L-rhamnose, 3) appearance within 8 to 16 hr after the addition of Con A, 4) elaboration by cells irradiated with 500 or 2000 rads, 5) production by highly purified T cells, 6) stability at pH 2.5 but instability at 56/sup o/C, and 7) m.w. of 60 to 80,000. These data indicate that after Con A activation, selected T cells not only become potent suppressor cells, but also generate a soluble saccharide-specific factor(s) that inhibits polyclonal immunoglobulin production by human B cells

  19. The Cucumber vein yellowing virus silencing suppressor P1b can functionally replace HCPro in Plum pox virus infection in a host-specific manner.

    Science.gov (United States)

    Carbonell, Alberto; Dujovny, Gabriela; García, Juan Antonio; Valli, Adrian

    2012-02-01

    Plant viruses of the genera Potyvirus and Ipomovirus (Potyviridae family) use unrelated RNA silencing suppressors (RSS) to counteract antiviral RNA silencing responses. HCPro is the RSS of Potyvirus spp., and its activity is enhanced by the upstream P1 protein. Distinctively, the ipomovirus Cucumber vein yellowing virus (CVYV) lacks HCPro but contains two P1 copies in tandem (P1aP1b), the second of which functions as RSS. Using chimeras based on the potyvirus Plum pox virus (PPV), we found that P1b can functionally replace HCPro in potyviral infections of Nicotiana plants. Interestingly, P1a, the CVYV protein homologous to potyviral P1, disrupted the silencing suppression activity of P1b and reduced the infection efficiency of PPV in Nicotiana benthamiana. Testing the influence of RSS in host specificity, we found that a P1b-expressing chimera poorly infected PPV's natural host, Prunus persica. Conversely, P1b conferred on PPV chimeras the ability to replicate locally in cucumber, CVYV's natural host. The deleterious effect of P1a on PPV infection is host dependent, because the P1aP1b-expressing PPV chimera accumulated in cucumber to higher levels than PPV expressing P1b alone. These results demonstrate that a potyvirus can use different RSS, and that particular RSS and upstream P1-like proteins contribute to defining the virus host range.

  20. The Polerovirus F box protein P0 targets ARGONAUTE1 to suppress RNA silencing.

    Science.gov (United States)

    Bortolamiol, Diane; Pazhouhandeh, Maghsoud; Marrocco, Katia; Genschik, Pascal; Ziegler-Graff, Véronique

    2007-09-18

    Plants employ post-transcriptional gene silencing (PTGS) as an antiviral defense response. In this mechanism, viral-derived small RNAs are incorporated into the RNA-induced silencing complex (RISC) to guide degradation of the corresponding viral RNAs. ARGONAUTE1 (AGO1) is a key component of RISC: it carries the RNA slicer activity. As a counter-defense, viruses have evolved various proteins that suppress PTGS. Recently, we showed that the Polerovirus P0 protein carries an F box motif required to form an SCF-like complex, which is also essential for P0's silencing suppressor function. Here, we investigate the molecular mechanism by which P0 impairs PTGS. First we show that P0's expression does not affect the biogenesis of primary siRNAs in an inverted repeat-PTGS assay, but it does affect their activity. Moreover, P0's expression in transformed Arabidopsis plants leads to various developmental abnormalities reminiscent of mutants affected in miRNA pathways, which is accompanied by enhanced levels of several miRNA-target transcripts, suggesting that P0 acts at the level of RISC. Interestingly, ectopic expression of P0 triggered AGO1 protein decay in planta. Finally, we provide evidence that P0 physically interacts with AGO1. Based on these results, we propose that P0 hijacks the host SCF machinery to modulate gene silencing by destabilizing AGO1.

  1. Alloantigen-specific suppressor T cells are not inhibited by cyclosporin A, but do require IL 2 for activation

    International Nuclear Information System (INIS)

    Bucy, R.P.

    1986-01-01

    Alloantigen-specific suppressor T cells are activated from normal murine spleen cells in mixed lymphocyte reactions (MLR). These T cells are radioresistant and suppress the activation of cytotoxic T lymphocytes (CTL) in second primary MLR cultures. This report demonstrates that cyclosporin A (CsA) blocks the activation of these suppressor cells at a dose of 1 microgram/ml. However, reconstitution of CsA blocked cultures with IL 2 restores the activation of the suppressor T cells, but fails to significantly restore the activation of CTL in these same cultures. This differential activation requirement was used to establish T cell lines that demonstrate enriched suppressor cell activity but depletion of CTL activity. These findings are discussed in terms of the mechanism of action of CsA in these distinct T cell subsets and the relevance to models of allograft unresponsiveness

  2. HC-Pro silencing suppressor significantly alters the gene expression profile in tobacco leaves and flowers

    Directory of Open Access Journals (Sweden)

    Lehto Kirsi

    2011-04-01

    Full Text Available Abstract Background RNA silencing is used in plants as a major defence mechanism against invasive nucleic acids, such as viruses. Accordingly, plant viruses have evolved to produce counter defensive RNA-silencing suppressors (RSSs. These factors interfere in various ways with the RNA silencing machinery in cells, and thereby disturb the microRNA (miRNA mediated endogene regulation and induce developmental and morphological changes in plants. In this study we have explored these effects using previously characterized transgenic tobacco plants which constitutively express (under CaMV 35S promoter the helper component-proteinase (HC-Pro derived from a potyviral genome. The transcript levels of leaves and flowers of these plants were analysed using microarray techniques (Tobacco 4 × 44 k, Agilent. Results Over expression of HC-Pro RSS induced clear phenotypic changes both in growth rate and in leaf and flower morphology of the tobacco plants. The expression of 748 and 332 genes was significantly changed in the leaves and flowers, respectively, in the HC-Pro expressing transgenic plants. Interestingly, these transcriptome alterations in the HC-Pro expressing tobacco plants were similar as those previously detected in plants infected with ssRNA-viruses. Particularly, many defense-related and hormone-responsive genes (e.g. ethylene responsive transcription factor 1, ERF1 were differentially regulated in these plants. Also the expression of several stress-related genes, and genes related to cell wall modifications, protein processing, transcriptional regulation and photosynthesis were strongly altered. Moreover, genes regulating circadian cycle and flowering time were significantly altered, which may have induced a late flowering phenotype in HC-Pro expressing plants. The results also suggest that photosynthetic oxygen evolution, sugar metabolism and energy levels were significantly changed in these transgenic plants. Transcript levels of S

  3. Metabolic activity is necessary for activation of T suppressor cells by B cells

    International Nuclear Information System (INIS)

    Elkins, K.L.; Stashak, P.W.; Baker, P.J.

    1990-01-01

    Ag-primed B cells must express cell-surface IgM, but not IgD or Ia Ag, and must remain metabolically active, in order to activate suppressor T cells (Ts) specific for type III pneumococcal polysaccharide. Ag-primed B cells that were gamma-irradiated with 1000r, or less, retained the ability to activate Ts; however, Ag-primed B cells exposed to UV light were not able to do so. gamma-Irradiated and UV-treated Ag-primed B cells both expressed comparable levels of cell-surface IgM, and both localized to the spleen after in vivo transfer; neither could proliferate in vitro in response to mitogens. By contrast, gamma-irradiated primed B cells were still able to synthesize proteins, whereas UV-treated primed B cells could not. These findings suggest that in order for Ag-primed B cells to activate Ts, they must (a) express cell-associated IgM (sIgM) antibody bearing the idiotypic determinants of antibody specific for type III pneumococcal polysaccharide, and (b) be able to synthesize protein for either the continued expression of sIgM after cell transfer, or for the elaboration of another protein molecule that is also required for the activation of Ts; this molecule does not appear to be Ia Ag

  4. PhOBF1, a petunia ocs element binding factor, plays an important role in antiviral RNA silencing.

    Science.gov (United States)

    Sun, Daoyang; Li, Shaohua; Niu, Lixin; Reid, Michael S; Zhang, Yanlong; Jiang, Cai-Zhong

    2017-02-01

    Virus-induced gene silencing (VIGS) is a common reverse genetics strategy for characterizing the function of genes in plants. The detailed mechanism governing RNA silencing efficiency triggered by viruses is largely unclear. Here, we reveal that a petunia (Petunia hybrida) ocs element binding factor, PhOBF1, one of the basic leucine zipper (bZIP) transcription factors, was up-regulated by Tobacco rattle virus (TRV) infection. Simultaneous silencing of PhOBF1 and a reporter gene, phytoene desaturase (PDS) or chalcone synthase (CHS), by TRV-based VIGS led to a failure of the development of leaf photobleaching or the white-corollas phenotype. PhOBF1 silencing caused down-regulation of RNA silencing-related genes, including RNA-dependent RNA polymerases (RDRs), Dicer-like RNase III enzymes (DCLs), and Argonautes (AGOs). After inoculation with the TRV-PhPDS, PhOBF1-RNAi lines exhibited a substantially impaired PDS silencing efficiency, whereas overexpression of PhOBF1 resulted in a recovery of the silencing phenotype (photobleaching) in systemic leaves. A compromised resistance to TRV and Tobacco mosaic virus was found in PhOBF1-RNAi lines, while PhOBF1-overexpressing lines displayed an enhanced resistance to their infections. Compared with wild-type plants, PhOBF1-silenced plants accumulated lower levels of free salicylic acid (SA), salicylic acid glucoside, and phenylalanine, contrarily to higher levels of those in plants overexpressing PhOBF1. Furthermore, transcripts of a number of genes associated with the shikimate and phenylpropanoid pathways were decreased or increased in PhOBF1-RNAi or PhOBF1-overexpressing lines, respectively. Taken together, the data suggest that PhOBF1 regulates TRV-induced RNA silencing efficiency through modulation of RDRs, DCLs, and AGOs mediated by the SA biosynthesis pathway. © The Author 2017. Published by Oxford University Press on behalf of the Society for Experimental Biology.

  5. A petunia ethylene-responsive element binding factor, PhERF2, plays an important role in antiviral RNA silencing.

    Science.gov (United States)

    Sun, Daoyang; Nandety, Raja Sekhar; Zhang, Yanlong; Reid, Michael S; Niu, Lixin; Jiang, Cai-Zhong

    2016-05-01

    Virus-induced RNA silencing is involved in plant antiviral defense and requires key enzyme components, including RNA-dependent RNA polymerases (RDRs), Dicer-like RNase III enzymes (DCLs), and Argonaute proteins (AGOs). However, the transcriptional regulation of these critical components is largely unknown. In petunia (Petunia hybrida), an ethylene-responsive element binding factor, PhERF2, is induced by Tobacco rattle virus (TRV) infection. Inclusion of a PhERF2 fragment in a TRV silencing construct containing reporter fragments of phytoene desaturase (PDS) or chalcone synthase (CHS) substantially impaired silencing efficiency of both the PDS and CHS reporters. Silencing was also impaired in PhERF2- RNAi lines, where TRV-PhPDS infection did not show the expected silencing phenotype (photobleaching). In contrast, photobleaching in response to infiltration with the TRV-PhPDS construct was enhanced in plants overexpressing PhERF2 Transcript abundance of the RNA silencing-related genes RDR2, RDR6, DCL2, and AGO2 was lower in PhERF2-silenced plants but higher in PhERF2-overexpressing plants. Moreover, PhERF2-silenced lines showed higher susceptibility to Cucumber mosaic virus (CMV) than wild-type (WT) plants, while plants overexpressing PhERF2 exhibited increased resistance. Interestingly, growth and development of PhERF2-RNAi lines were substantially slower, whereas the overexpressing lines were more vigorous than the controls. Taken together, our results indicate that PhERF2 functions as a positive regulator in antiviral RNA silencing. © The Author 2016. Published by Oxford University Press on behalf of the Society for Experimental Biology.

  6. Argonaute Utilization for miRNA Silencing Is Determined by Phosphorylation-Dependent Recruitment of LIM-Domain-Containing Proteins

    Directory of Open Access Journals (Sweden)

    Katherine S. Bridge

    2017-07-01

    Full Text Available As core components of the microRNA-induced silencing complex (miRISC, Argonaute (AGO proteins interact with TNRC6 proteins, recruiting other effectors of translational repression/mRNA destabilization. Here, we show that LIMD1 coordinates the assembly of an AGO-TNRC6 containing miRISC complex by binding both proteins simultaneously at distinct interfaces. Phosphorylation of AGO2 at Ser 387 by Akt3 induces LIMD1 binding, which in turn enables AGO2 to interact with TNRC6A and downstream effector DDX6. Conservation of this serine in AGO1 and 4 indicates this mechanism may be a fundamental requirement for AGO function and miRISC assembly. Upon CRISPR-Cas9-mediated knockout of LIMD1, AGO2 miRNA-silencing function is lost and miRNA silencing becomes dependent on a complex formed by AGO3 and the LIMD1 family member WTIP. The switch to AGO3 utilization occurs due to the presence of a glutamic acid residue (E390 on the interaction interface, which allows AGO3 to bind to LIMD1, AJUBA, and WTIP irrespective of Akt signaling.

  7. Concanavalin A-induced and spontaneous suppressor cell activities in peripheral blood lymphocytes and spleen cells from gastric cancer patients.

    Science.gov (United States)

    Toge, T; Hamamoto, S; Itagaki, E; Yajima, K; Tanada, M; Nakane, H; Kohno, H; Nakanishi, K; Hattori, T

    1983-11-01

    In 173 gastric cancer patients, activities of Concanavalin-A-induced suppressor cells (Con-AS) and spontaneous suppressor cells (SpS) in peripheral blood lymphocytes (PBL), splenic vein lymphocytes (SVL), and spleen cells (SCs) were investigated. Suppressions by Con-AS in PBL were significantly effective in patients of Stages III and IV, while suppressions by SpS were effective in patients with recurrent tumors. Thus, in PBLs of cancer patients, suppressor precursors, which are considered to be activated in vitro by Concanavalin-A, seemed to appear with the advances of the disease, and SpS activities, which could be already activated in vivo, seemed to increase in the terminal stage. In SCs, increased activities of Con-AS, but normal activities of SpS, were observed, and these suppressor-cell populations consisted of glass nonadherent cells. Suppressor activities of SCs would be due to suppressor T-cells, not to other types of cells. Furthermore, Con-AS existed in the medium-sized lymphocytes, which were fractionated on the basis of cell size, while SpS in the large-sized lymphocytes. A higher proportion of T-cells, bearing Fc receptors for IgG, was observed in the larger-sized lymphocyte fractions. Cell numbers in the large-sized lymphocyte fraction tended to increase with the advances of tumors. From these results, it is suggested that higher presence of suppressor precursors and the increase of SpS activities may occur in cancer patients, depending on the tumor advancing.

  8. Ring structure amino acids affect the suppressor activity of melon aphid-borne yellows virus P0 protein.

    Science.gov (United States)

    Han, Yan-Hong; Xiang, Hai-Ying; Wang, Qian; Li, Yuan-Yuan; Wu, Wen-Qi; Han, Cheng-Gui; Li, Da-Wei; Yu, Jia-Lin

    2010-10-10

    Melon aphid-borne yellows virus (MABYV) is a newly identified polerovirus occurring in China. Here, we demonstrate that the MABYV encoded P0 (P0(MA)) protein is a strong suppressor of post-transcriptional gene silencing (PTGS) with activity comparable to tobacco etch virus (TEV) HC-Pro. In addition we have shown that the LP F-box motif present at the N-terminus of P0(MA) is required for suppressor activity. Detailed mutational analyses on P0(MA) revealed that changing the conserved Trp 212 with non-ring structured amino acids altered silencing suppressor functions. Ala substitutions at positions 12 and 211 for Phe had no effect on P0 suppression-activity, whereas Arg and Glu substitutions had greatly decreased suppressor activity. Furthermore, substitutions targeting Phe at position 30 also resulted in reduced P0 suppression-activity. Altogether, these results suggest that ring structured Trp/Phe residues in P0 have important roles in suppressor activity. Copyright © 2010 Elsevier Inc. All rights reserved.

  9. Molecular chaperone complexes with antagonizing activities regulate stability and activity of the tumor suppressor LKB1.

    Science.gov (United States)

    Gaude, H; Aznar, N; Delay, A; Bres, A; Buchet-Poyau, K; Caillat, C; Vigouroux, A; Rogon, C; Woods, A; Vanacker, J-M; Höhfeld, J; Perret, C; Meyer, P; Billaud, M; Forcet, C

    2012-03-22

    LKB1 is a tumor suppressor that is constitutionally mutated in a cancer-prone condition, called Peutz-Jeghers syndrome, as well as somatically inactivated in a sizeable fraction of lung and cervical neoplasms. The LKB1 gene encodes a serine/threonine kinase that associates with the pseudokinase STRAD (STE-20-related pseudokinase) and the scaffolding protein MO25, the formation of this heterotrimeric complex promotes allosteric activation of LKB1. We have previously reported that the molecular chaperone heat shock protein 90 (Hsp90) binds to and stabilizes LKB1. Combining pharmacological studies and RNA interference approaches, we now provide evidence that the co-chaperone Cdc37 participates to the regulation of LKB1 stability. It is known that the Hsp90-Cdc37 complex recognizes a surface within the N-terminal catalytic lobe of client protein kinases. In agreement with this finding, we found that the chaperones Hsp90 and Cdc37 interact with an LKB1 isoform that differs in the C-terminal region, but not with a novel LKB1 variant that lacks a portion of the kinase N-terminal lobe domain. Reconstitution of the two complexes LKB1-STRAD and LKB1-Hsp90-Cdc37 with recombinant proteins revealed that the former is catalytically active whereas the latter is inactive. Furthermore, consistent with a documented repressor function of Hsp90, LKB1 kinase activity was transiently stimulated upon dissociation of Hsp90. Finally, disruption of the LKB1-Hsp90 complex favors the recruitment of both Hsp/Hsc70 and the U-box dependent E3 ubiquitin ligase CHIP (carboxyl terminus of Hsc70-interacting protein) that triggers LKB1 degradation. Taken together, our results establish that the Hsp90-Cdc37 complex controls both the stability and activity of the LKB1 kinase. This study further shows that two chaperone complexes with antagonizing activities, Hsp90-Cdc37 and Hsp/Hsc70-CHIP, finely control the cellular level of LKB1 protein.

  10. Tumor Suppressor Genes within Common Fragile Sites Are Active Players in the DNA Damage Response.

    Directory of Open Access Journals (Sweden)

    Idit Hazan

    2016-12-01

    Full Text Available The role of common fragile sites (CFSs in cancer remains controversial. Two main views dominate the discussion: one suggests that CFS loci are hotspots of genomic instability leading to inactivation of genes encoded within them, while the other view proposes that CFSs are functional units and that loss of the encoded genes confers selective pressure, leading to cancer development. The latter view is supported by emerging evidence showing that expression of a given CFS is associated with genome integrity and that inactivation of CFS-resident tumor suppressor genes leads to dysregulation of the DNA damage response (DDR and increased genomic instability. These two viewpoints of CFS function are not mutually exclusive but rather coexist; when breaks at CFSs are not repaired accurately, this can lead to deletions by which cells acquire growth advantage because of loss of tumor suppressor activities. Here, we review recent advances linking some CFS gene products with the DDR, genomic instability, and carcinogenesis and discuss how their inactivation might represent a selective advantage for cancer cells.

  11. Tumor suppressors: enhancers or suppressors of regeneration?

    Science.gov (United States)

    Pomerantz, Jason H.; Blau, Helen M.

    2013-01-01

    Tumor suppressors are so named because cancers occur in their absence, but these genes also have important functions in development, metabolism and tissue homeostasis. Here, we discuss known and potential functions of tumor suppressor genes during tissue regeneration, focusing on the evolutionarily conserved tumor suppressors pRb1, p53, Pten and Hippo. We propose that their activity is essential for tissue regeneration. This is in contrast to suggestions that tumor suppression is a trade-off for regenerative capacity. We also hypothesize that certain aspects of tumor suppressor pathways inhibit regenerative processes in mammals, and that transient targeted modification of these pathways could be fruitfully exploited to enhance processes that are important to regenerative medicine. PMID:23715544

  12. Oral cancer cells may rewire alternative metabolic pathways to survive from siRNA silencing of metabolic enzymes

    International Nuclear Information System (INIS)

    Zhang, Min; Chai, Yang D; Brumbaugh, Jeffrey; Liu, Xiaojun; Rabii, Ramin; Feng, Sizhe; Misuno, Kaori; Messadi, Diana; Hu, Shen

    2014-01-01

    Cancer cells may undergo metabolic adaptations that support their growth as well as drug resistance properties. The purpose of this study is to test if oral cancer cells can overcome the metabolic defects introduced by using small interfering RNA (siRNA) to knock down their expression of important metabolic enzymes. UM1 and UM2 oral cancer cells were transfected with siRNA to transketolase (TKT) or siRNA to adenylate kinase (AK2), and Western blotting was used to confirm the knockdown. Cellular uptake of glucose and glutamine and production of lactate were compared between the cancer cells with either TKT or AK2 knockdown and those transfected with control siRNA. Statistical analysis was performed with student T-test. Despite the defect in the pentose phosphate pathway caused by siRNA knockdown of TKT, the survived UM1 or UM2 cells utilized more glucose and glutamine and secreted a significantly higher amount of lactate than the cells transferred with control siRNA. We also demonstrated that siRNA knockdown of AK2 constrained the proliferation of UM1 and UM2 cells but similarly led to an increased uptake of glucose/glutamine and production of lactate by the UM1 or UM2 cells survived from siRNA silencing of AK2. Our results indicate that the metabolic defects introduced by siRNA silencing of metabolic enzymes TKT or AK2 may be compensated by alternative feedback metabolic mechanisms, suggesting that cancer cells may overcome single defective pathways through secondary metabolic network adaptations. The highly robust nature of oral cancer cell metabolism implies that a systematic medical approach targeting multiple metabolic pathways may be needed to accomplish the continued improvement of cancer treatment

  13. F-box-like domain in the polerovirus protein P0 is required for silencing suppressor function

    Science.gov (United States)

    Pazhouhandeh, Maghsoud; Dieterle, Monika; Marrocco, Katia; Lechner, Esther; Berry, Bassam; Brault, Véronique; Hemmer, Odile; Kretsch, Thomas; Richards, Kenneth E.; Genschik, Pascal; Ziegler-Graff, Véronique

    2006-01-01

    Plants employ small RNA-mediated posttranscriptional gene silencing as a virus defense mechanism. In response, plant viruses encode proteins that can suppress RNA silencing, but the mode of action of most such proteins is poorly understood. Here, we show that the silencing suppressor protein P0 of two Arabidopsis-infecting poleroviruses interacts by means of a conserved minimal F-box motif with Arabidopsis thaliana orthologs of S-phase kinase-related protein 1 (SKP1), a component of the SCF family of ubiquitin E3 ligases. Point mutations in the F-box-like motif abolished the P0–SKP1 ortholog interaction, diminished virus pathogenicity, and inhibited the silencing suppressor activity of P0. Knockdown of expression of a SKP1 ortholog in Nicotiana benthamiana rendered the plants resistant to polerovirus infection. Together, the results support a model in which P0 acts as an F-box protein that targets an essential component of the host posttranscriptional gene silencing machinery. PMID:16446454

  14. Host natural suppressor activity regulates hemopoietic engraftment kinetics in antibody-conditioned recipient mice

    International Nuclear Information System (INIS)

    Sadelain, M.W.; Green, D.R.; Wegmann, T.G.

    1990-01-01

    Resistance to semi-allogeneic or syngeneic hemopoietic stem cell engraftment can be reduced by treating the unirradiated host with anti-class I MHC antibody. In our previous studies we showed a direct correlation between such resistance and the level of natural suppressor (NS) activity in the host. Thus newborn mice that have high NS activity are very resistant to marrow engraftment, as are adults pretreated with CFA that increases NS activity in the bone marrow. We have now devised a method that allows us to follow hemopoietic engraftment kinetics within the marrow cavity itself by assaying individual CFU-granulocyte/macrophage progenitor cells for their host or donor origin over the immediate post-transplant period. By using this method, we find a close correlation between the rate of marrow engraftment and reduction in host NS activity. Marrow engraftment does not correlate with the reduction of either total host bone marrow cellular content or CFU-granulocyte/macrophage progenitor cell levels. NS activity is mediated by Thy-1-, partially radiosensitive, nylon wool nonadherent cells without NK activity. Adoptively transferred Thy-1-, irradiated spleen cells containing NS activity induced by pretreatment with CFA delayed engraftment kinetics in the marrow cavity. Thus hemopoietic engraftment in the marrow cavity appears to be controlled by an inhibitory regulatory activity that is reflected in the in vitro NS assay. These studies suggest new regulatory targets for selective host conditioning to eliminate resistance to marrow transplantation

  15. Activated T cells sustain myeloid-derived suppressor cell-mediated immune suppression

    Science.gov (United States)

    Damuzzo, Vera; Francescato, Samuela; Pozzuoli, Assunta; Berizzi, Antonio; Mocellin, Simone; Rossi, Carlo Riccardo; Bronte, Vincenzo; Mandruzzato, Susanna

    2016-01-01

    The expansion of myeloid derived suppressor cells (MDSCs), a suppressive population able to hamper the immune response against cancer, correlates with tumor progression and overall survival in several cancer types. We have previously shown that MDSCs can be induced in vitro from precursors present in the bone marrow and observed that these cells are able to actively proliferate in the presence of activated T cells, whose activation level is critical to drive the suppressive activity of MDSCs. Here we investigated at molecular level the mechanisms involved in the interplay between MDSCs and activated T cells. We found that activated T cells secrete IL-10 following interaction with MDSCs which, in turn, activates STAT3 phosphorylation on MDSCs then leading to B7-H1 expression. We also demonstrated that B7-H1+ MDSCs are responsible for immune suppression through a mechanism involving ARG-1 and IDO expression. Finally, we show that the expression of ligands B7-H1 and MHC class II both on in vitro-induced MDSCs and on MDSCs in the tumor microenvironment of cancer patients is paralleled by an increased expression of their respective receptors PD-1 and LAG-3 on T cells, two inhibitory molecules associated with T cell dysfunction. These findings highlight key molecules and interactions responsible for the extensive cross-talk between MDSCs and activated T cells that are at the basis of immune suppression. PMID:26700461

  16. Immune suppressor factor confers stromal cell line with enhanced supporting activity for hematopoietic stem cells

    International Nuclear Information System (INIS)

    Nakajima, Hideaki; Shibata, Fumi; Fukuchi, Yumi; Goto-Koshino, Yuko; Ito, Miyuki; Urano, Atsushi; Nakahata, Tatsutoshi; Aburatani, Hiroyuki; Kitamura, Toshio

    2006-01-01

    Immune suppressor factor (ISF) is a subunit of the vacuolar ATPase proton pump. We earlier identified a short form of ISF (ShIF) as a stroma-derived factor that supports cytokine-independent growth of mutant Ba/F3 cells. Here, we report that ISF/ShIF supports self-renewal and expansion of primary hematopoietic stem cells (HSCs). Co-culture of murine bone marrow cells with a stromal cell line overexpressing ISF or ShIF (MS10/ISF or MS10/ShIF) not only enhanced their colony-forming activity and the numbers of long-term culture initiating cells, but also maintained the competitive repopulating activity of HSC. This stem cell supporting activity depended on the proton-transfer function of ISF/ShIF. Gene expression analysis of ISF/ShIF-transfected cell lines revealed down-regulation of secreted frizzled-related protein-1 and tissue inhibitor of metalloproteinase-3, and the restoration of their expressions in MS10/ISF cells partially reversed its enhanced LTC-IC supporting activity to a normal level. These results suggest that ISF/ShIF confers stromal cells with enhanced supporting activities for HSCs by modulating Wnt-activity and the extracellular matrix

  17. Targeting myeloid-derived suppressor cells augments antitumor activity against lung cancer

    Directory of Open Access Journals (Sweden)

    Srivastava MK

    2012-10-01

    Full Text Available Minu K Srivastava,1,2 Li Zhu,1,2 Marni Harris-White,2 Min Huang,1–3 Maie St John,1,3 Jay M Lee,1,3 Ravi Salgia,4 Robert B Cameron,1,3,5 Robert Strieter,6 Steven Dubinett,1–3 Sherven Sharma1–31Department of Medicine, UCLA Lung Cancer Research Program, David Geffen School of Medicine at UCLA, Los Angeles, CA, 2Molecular Gene Medicine Laboratory, Veterans Affairs Greater Los Angeles Healthcare System, Los Angeles, CA, 3Jonsson Comprehensive Cancer Center, David Geffen School of Medicine at UCLA, Los Angeles, CA, 4Department of Medicine, University of Chicago, Chicago, IL, 5Department of Surgery, Veterans Affairs Greater Los Angeles Healthcare System, Los Angeles, CA, 6Department of Medicine, University of Virginia, Charlottesville, VA, USAAbstract: Lung cancer evades host immune surveillance by dysregulating inflammation. Tumors and their surrounding stromata produce growth factors, cytokines, and chemokines that recruit, expand, and/or activate myeloid-derived suppressor cells (MDSCs. MDSCs regulate immune responses and are frequently found in malignancy. In this review the authors discuss tumor-MDSC interactions that suppress host antitumor activities and the authors' recent findings regarding MDSC depletion that led to improved therapeutic vaccination responses against lung cancer. Despite the identification of a repertoire of tumor antigens, hurdles persist for immune-based anticancer therapies. It is likely that combined therapies that address the multiple immune deficits in cancer patients will be required for effective therapy. MDSCs play a major role in the suppression of T-cell activation and they sustain tumor growth, proliferation, and metastases. Regulation of MDSC recruitment, differentiation or expansion, and inhibition of the MDSC suppressive function with pharmacologic agents will be useful in the control of cancer growth and progression. Pharmacologic agents that regulate MDSCs may be more effective when combined with

  18. ZFP226 is a novel artificial transcription factor for selective activation of tumor suppressor KIBRA.

    Science.gov (United States)

    Schelleckes, Katrin; Schmitz, Boris; Lenders, Malte; Mewes, Mirja; Brand, Stefan-Martin; Brand, Eva

    2018-03-09

    KIBRA has been suggested as a key regulator of the hippo pathway, regulating organ size, cell contact inhibition as well as tissue regeneration and tumorigenesis. Recently, alterations of KIBRA expression caused by promotor methylation have been reported for several types of cancer. Our current study aimed to design an artificial transcription factor capable of re-activating expression of the tumor suppressor KIBRA and the hippo pathway. We engineered a new gene named 'ZFP226' encoding for a ~23 kDa fusion protein. ZFP226 belongs to the Cys2-His2 zinc finger type and recognizes a nine base-pair DNA sequence 5'-GGC-GGC-GGC-3' in the KIBRA core promoter P1a. ZFP226 showed nuclear localization in human immortalized kidney epithelial cells and activated the KIBRA core promoter (p < 0.001) resulting in significantly increased KIBRA mRNA and protein levels (p < 0.001). Furthermore, ZFP226 led to activation of hippo signaling marked by elevated YAP and LATS phosphorylation. In Annexin V flow cytometry assays ZFP226 overexpression showed strong pro-apoptotic capacity on MCF-7 breast cancer cells (p < 0.01 early-, p < 0.001 late-apoptotic cells). We conclude that the artificial transcription factor ZFP226 can be used for target KIBRA and hippo pathway activation. This novel molecule may represent a molecular tool for the development of future applications in cancer treatment.

  19. Tumor Suppressor Activity of the EphB2 Receptor in Prostate Cancer

    National Research Council Canada - National Science Library

    Pasquale, Elena B

    2007-01-01

    Mutations have been recently identified in the EphB2 receptor gene in prostate cancer suggesting that EphB2, a member of the large Eph receptor tyrosine kinase family, is a tumor suppressor in prostate cancer...

  20. Tumor Suppressor Activity of the EphB2 Receptor in Prostate Cancer

    National Research Council Canada - National Science Library

    Pasquale, Elena B

    2006-01-01

    Mutations have been recently identified in the EphB2 receptor gene in prostate cancer suggesting that EphB2, a member of the large Eph receptor tyrosine kinase family, is a tumor suppressor in prostate cancer...

  1. Regulation of the activity of the tumor suppressor PTEN by thioredoxin in Drosophila melanogaster

    International Nuclear Information System (INIS)

    Song, Zuohe; Saghafi, Negin; Gokhale, Vijay; Brabant, Marc; Meuillet, Emmanuelle J.

    2007-01-01

    Human Thioredoxin-1 (hTrx-1) is a small redox protein with a molecular weight of 12 kDa that contains two cysteine residues found in its catalytic site. HTrx-1 plays an important role in cell growth, apoptosis, and cancer patient prognosis. Recently, we have demonstrated that hTrx-1 binds to the C2 domain of the human tumor suppressor, PTEN, in a redox dependent manner. This binding leads to the inhibition of PTEN lipid phosphatase activity in mammalian tissue culture systems. In this study, we show that over-expression of hTrx-1 in Drosophila melanogaster promotes cell growth and proliferation during eye development as measured by eye size and ommatidia size. Furthermore, hTrx-1 rescues the small eye phenotype induced by the over-expression of PTEN. We demonstrate that this rescue of the PTEN-induced eye size phenotype requires cysteine-218 in the C2 domain of PTEN. We also show that hTrx-1 over-expression results in increased Akt phosphorylation in fly head extracts supporting our observations that the hTrx-1-induced eye size increase results from the inhibition of PTEN activity. Our study confirms the redox regulation of PTEN through disulfide bond formation with the hTrx-1 in Drosophila and suggests conserved mechanisms for thioredoxins and their interactions with the phosphatidylinositol-3-kinase signaling pathway in humans and fruit flies

  2. Characterization of membrane determinant in old T-cells with suppressor activity

    International Nuclear Information System (INIS)

    Hendricks, L.C.; Heidrick, M.L.

    1986-01-01

    T-cell function declines with age. Many T-cell functions are initiated at the cell membrane; therefore, age-related membrane alterations may contribute to loss of function. They have previously reported developing a monoclonal antibody, HH-AGE-T(1), which recognizes a cell with suppressor activity and binds to 15-20% of the T-cells from old BC3F 1 mice, but only to 0-4% of young T-cells. To further characterize the determinant recognized by HH-AGE-T(1), they analyzed immunoprecipitates (IP) of young and old T-cell membranes by 2D-SDS PAGE, followed by Western blotting. Immunodetection of the blots showed that HH-AGE-T(1) bound a heterodimer (66 kD, pI 8.44 and 36 kD, pI 5.82-7.12 subunits) in IP from old mice; but not young mice. Monoclonal anti-Lyt 2 antibody did not bind the determinant. When IP of iodinated T-cells were run on SDS-PAGE gels followed by blotting and autoradiography of the blots, very prominent bands were detected in the old sample and faint bands were detected in the young sample. These results suggest that HH-AGE-T(1) recognizes a membrane protein which is present in small amounts on young T-cells but which increases markedly with age. Further studies are needed to determine the significance of this age-related membrane change

  3. MUC1-C activates polycomb repressive complexes and downregulates tumor suppressor genes in human cancer cells.

    Science.gov (United States)

    Rajabi, Hasan; Hiraki, Masayuki; Kufe, Donald

    2018-04-01

    The PRC2 and PRC1 complexes are aberrantly expressed in human cancers and have been linked to decreases in patient survival. MUC1-C is an oncoprotein that is also overexpressed in diverse human cancers and is associated with a poor prognosis. Recent studies have supported a previously unreported function for MUC1-C in activating PRC2 and PRC1 in cancer cells. In the regulation of PRC2, MUC1-C (i) drives transcription of the EZH2 gene, (ii) binds directly to EZH2, and (iii) enhances occupancy of EZH2 on target gene promoters with an increase in H3K27 trimethylation. Regarding PRC1, which is recruited to PRC2 sites in the hierarchical model, MUC1-C induces BMI1 transcription, forms a complex with BMI1, and promotes H2A ubiquitylation. MUC1-C thereby contributes to the integration of PRC2 and PRC1-mediated repression of tumor suppressor genes, such as CDH1, CDKN2A, PTEN and BRCA1. Like PRC2 and PRC1, MUC1-C is associated with the epithelial-mesenchymal transition (EMT) program, cancer stem cell (CSC) state, and acquisition of anticancer drug resistance. In concert with these observations, targeting MUC1-C downregulates EZH2 and BMI1, inhibits EMT and the CSC state, and reverses drug resistance. These findings emphasize the significance of MUC1-C as a therapeutic target for inhibiting aberrant PRC function and reprogramming the epigenome in human cancers.

  4. Persistent ER stress induces the spliced leader RNA silencing pathway (SLS, leading to programmed cell death in Trypanosoma brucei.

    Directory of Open Access Journals (Sweden)

    Hanoch Goldshmidt

    2010-01-01

    Full Text Available Trypanosomes are parasites that cycle between the insect host (procyclic form and mammalian host (bloodstream form. These parasites lack conventional transcription regulation, including factors that induce the unfolded protein response (UPR. However, they possess a stress response mechanism, the spliced leader RNA silencing (SLS pathway. SLS elicits shut-off of spliced leader RNA (SL RNA transcription by perturbing the binding of the transcription factor tSNAP42 to its cognate promoter, thus eliminating trans-splicing of all mRNAs. Induction of endoplasmic reticulum (ER stress in procyclic trypanosomes elicits changes in the transcriptome similar to those induced by conventional UPR found in other eukaryotes. The mechanism of up-regulation under ER stress is dependent on differential stabilization of mRNAs. The transcriptome changes are accompanied by ER dilation and elevation in the ER chaperone, BiP. Prolonged ER stress induces SLS pathway. RNAi silencing of SEC63, a factor that participates in protein translocation across the ER membrane, or SEC61, the translocation channel, also induces SLS. Silencing of these genes or prolonged ER stress led to programmed cell death (PCD, evident by exposure of phosphatidyl serine, DNA laddering, increase in reactive oxygen species (ROS production, increase in cytoplasmic Ca(2+, and decrease in mitochondrial membrane potential, as well as typical morphological changes observed by transmission electron microscopy (TEM. ER stress response is also induced in the bloodstream form and if the stress persists it leads to SLS. We propose that prolonged ER stress induces SLS, which serves as a unique death pathway, replacing the conventional caspase-mediated PCD observed in higher eukaryotes.

  5. Antiviral RNA silencing initiated in the absence of RDE-4, a double-stranded RNA binding protein, in Caenorhabditis elegans.

    Science.gov (United States)

    Guo, Xunyang; Zhang, Rui; Wang, Jeffrey; Lu, Rui

    2013-10-01

    Small interfering RNAs (siRNAs) processed from double-stranded RNA (dsRNA) of virus origins mediate potent antiviral defense through a process referred to as RNA interference (RNAi) or RNA silencing in diverse organisms. In the simple invertebrate Caenorhabditis elegans, the RNAi process is initiated by a single Dicer, which partners with the dsRNA binding protein RDE-4 to process dsRNA into viral siRNAs (viRNAs). Notably, in C. elegans this RNA-directed viral immunity (RDVI) also requires a number of worm-specific genes for its full antiviral potential. One such gene is rsd-2 (RNAi spreading defective 2), which was implicated in RDVI in our previous studies. In the current study, we first established an antiviral role by showing that rsd-2 null mutants permitted higher levels of viral RNA accumulation, and that this enhanced viral susceptibility was reversed by ectopic expression of RSD-2. We then examined the relationship of rsd-2 with other known components of RNAi pathways and established that rsd-2 functions in a novel pathway that is independent of rde-4 but likely requires the RNA-dependent RNA polymerase RRF-1, suggesting a critical role for RSD-2 in secondary viRNA biogenesis, likely through coordinated action with RRF-1. Together, these results suggest that RDVI in the single-Dicer organism C. elegans depends on the collective actions of both RDE-4-dependent and RDE-4-independent mechanisms to produce RNAi-inducing viRNAs. Our study reveals, for the first time, a novel siRNA-producing mechanism in C. elegans that bypasses the need for a dsRNA-binding protein.

  6. Reconstruction of gene regulatory modules from RNA silencing of IFN-α modulators: experimental set-up and inference method.

    Science.gov (United States)

    Grassi, Angela; Di Camillo, Barbara; Ciccarese, Francesco; Agnusdei, Valentina; Zanovello, Paola; Amadori, Alberto; Finesso, Lorenzo; Indraccolo, Stefano; Toffolo, Gianna Maria

    2016-03-12

    Inference of gene regulation from expression data may help to unravel regulatory mechanisms involved in complex diseases or in the action of specific drugs. A challenging task for many researchers working in the field of systems biology is to build up an experiment with a limited budget and produce a dataset suitable to reconstruct putative regulatory modules worth of biological validation. Here, we focus on small-scale gene expression screens and we introduce a novel experimental set-up and a customized method of analysis to make inference on regulatory modules starting from genetic perturbation data, e.g. knockdown and overexpression data. To illustrate the utility of our strategy, it was applied to produce and analyze a dataset of quantitative real-time RT-PCR data, in which interferon-α (IFN-α) transcriptional response in endothelial cells is investigated by RNA silencing of two candidate IFN-α modulators, STAT1 and IFIH1. A putative regulatory module was reconstructed by our method, revealing an intriguing feed-forward loop, in which STAT1 regulates IFIH1 and they both negatively regulate IFNAR1. STAT1 regulation on IFNAR1 was object of experimental validation at the protein level. Detailed description of the experimental set-up and of the analysis procedure is reported, with the intent to be of inspiration for other scientists who want to realize similar experiments to reconstruct gene regulatory modules starting from perturbations of possible regulators. Application of our approach to the study of IFN-α transcriptional response modulators in endothelial cells has led to many interesting novel findings and new biological hypotheses worth of validation.

  7. Short Hairpin RNA Silencing of PHD-2 Improves Neovascularization and Functional Outcomes in Diabetic Wounds and Ischemic Limbs.

    Directory of Open Access Journals (Sweden)

    Kevin J Paik

    Full Text Available The transcription factor hypoxia-inducible factor 1-alpha (HIF-1α is responsible for the downstream expression of over 60 genes that regulate cell survival and metabolism in hypoxic conditions as well as those that enhance angiogenesis to alleviate hypoxia. However, under normoxic conditions, HIF-1α is hydroxylated by prolyl hydroxylase 2, and subsequently degraded, with a biological half-life of less than five minutes. Here we investigated the therapeutic potential of inhibiting HIF-1α degradation through short hairpin RNA silencing of PHD-2 in the setting of diabetic wounds and limb ischemia. Treatment of diabetic mouse fibroblasts with shPHD-2 in vitro resulted in decreased levels of PHD-2 transcript demonstrated by qRT-PCR, higher levels of HIF-1α as measured by western blot, and higher expression of the downstream angiogenic genes SDF-1 and VEGFα, as measured by qRT-PCR. In vivo, shPHD-2 accelerated healing of full thickness excisional wounds in diabetic mice compared to shScr control, (14.33 ± 0.45 days vs. 19 ± 0.33 days and was associated with an increased vascular density. Delivery of shPHD-2 also resulted in improved perfusion of ischemic hind limbs compared to shScr, prevention of distal digit tip necrosis, and increased survival of muscle tissue. Knockdown of PHD-2 through shRNA treatment has the potential to stimulate angiogenesis through overexpression of HIF-1α and upregulation of pro-angiogenic genes downstream of HIF-1α, and may represent a viable, non-viral approach to gene therapy for ischemia related applications.

  8. An intronic microRNA silences genes that are functionally antagonistic to its host gene.

    Science.gov (United States)

    Barik, Sailen

    2008-09-01

    MicroRNAs (miRNAs) are short noncoding RNAs that down-regulate gene expression by silencing specific target mRNAs. While many miRNAs are transcribed from their own genes, nearly half map within introns of 'host' genes, the significance of which remains unclear. We report that transcriptional activation of apoptosis-associated tyrosine kinase (AATK), essential for neuronal differentiation, also generates miR-338 from an AATK gene intron that silences a family of mRNAs whose protein products are negative regulators of neuronal differentiation. We conclude that an intronic miRNA, transcribed together with the host gene mRNA, may serve the interest of its host gene by silencing a cohort of genes that are functionally antagonistic to the host gene itself.

  9. Knockdown of TFIIS by RNA silencing inhibits cancer cell proliferation and induces apoptosis

    International Nuclear Information System (INIS)

    Hubbard, Kyle; Catalano, Jennifer; Puri, Raj K; Gnatt, Averell

    2008-01-01

    A common element among cancer cells is the presence of improperly controlled transcription. In these cells, the degree of specific activation of some genes is abnormal, and altering the aberrant transcription may therefore directly target cancer. TFIIS is a transcription elongation factor, which directly binds the transcription motor, RNA Polymerase II and allows it to read through various transcription arrest sites. We report on RNA interference of TFIIS, a transcription elongation factor, and its affect on proliferation of cancer cells in culture. RNA interference was performed by transfecting siRNA to specifically knock down TFIIS expression in MCF7, MCF10A, PL45 and A549 cells. Levels of TFIIS expression were determined by the Quantigene method, and relative protein levels of TFIIS, c-myc and p53 were determined by C-ELISA. Induction of apoptosis was determined by an enzymatic Caspase 3/7 assay, as well as a non-enzymatic assay detecting cytoplasmic mono- and oligonucleosomes. A gene array analysis was conducted for effects of TFIIS siRNA on MCF7 and MCF10A cell lines. Knockdown of TFIIS reduced cancer cell proliferation in breast, lung and pancreatic cancer cell lines. More specifically, TFIIS knockdown in the MCF7 breast cancer cell line induced cancer cell death and increased c-myc and p53 expression whereas TFIIS knockdown in the non-cancerous breast cell line MCF10A was less affected. Differential effects of TFIIS knockdown in MCF7 and MCF10A cells included the estrogenic, c-myc and p53 pathways, as observed by C-ELISA and gene array, and were likely involved in MCF7 cell-death. Although transcription is a fundamental process, targeting select core transcription factors may provide for a new and potent avenue for cancer therapeutics. In the present study, knockdown of TFIIS inhibited cancer cell proliferation, suggesting that TFIIS could be studied as a potential cancer target within the transcription machinery

  10. Natural killer activity and suppressor cells in irradiated mice repopulated with a mixture of cells from normal and 89Sr-treated donors

    International Nuclear Information System (INIS)

    Levy, E.M.; Kumar, V.; Bennett, M.

    1981-01-01

    Mice that have been injected with 89 Sr have fairly normal B and T cell function, but are abnormal in that they lack natural killer (NK) activity and other functions that require an intact bone marrow. These mice also have an increased potential for suppressor cell activity. We had previously shown that spleen cells from 89 Sr-treated mice could transfer low NK activity and increased suppressor cell function to lethally irradiated syngeneic recipients. To investigate the mechanisms involved in perpetuating these defects, groups of normal spleen or bone marrow cells. Recipients were assayed for their NK activity and suppressor cell function 5 to 14 wk later. it was found that the addition of normal cells in the donor inoculum resulted in normal NK activity. This indicates that low NK activity in 89 Sr-treated mice was not due to the presence of a suppressor cell that prevented NK cell generation. It was additionally found that low NK activity in recipient mice could be boosted by interferon inducers. This would indicate that NK activity in the recipients was not due to a lack of interferon-sensitive pre-NK cells. Suppressor cell function in recipient mice depended on the type and number of normal cells in the donor inoculum. Bone marrow cells were very efficient in overcoming the tendency to produce suppressor cells. It took approximately 20 times more normal spleen cells to produce the same results. The implications of these findings are discussed

  11. The RNA silencing enzyme RNA polymerase v is required for plant immunity.

    Directory of Open Access Journals (Sweden)

    Ana López

    2011-12-01

    Full Text Available RNA-directed DNA methylation (RdDM is an epigenetic control mechanism driven by small interfering RNAs (siRNAs that influence gene function. In plants, little is known of the involvement of the RdDM pathway in regulating traits related to immune responses. In a genetic screen designed to reveal factors regulating immunity in Arabidopsis thaliana, we identified NRPD2 as the OVEREXPRESSOR OF CATIONIC PEROXIDASE 1 (OCP1. NRPD2 encodes the second largest subunit of the plant-specific RNA Polymerases IV and V (Pol IV and Pol V, which are crucial for the RdDM pathway. The ocp1 and nrpd2 mutants showed increases in disease susceptibility when confronted with the necrotrophic fungal pathogens Botrytis cinerea and Plectosphaerella cucumerina. Studies were extended to other mutants affected in different steps of the RdDM pathway, such as nrpd1, nrpe1, ago4, drd1, rdr2, and drm1drm2 mutants. Our results indicate that all the mutants studied, with the exception of nrpd1, phenocopy the nrpd2 mutants; and they suggest that, while Pol V complex is required for plant immunity, Pol IV appears dispensable. Moreover, Pol V defective mutants, but not Pol IV mutants, show enhanced disease resistance towards the bacterial pathogen Pseudomonas syringae DC3000. Interestingly, salicylic acid (SA-mediated defenses effective against PsDC3000 are enhanced in Pol V defective mutants, whereas jasmonic acid (JA-mediated defenses that protect against fungi are reduced. Chromatin immunoprecipitation analysis revealed that, through differential histone modifications, SA-related defense genes are poised for enhanced activation in Pol V defective mutants and provide clues for understanding the regulation of gene priming during defense. Our results highlight the importance of epigenetic control as an additional layer of complexity in the regulation of plant immunity and point towards multiple components of the RdDM pathway being involved in plant immunity based on genetic evidence

  12. The RNA silencing enzyme RNA polymerase v is required for plant immunity.

    Science.gov (United States)

    López, Ana; Ramírez, Vicente; García-Andrade, Javier; Flors, Victor; Vera, Pablo

    2011-12-01

    RNA-directed DNA methylation (RdDM) is an epigenetic control mechanism driven by small interfering RNAs (siRNAs) that influence gene function. In plants, little is known of the involvement of the RdDM pathway in regulating traits related to immune responses. In a genetic screen designed to reveal factors regulating immunity in Arabidopsis thaliana, we identified NRPD2 as the OVEREXPRESSOR OF CATIONIC PEROXIDASE 1 (OCP1). NRPD2 encodes the second largest subunit of the plant-specific RNA Polymerases IV and V (Pol IV and Pol V), which are crucial for the RdDM pathway. The ocp1 and nrpd2 mutants showed increases in disease susceptibility when confronted with the necrotrophic fungal pathogens Botrytis cinerea and Plectosphaerella cucumerina. Studies were extended to other mutants affected in different steps of the RdDM pathway, such as nrpd1, nrpe1, ago4, drd1, rdr2, and drm1drm2 mutants. Our results indicate that all the mutants studied, with the exception of nrpd1, phenocopy the nrpd2 mutants; and they suggest that, while Pol V complex is required for plant immunity, Pol IV appears dispensable. Moreover, Pol V defective mutants, but not Pol IV mutants, show enhanced disease resistance towards the bacterial pathogen Pseudomonas syringae DC3000. Interestingly, salicylic acid (SA)-mediated defenses effective against PsDC3000 are enhanced in Pol V defective mutants, whereas jasmonic acid (JA)-mediated defenses that protect against fungi are reduced. Chromatin immunoprecipitation analysis revealed that, through differential histone modifications, SA-related defense genes are poised for enhanced activation in Pol V defective mutants and provide clues for understanding the regulation of gene priming during defense. Our results highlight the importance of epigenetic control as an additional layer of complexity in the regulation of plant immunity and point towards multiple components of the RdDM pathway being involved in plant immunity based on genetic evidence, but whether

  13. Insulin induces suppressor of cytokine signaling-3 tyrosine phosphorylation through janus-activated kinase

    NARCIS (Netherlands)

    Peraldi, P; Filloux, C; Emanuelli, B; Hilton, DJ; Van Obberghen, E

    2001-01-01

    Suppressor of cytokine signaling (SOCS) proteins were originally described as cytokine-induced molecules involved in negative feedback loops. We have shown that SOCS-3 is also a component of the insulin signaling network (1), Indeed, insulin leads to SOCS-3 expression in 3T3-L1 adipocytes. Once

  14. Adsorption behavior of redox-active suppressor additives: Combined electrochemical and STM studies

    International Nuclear Information System (INIS)

    Hai, N.T.M.; Huynh, T.M.T.; Fluegel, A.; Mayer, D.; Broekmann, P.

    2011-01-01

    Highlights: → Janus Green B and safranine are prototypical redox-active leveler additives for copper electroplating. → Their redox-transitions lie within the copper potential window. → Reduced additives are identified as active species for the leveling effect. → Electro-reduction affects in particular the central aromatic cores of the additives. - Abstract: The redox chemistry and the related surface phase behavior of Safranine (SAF) and Janus Green B (JGB) have been studied by means of cyclic voltammetry in combination with in situ Scanning Tunneling Microscopy using HOPG (Highly Oriented Pyrolytic Graphite) and single crystalline Cu(1 0 0) as model substrates, both revealing different widths of the accessible potential windows. JGB and SAF serve as prototypical heterocyclic suppressor/leveler additives that are used for the metallization of 3D-TSVs (3D Through Silicon Vias) following a classical 'leveling' concept. SAF can be considered as the reductive decomposition product of JGB that is formed at the copper/electrolyte interface upon electroplating. Both additives reveal a pronounced pH-dependent redox-chemistry with redox-transitions lying close to or even beyond the anodic limit of the copper potential window. Affected by these redox-processes are in particular the aromatic cores of those heterocycles that can be (quasi)reversibly reduced by a two electron transfer process within the potential window of copper. Therefore we identify the reduced form of those dyes as the active components for the suppressing/leveling effect in copper plating. STM data clearly shows a dye surface phase behavior that is crucially determined by its potential-dependent redox-chemistry. This will be exemplarily discussed for the SAF dye. On chloride-modified Cu(1 0 0) mono-reduced SAF forms a structurally well-defined monolayer of cationic stacking polymers. However, this coupled anion/cation layer reveals only minor suppressing capabilities with respect to the copper

  15. The wip1 phosphatase (PPM1D) antagonizes activation of the CHK2 tumor suppressor kinase

    Energy Technology Data Exchange (ETDEWEB)

    Manet, Oliva-Trastoy; Berthonaud, V.; Chevalier, A.; Ducrot, C.; Marsolier-Kergoat, M.C.; Mann, C.; Leteurtre, F. [CEA Saclay, DSV, DBJC, SBGM, Lab. du Controle du Cycle Cellulaire, 91 - Gif-sur-Yvette (France)

    2006-07-01

    The DNA checkpoints are signal transduction pathways that sense DNA damage and coordinate various responses such as cell cycle arrests, DNA repair or cell death. These pathways are particularly well conserved in eukaryotes and the family of the 'Checkpoint Kinases 2' genes (or CHK2) plays a major role in them. This family includes the Rad53 protein of the yeast Saccharomyces cerevisiae and its Chk2 human homologue. Rad53 plays a central part in DNA checkpoint: rad53d mutants (whose RAD53 gene has been deleted) are hypersensitive to all genotoxic stresses. Mice Chk2-1- cells are defective in the G1, the intra-S, and the G2/M checkpoints. Mutations in CHK2 have been associated to many forms o f cancer, either sporadic or hereditary which demonstrates Chk2 tumor suppressor function. Chk2 proteins are characterized by several conserved elements: (i) an N-terminal domain with a series of SQ/TQ motifs, preferential phosphorylation sites for the ATM/ATR kinases, (ii) an FHA domain (ForkHead Associated) that binds specifically to phosphorylated residues within TXXY motifs (with the Y residue depending on the FHA domain and conferring an extra specificity) and (iii) a kinase domain including an activation loop. The Chk2 protein is activated by phosphorylation of its threonine T68, mainly by ATM, upon DNA double-strand breaks. This phosphorylation allows for the homo-dimerization of Chk2 through the binding of phospho-T68 from one molecule to the FHA domain of another molecule. It results in trans auto-phosphorylations, especially at threonines T383 and T387 in the activation T-loop. Fully active Chk2 becomes monomeric and, diffusing through the whole nucleus, phosphorylates its targets (CDC25 A and CDC25C/cell cycle arrest; p53, E2F, PML/apoptosis; BRCA2/DNA repair). Chk2/Rad53 inactivation occurs in two cases: once the DNA lesions have been repaired (it is called recovery) or, under certain conditions, in the presence of unrepaired DNA damage (it is then called

  16. The wip1 phosphatase (PPM1D) antagonizes activation of the CHK2 tumor suppressor kinase

    International Nuclear Information System (INIS)

    Manet, Oliva-Trastoy; Berthonaud, V.; Chevalier, A.; Ducrot, C.; Marsolier-Kergoat, M.C.; Mann, C.; Leteurtre, F.

    2006-01-01

    The DNA checkpoints are signal transduction pathways that sense DNA damage and coordinate various responses such as cell cycle arrests, DNA repair or cell death. These pathways are particularly well conserved in eukaryotes and the family of the 'Checkpoint Kinases 2' genes (or CHK2) plays a major role in them. This family includes the Rad53 protein of the yeast Saccharomyces cerevisiae and its Chk2 human homologue. Rad53 plays a central part in DNA checkpoint: rad53d mutants (whose RAD53 gene has been deleted) are hypersensitive to all genotoxic stresses. Mice Chk2-1- cells are defective in the G1, the intra-S, and the G2/M checkpoints. Mutations in CHK2 have been associated to many forms o f cancer, either sporadic or hereditary which demonstrates Chk2 tumor suppressor function. Chk2 proteins are characterized by several conserved elements: (i) an N-terminal domain with a series of SQ/TQ motifs, preferential phosphorylation sites for the ATM/ATR kinases, (ii) an FHA domain (ForkHead Associated) that binds specifically to phosphorylated residues within TXXY motifs (with the Y residue depending on the FHA domain and conferring an extra specificity) and (iii) a kinase domain including an activation loop. The Chk2 protein is activated by phosphorylation of its threonine T68, mainly by ATM, upon DNA double-strand breaks. This phosphorylation allows for the homo-dimerization of Chk2 through the binding of phospho-T68 from one molecule to the FHA domain of another molecule. It results in trans auto-phosphorylations, especially at threonines T383 and T387 in the activation T-loop. Fully active Chk2 becomes monomeric and, diffusing through the whole nucleus, phosphorylates its targets (CDC25 A and CDC25C/cell cycle arrest; p53, E2F, PML/apoptosis; BRCA2/DNA repair). Chk2/Rad53 inactivation occurs in two cases: once the DNA lesions have been repaired (it is called recovery) or, under certain conditions, in the presence of unrepaired DNA damage (it is then called adaptation

  17. Viral RNA silencing suppression

    NARCIS (Netherlands)

    Hedil, Marcio; Kormelink, Richard

    2016-01-01

    The Bunyaviridae is a family of arboviruses including both plant-and vertebrate-infecting representatives. The Tospovirus genus accommodates plant-infecting bunyaviruses, which not only replicate in their plant host, but also in their insect thrips vector during persistent propagative

  18. SIGNALING TO THE P53 TUMOR SUPPRESSOR THROUGH PATHWAYS ACTIVATED BY GENOTOXIC AND NON-GENOTOXIC STRESSES.

    Energy Technology Data Exchange (ETDEWEB)

    ANDERSON,C.W.APPELLA,E.

    2002-07-01

    The p53 tumor suppressor is a tetrameric transcription factor that is post-translational modified at {approx}18 different sites by phosphorylation, acetylation, or sumoylation in response to various cellular stress conditions. Specific posttranslational modifications, or groups of modifications, that result from the activation of different stress-induced signaling pathways are thought to modulate p53 activity to regulate cell fate by inducing cell cycle arrest, apoptosis, or cellular senescence. Here we review the posttranslational modifications to p53 and the pathways that produce them in response to both genotoxic and non-genotoxic stresses.

  19. CMTM5 exhibits tumor suppressor activity through promoter methylation in oral squamous cell carcinoma

    International Nuclear Information System (INIS)

    Zhang, Heyu; Nan, Xu; Li, Xuefen; Chen, Yan; Zhang, Jianyun; Sun, Lisha; Han, Wenlin; Li, Tiejun

    2014-01-01

    Highlights: • Down-regulation of CMTM5 expression in OSCC tissues was found. • The promoter methylation status of CMTM5 was measured. • CMTM5-v1 inhibited cell proliferation and migration and induced apoptosis. • CMTM5 might act as a putative tumor suppressor gene in OSCC. - Abstract: Oral squamous cell carcinoma (OSCC) is one of the most common types of malignancies in the head and neck region. CKLF-like MARVEL transmembrane domain-containing member 5 (CMTM5) has been recently implicated as a tumor suppressor gene in several cancer types. Herein, we examined the expression and function of CMTM5 in oral squamous cell carcinoma. CMTM5 was down-regulated in oral squamous cell lines and tumor samples from patients with promoter methylation. Treatment with the demethylating agent 5-aza-2′-deoxycytidine restored CMTM5 expression. In the OSCC cell lines CAL27 and GNM, the ectopic expression of CMTM5-v1 strongly inhibited cell proliferation and migration and induced apoptosis. In addition, CMTM5-v1 inhibited tumor formation in vivo. Therefore, CMTM5 might act as a putative tumor suppressor gene through promoter methylation in oral squamous cell carcinoma

  20. CMTM5 exhibits tumor suppressor activity through promoter methylation in oral squamous cell carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Heyu [Central Laboratory, Peking University School of Stomatology, Beijing (China); Nan, Xu [Center for Human Disease Genomics, Department of Immunology, Key Laboratory of Medical Immunology, Ministry of Health, School of Basic Medical Sciences, Peking University, Beijing (China); Li, Xuefen [Central Laboratory, Peking University School of Stomatology, Beijing (China); Chen, Yan; Zhang, Jianyun [Department of Oral Pathology, Peking University School of Stomatology, Beijing (China); Sun, Lisha [Central Laboratory, Peking University School of Stomatology, Beijing (China); Han, Wenlin [Center for Human Disease Genomics, Department of Immunology, Key Laboratory of Medical Immunology, Ministry of Health, School of Basic Medical Sciences, Peking University, Beijing (China); Li, Tiejun, E-mail: litiejun22@vip.sina.com [Department of Oral Pathology, Peking University School of Stomatology, Beijing (China)

    2014-05-02

    Highlights: • Down-regulation of CMTM5 expression in OSCC tissues was found. • The promoter methylation status of CMTM5 was measured. • CMTM5-v1 inhibited cell proliferation and migration and induced apoptosis. • CMTM5 might act as a putative tumor suppressor gene in OSCC. - Abstract: Oral squamous cell carcinoma (OSCC) is one of the most common types of malignancies in the head and neck region. CKLF-like MARVEL transmembrane domain-containing member 5 (CMTM5) has been recently implicated as a tumor suppressor gene in several cancer types. Herein, we examined the expression and function of CMTM5 in oral squamous cell carcinoma. CMTM5 was down-regulated in oral squamous cell lines and tumor samples from patients with promoter methylation. Treatment with the demethylating agent 5-aza-2′-deoxycytidine restored CMTM5 expression. In the OSCC cell lines CAL27 and GNM, the ectopic expression of CMTM5-v1 strongly inhibited cell proliferation and migration and induced apoptosis. In addition, CMTM5-v1 inhibited tumor formation in vivo. Therefore, CMTM5 might act as a putative tumor suppressor gene through promoter methylation in oral squamous cell carcinoma.

  1. Deep sequencing uncovers commonality in small RNA profiles between transgene-induced and naturally occurring RNA silencing of chalcone synthase-A gene in petunia.

    Science.gov (United States)

    Kasai, Megumi; Matsumura, Hideo; Yoshida, Kentaro; Terauchi, Ryohei; Taneda, Akito; Kanazawa, Akira

    2013-01-30

    Introduction of a transgene that transcribes RNA homologous to an endogenous gene in the plant genome can induce silencing of both genes, a phenomenon termed cosuppression. Cosuppression was first discovered in transgenic petunia plants transformed with the CHS-A gene encoding chalcone synthase, in which nonpigmented sectors in flowers or completely white flowers are produced. Some of the flower-color patterns observed in transgenic petunias having CHS-A cosuppression resemble those in existing nontransgenic varieties. Although the mechanism by which white sectors are generated in nontransgenic petunia is known to be due to RNA silencing of the CHS-A gene as in cosuppression, whether the same trigger(s) and/or pattern of RNA degradation are involved in these phenomena has not been known. Here, we addressed this question using deep-sequencing and bioinformatic analyses of small RNAs. We analyzed short interfering RNAs (siRNAs) produced in nonpigmented sectors of petal tissues in transgenic petunia plants that have CHS-A cosuppression and a nontransgenic petunia variety Red Star, that has naturally occurring CHS-A RNA silencing. In both silencing systems, 21-nt and 22-nt siRNAs were the most and the second-most abundant size classes, respectively. CHS-A siRNA production was confined to exon 2, indicating that RNA degradation through the RNA silencing pathway occurred in this exon. Common siRNAs were detected in cosuppression and naturally occurring RNA silencing, and their ranks based on the number of siRNAs in these plants were correlated with each other. Noticeably, highly abundant siRNAs were common in these systems. Phased siRNAs were detected in multiple phases at multiple sites, and some of the ends of the regions that produced phased siRNAs were conserved. The features of siRNA production found to be common to cosuppression and naturally occurring silencing of the CHS-A gene indicate mechanistic similarities between these silencing systems especially in the

  2. Pharmacological activation of tumor suppressor, wild-type p53 as a promising strategy to fight cancer

    Directory of Open Access Journals (Sweden)

    Alicja Sznarkowska

    2010-08-01

    Full Text Available A powerful tumor suppressor – p53 protein is a transcription factor which plays a critical role in eliciting cellular responses to a variety of stress signals, including DNA damage, hypoxia and aberrant proliferative signals, such as oncogene activation. Since its discovery thirty one years ago, p53 has been connected to tumorigenesis as it accumulates in the transformed tumor cells. Cellular stress induces stabilization of p53 and promotes, depending on the stress level, cell cycle arrest or apoptosis in the irreversibly damaged cells. The p53 protein is found inactive in more than 50�0of human tumors either by enhanced proteasomal degradation or due to the inactivating point mutations in its gene. Numerous data indicate that low molecular weight compounds, identified by molecular modeling or in the functional, cell-based assays, efficiently activate non-mutated p53 in cancer cells which in consequence leads to their elimination due to p53-dependent apoptosis. In this work we describe the structure and cellular function of p53 as well as the latest discoveries on the compounds with high anti-tumor activities aiming at reactivation of the tumor suppressor function of p53.

  3. Epigenetic identification of ZNF545 as a functional tumor suppressor in multiple myeloma via activation of p53 signaling pathway

    Energy Technology Data Exchange (ETDEWEB)

    Fan, Yu [Chongqing Key Laboratory of Molecular Oncology and Epigenetics, The First Affiliated Hospital of Chongqing Medical University, Chongqing (China); Zhan, Qian [The Center for Clinical Molecular Medical Detection, The First Affiliated Hospital of Chongqing Medical University, Chongqing (China); Xu, Hongying [Chongqing Key Laboratory of Molecular Oncology and Epigenetics, The First Affiliated Hospital of Chongqing Medical University, Chongqing (China); Li, Lili; Li, Chen [Cancer Epigenetics Laboratory, Department of Clinical Oncology, Sir YK Pao Center for Cancer and Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong and CUHK Shenzhen Research Institute (Hong Kong); Xiao, Qian; Xiang, Shili; Hui, Tianli [Chongqing Key Laboratory of Molecular Oncology and Epigenetics, The First Affiliated Hospital of Chongqing Medical University, Chongqing (China); Xiang, Tingxiu, E-mail: larissaxiang@163.com [Chongqing Key Laboratory of Molecular Oncology and Epigenetics, The First Affiliated Hospital of Chongqing Medical University, Chongqing (China); Ren, Guosheng, E-mail: rengs726@126.com [Chongqing Key Laboratory of Molecular Oncology and Epigenetics, The First Affiliated Hospital of Chongqing Medical University, Chongqing (China)

    2016-06-10

    The KRAB–zinc-finger protein ZNF545 was recently identified as a potential suppressor gene in several tumors. However, the regulatory mechanisms of ZNF545 in tumorigenesis remain unclear. In this study, we investigated the expression and roles of ZNF545 in multiple myeloma (MM). ZNF545 was frequently downregulated in MM tissues compared with non-tumor bone marrow tissues. ZNF545 expression was silenced by promoter methylation in MM cell lines, and could be restored by demethylation treatment. ZNF545 methylation was detected in 28.3% of MM tissues, compared with 4.3% of normal bone marrow tissues. ZNF545 transcriptionally activated the p53 signaling pathway but had no effect on Akt in MM, whereas ectopic expression of ZNF545 in silenced cells suppressed their proliferation and induced apoptosis. We therefore identified ZNF545 as a novel tumor suppressor inhibiting tumor growth through activation of the p53 pathway in MM. Moreover, tumor-specific methylation of ZNF545 may represent an epigenetic biomarker for MM diagnosis, and a potential target for specific therapy. -- Highlights: •Downregulated ZNF545 in MM tissues and cell lines and ectopic expression of ZNF545 suppresses tumor growth. •Tumor-specific methylation of ZNF545 represents an epigenetic biomarker for MM diagnosis, and a potential target for specific therapy. •ZNF545 exerts its tumor suppressive effects via transcriptional activating p53 pathway.

  4. Epigenetic identification of ZNF545 as a functional tumor suppressor in multiple myeloma via activation of p53 signaling pathway

    International Nuclear Information System (INIS)

    Fan, Yu; Zhan, Qian; Xu, Hongying; Li, Lili; Li, Chen; Xiao, Qian; Xiang, Shili; Hui, Tianli; Xiang, Tingxiu; Ren, Guosheng

    2016-01-01

    The KRAB–zinc-finger protein ZNF545 was recently identified as a potential suppressor gene in several tumors. However, the regulatory mechanisms of ZNF545 in tumorigenesis remain unclear. In this study, we investigated the expression and roles of ZNF545 in multiple myeloma (MM). ZNF545 was frequently downregulated in MM tissues compared with non-tumor bone marrow tissues. ZNF545 expression was silenced by promoter methylation in MM cell lines, and could be restored by demethylation treatment. ZNF545 methylation was detected in 28.3% of MM tissues, compared with 4.3% of normal bone marrow tissues. ZNF545 transcriptionally activated the p53 signaling pathway but had no effect on Akt in MM, whereas ectopic expression of ZNF545 in silenced cells suppressed their proliferation and induced apoptosis. We therefore identified ZNF545 as a novel tumor suppressor inhibiting tumor growth through activation of the p53 pathway in MM. Moreover, tumor-specific methylation of ZNF545 may represent an epigenetic biomarker for MM diagnosis, and a potential target for specific therapy. -- Highlights: •Downregulated ZNF545 in MM tissues and cell lines and ectopic expression of ZNF545 suppresses tumor growth. •Tumor-specific methylation of ZNF545 represents an epigenetic biomarker for MM diagnosis, and a potential target for specific therapy. •ZNF545 exerts its tumor suppressive effects via transcriptional activating p53 pathway.

  5. Presence of activating KRAS mutations correlates significantly with expression of tumour suppressor genes DCN and TPM1 in colorectal cancer

    Directory of Open Access Journals (Sweden)

    Rems Miran

    2009-08-01

    Full Text Available Abstract Background Despite identification of the major genes and pathways involved in the development of colorectal cancer (CRC, it has become obvious that several steps in these pathways might be bypassed by other as yet unknown genetic events that lead towards CRC. Therefore we wanted to improve our understanding of the genetic mechanisms of CRC development. Methods We used microarrays to identify novel genes involved in the development of CRC. Real time PCR was used for mRNA expression as well as to search for chromosomal abnormalities within candidate genes. The correlation between the expression obtained by real time PCR and the presence of the KRAS mutation was investigated. Results We detected significant previously undescribed underexpression in CRC for genes SLC26A3, TPM1 and DCN, with a suggested tumour suppressor role. We also describe the correlation between TPM1 and DCN expression and the presence of KRAS mutations in CRC. When searching for chromosomal abnormalities, we found deletion of the TPM1 gene in one case of CRC, but no deletions of DCN and SLC26A3 were found. Conclusion Our study provides further evidence of decreased mRNA expression of three important tumour suppressor genes in cases of CRC, thus implicating them in the development of this type of cancer. Moreover, we found underexpression of the TPM1 gene in a case of CRCs without KRAS mutations, showing that TPM1 might serve as an alternative path of development of CRC. This downregulation could in some cases be mediated by deletion of the TPM1 gene. On the other hand, the correlation of DCN underexpression with the presence of KRAS mutations suggests that DCN expression is affected by the presence of activating KRAS mutations, lowering the amount of the important tumour suppressor protein decorin.

  6. Catalytic activity of matrix metalloproteinase-19 is essential for tumor suppressor and anti-angiogenic activities in nasopharyngeal carcinoma

    Czech Academy of Sciences Publication Activity Database

    Chan, K.C.; Ko, J.M.; Lung, H.L.; Sedláček, Radislav; Zhang, Z.F.; Luo, D.Z.; Feng, Z.B.; Chen, S.; Chen, H.; Chan, K.W.; Tsao, S.W.; Chua, D.T.; Zabarovsky, E.R.; Stanbridge, E.J.; Lung, M.L.

    2011-01-01

    Roč. 129, č. 8 (2011), s. 1826-1837 ISSN 0020-7136 Grant - others:Research Grants Council of the Hong Kong Special Administrative Region(CN) HKU661708M Institutional research plan: CEZ:AV0Z50520514 Keywords : MMP19 * nasopharyngeal carcinoma * tumor suppressor gene * angiogenesis Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.444, year: 2011

  7. Enhancement of the RAD51 Recombinase Activity by the Tumor Suppressor PALB2

    Energy Technology Data Exchange (ETDEWEB)

    Dray, Eloise; Etchin, Julia; Wiese, Claudia; Saro, Dorina; Williams, Gareth J.; Hammel, Michal; Yu, Xiong; Galkin, Vitold E.; Liu, Dongqing; Tsai, Miaw-Sheue; Sy, Shirley M-H.; Egelman, Edward; Chen, Junjie; Sung, Patrick; Schild, D.

    2010-08-24

    Homologous recombination mediated by the RAD51 recombinase helps eliminate chromosomal lesions, such as DNA double-stranded breaks induced by radiation or arising from injured DNA replication forks. The tumor suppressors BRCA2 and PALB2 act together to deliver RAD51 to chromosomal lesions to initiate repair. Here we document a new function of PALB2 in the enhancement of RAD51's ability to form the D-loop. We show that PALB2 binds DNA and physically interacts with RAD51. Importantly, while PALB2 alone stimulates D-loop formation, a cooperative effect is seen with RAD51AP1, an enhancer of RAD51. This stimulation stems from PALB2's ability to function with RAD51 and RAD51AP1 to assemble the synaptic complex. Our results help unveil a multi-faceted role of PALB2 in chromosome damage repair. Since PALB2 mutations can cause breast and other tumors or lead to Fanconi anemia, our findings are important for understanding the mechanism of tumor suppression in humans.

  8. Peroxisome proliferator-activated receptor gamma coactivator-1 alpha acts as a tumor suppressor in hepatocellular carcinoma.

    Science.gov (United States)

    Liu, Rui; Zhang, Haiyang; Zhang, Yan; Li, Shuang; Wang, Xinyi; Wang, Xia; Wang, Cheng; Liu, Bin; Zen, Ke; Zhang, Chen-Yu; Zhang, Chunni; Ba, Yi

    2017-04-01

    suppressor in the formation and development of hepatocellular carcinoma and that peroxisome proliferator-activated receptor gamma coactivator-1 alpha may be a potential therapeutic target for hepatocellular carcinoma.

  9. Activated NKT cells facilitated functional switch of myeloid-derived suppressor cells at inflammation sites in fulminant hepatitis mice.

    Science.gov (United States)

    Wu, Danxiao; Shi, Yu; Wang, Cheng; Chen, Hanwen; Liu, Qiaoyun; Liu, Jianhua; Zhang, Lihuang; Wu, Yihua; Xia, Dajing

    2017-02-01

    Myeloid-derived suppressor cells (MDSCs) confer immunosuppressive properties, but their roles in fulminant hepatitis have not been well defined. In this study, we systematically examined the distribution of MDSCs in bone marrow (BM), liver and spleen, and their functional and differentiation status in an acute fulminant hepatitis mouse model induced by lipopolysaccharide and D-galactosamine (LPS-GalN). Moreover, the interaction between NKT cells and MDSCs was determined. Our study revealed that BM contained the largest pool of MDSCs during pathogenesis of fulminant hepatitis compared with liver and spleen. MDSCs in liver/spleen expressed higher levels of chemokine receptors such as CCR2, CX3CR1 and CXCR2. At inflamed tissues such as liver or spleen, activated NKT cells induced differentiation of MDSCs through cell-cell interaction, which markedly dampened the immunosuppressive effects and promoted MDSCs to produce pro-inflammatory cytokines and activate inflammatory cells. Our findings thus demonstrated an unexpected pro-inflammatory state for MDSCs, which was mediated by the activated NKT cells that precipitated the differentiation and functional evolution of these MDSCs at sites of inflammation. Copyright © 2016. Published by Elsevier GmbH.

  10. Myeloid-Derived Suppressor Cells Specifically Suppress IFN-γ Production and Antitumor Cytotoxic Activity of Vδ2 T Cells

    Directory of Open Access Journals (Sweden)

    Alessandra Sacchi

    2018-06-01

    Full Text Available γδ T cells represent less than 5% of circulating T cells; they exert a potent cytotoxic function against tumor or infected cells and secrete cytokines like conventional αβ T cells. As αβ T cells γδ T cells reside in the typical T cell compartments (the lymph nodes and spleen, but are more widely distributed in tissues throughout the body. For these reasons, some investigators are exploring the possibility of immunotherapies aimed to expand and activate Vδ2 T cells, or using them as Chimeric Antigen Receptor carriers. However, the role of immunosuppressive microenvironment on Vδ2 T cells during infections and cancers has not been completely elucidated. In particular, the effects of myeloid-derived suppressor cells (MDSC, largely expanded in such pathologies, were not explored. In the present work, we demonstrated that MDSC may inhibit IFN-γ production and degranulation of phosphoantigen-activated Vδ2 T cells. Moreover, the Vδ2 T cells cytotoxic activity against the Burkitt lymphoma cell line Daudi and Jurkat cell line were impaired by MDSC. The Arginase I seems to be involved in the impairment of Vδ2 T cell function induced by both tumor cells and MDSC. These data open a key issue in the context of Vδ2-targeted immunoteraphy, suggesting the need of combined strategies aimed to boost Vδ2 T cells circumventing tumor- and MDSC-induced Vδ2 T cells suppression.

  11. AICAR Antiproliferative Properties Involve the AMPK-Independent Activation of the Tumor Suppressors LATS 1 and 2.

    Science.gov (United States)

    Philippe, Chloé; Pinson, Benoît; Dompierre, Jim; Pantesco, Véronique; Viollet, Benoît; Daignan-Fornier, Bertrand; Moenner, Michel

    2018-06-01

    AICAR (Acadesine) is a pharmacological precursor of purine nucleotide biosynthesis with anti-tumoral properties. Although recognized as an AMP mimetic activator of the protein kinase AMPK, the AICAR monophosphate derivative ZMP was also shown to mediate AMPK-independent effects. In order to unveil these AMPK-independent functions, we performed a transcriptomic analysis in AMPKα1/α2 double knockout murine embryonic cells. Kinetic analysis of the cellular response to AICAR revealed the up-regulation of the large tumor suppressor kinases (Lats) 1 and 2 transcripts, followed by the repression of numerous genes downstream of the transcriptional regulators Yap1 and Taz. This transcriptional signature, together with the observation of increased levels in phosphorylation of Lats1 and Yap1 proteins, suggested that the Hippo signaling pathway was activated by AICAR. This effect was observed in both fibroblasts and epithelial cells. Knockdown of Lats1/2 prevented the cytoplasmic delocalization of Yap1/Taz proteins in response to AICAR and conferred a higher resistance to the drug. These results indicate that activation of the most downstream steps of the Hippo cascade participates to the antiproliferative effects of AICAR. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

  12. The Putative PAX8/PPARγ Fusion Oncoprotein Exhibits Partial Tumor Suppressor Activity through Up-Regulation of Micro-RNA-122 and Dominant-Negative PPARγ Activity.

    Science.gov (United States)

    Reddi, Honey V; Madde, Pranathi; Milosevic, Dragana; Hackbarth, Jennifer S; Algeciras-Schimnich, Alicia; McIver, Bryan; Grebe, Stefan K G; Eberhardt, Norman L

    2011-01-01

    In vitro studies have demonstrated that the PAX8/PPARγ fusion protein (PPFP), which occurs frequently in follicular thyroid carcinomas (FTC), exhibits oncogenic activity. However, paradoxically, a meta-analysis of extant tumor outcome studies indicates that 68% of FTC-expressing PPFP are minimally invasive compared to only 32% of those lacking PPFP (χ(2) = 6.86, P = 0.008), suggesting that PPFP favorably impacts FTC outcomes. In studies designed to distinguish benign thyroid neoplasms from thyroid carcinomas, the previously identified tumor suppressor miR-122, a major liver micro-RNA (miR) that is decreased in hepatocellular carcinoma, was increased 8.9-fold (P negative PPARγ mutant in WRO cells was less effective than PPFP at inhibiting xenograft tumor progression (1.8-fold [P negative PPARγ activity. Up-regulation of miR-122 negatively regulates ADAM-17, a known downstream target, in thyroid cells, suggesting an antiangiogenic mechanism in thyroid carcinoma. This latter inference is directly supported by reduced CD-31 expression in WRO xenografts expressing PPFP, miR-122, and DN-PPARγ. We conclude that, in addition to its apparent oncogenic potential in vitro, PPFP exhibits paradoxical tumor suppressor activity in vivo, mediated by multiple mechanisms including up-regulation of miR-122 and dominant-negative inhibition of PPARγ activity.

  13. Viral RNAi suppressor reversibly binds siRNA to outcompete Dicer and RISC via multiple turnover.

    Science.gov (United States)

    Rawlings, Renata A; Krishnan, Vishalakshi; Walter, Nils G

    2011-04-29

    RNA interference is a conserved gene regulatory mechanism employed by most eukaryotes as a key component of their innate immune response to viruses and retrotransposons. During viral infection, the RNase-III-type endonuclease Dicer cleaves viral double-stranded RNA into small interfering RNAs (siRNAs) 21-24 nucleotides in length and helps load them into the RNA-induced silencing complex (RISC) to guide the cleavage of complementary viral RNA. As a countermeasure, many viruses have evolved viral RNA silencing suppressors (RSS) that tightly, and presumably quantitatively, bind siRNAs to thwart RNA-interference-mediated degradation. Viral RSS proteins also act across kingdoms as potential immunosuppressors in gene therapeutic applications. Here we report fluorescence quenching and electrophoretic mobility shift assays that probe siRNA binding by the dimeric RSS p19 from Carnation Italian Ringspot Virus, as well as by human Dicer and RISC assembly complexes. We find that the siRNA:p19 interaction is readily reversible, characterized by rapid binding [(1.69 ± 0.07) × 10(8) M(-)(1) s(-1)] and marked dissociation (k(off)=0.062 ± 0.002 s(-1)). We also observe that p19 efficiently competes with recombinant Dicer and inhibits the formation of RISC-related assembly complexes found in human cell extract. Computational modeling based on these results provides evidence for the transient formation of a ternary complex between siRNA, human Dicer, and p19. An expanded model of RNA silencing indicates that multiple turnover by reversible binding of siRNAs potentiates the efficiency of the suppressor protein. Our predictive model is expected to be applicable to the dosing of p19 as a silencing suppressor in viral gene therapy. Copyright © 2011 Elsevier Ltd. All rights reserved.

  14. Structure-activity studies of peptidomimetics based on kinase-inhibitory region of suppressors of cytokine signaling 1.

    Science.gov (United States)

    La Manna, Sara; Lopez-Sanz, Laura; Leone, Marilisa; Brandi, Paola; Scognamiglio, Pasqualina Liana; Morelli, Giancarlo; Novellino, Ettore; Gomez-Guerrero, Carmen; Marasco, Daniela

    2017-11-20

    Suppressors of Cytokine Signaling (SOCS) proteins are negative regulators of JAK proteins that are receptor-associated tyrosine kinases, which play key roles in the phosphorylation and subsequent activation of several transcription factors named STATs. Unlike the other SOCS proteins, SOCS1 and 3 show, in the N-terminal portion, a small kinase inhibitory region (KIR) involved in the inhibition of JAK kinases. Drug discovery processes of compounds based on KIR sequence demonstrated promising in functional in vitro and in inflammatory animal models and we recently developed a peptidomimetic called PS5, as lead compound. Here, we investigated the cellular ability of PS5 to mimic SOCS1 biological functions in vascular smooth muscle cells and simultaneously we set up a new binding assay for the screening and identification of JAK2 binders based on a SPR experiment that revealed more robust with respect to previous ELISAs. On this basis, we designed several peptidomimetics bearing new structural constraints that were analyzed in both affinities toward JAK2 and conformational features through Circular Dichroism and NMR spectroscopies. Introduced chemical modifications provided an enhancement of serum stabilities of new sequences that could aid the design of future mimetic molecules of SOCS1 as novel anti-inflammatory compounds. © 2017 Wiley Periodicals, Inc.

  15. Immortalization of MEF is characterized by the deregulation of specific miRNAs with potential tumor suppressor activity.

    Science.gov (United States)

    Rizzo, Milena; Evangelista, Monica; Simili, Marcella; Mariani, Laura; Pitto, Letizia; Rainaldi, Giuseppe

    2011-07-01

    The life span (Hayflick limit) of primary mouse embryo fibroblasts (MEF) in culture is variable but it is still unclear if the escape of the Hayflick limit is also variable. To address this point MEF were expanded every fifteen days (6T15) instead of every three days (6T3) until they became immortal. With this protocol MEF lifespan was extended and immortalization accordingly delayed. By testing a panel of genes (p19ARF, p16, p21) and miRNAs (miR-20a, miR-21, miR-28, miR-290) related to primary MEF senescence, a switch of p21 from up to down regulation, the down regulation of specific miRNAs as well as a massive shift from diploidy to hyperdiploidy were observed in coincidence with the resumption of cell proliferation. Collectively, these data indicate that the inactivation of genes and miRNAs, important in controlling cell proliferation, might be determinant for the escape from the Hayflick limit. In support of this hypothesis was the finding that some of the down regulated miRNAs transfected in immortalized MEF inhibited cell proliferation thus displaying a tumor suppressor-like activity.

  16. Myeloid-derived suppressor activity is mediated by monocytic lineages maintained by continuous inhibition of extrinsic and intrinsic death pathways.

    Science.gov (United States)

    Haverkamp, Jessica M; Smith, Amber M; Weinlich, Ricardo; Dillon, Christopher P; Qualls, Joseph E; Neale, Geoffrey; Koss, Brian; Kim, Young; Bronte, Vincenzo; Herold, Marco J; Green, Douglas R; Opferman, Joseph T; Murray, Peter J

    2014-12-18

    Nonresolving inflammation expands a heterogeneous population of myeloid suppressor cells capable of inhibiting T cell function. This heterogeneity has confounded the functional dissection of individual myeloid subpopulations and presents an obstacle for antitumor immunity and immunotherapy. Using genetic manipulation of cell death pathways, we found the monocytic suppressor-cell subset, but not the granulocytic subset, requires continuous c-FLIP expression to prevent caspase-8-dependent, RIPK3-independent cell death. Development of the granulocyte subset requires MCL-1-mediated control of the intrinsic mitochondrial death pathway. Monocytic suppressors tolerate the absence of MCL-1 provided cytokines increase expression of the MCL-1-related protein A1. Monocytic suppressors mediate T cell suppression, whereas their granulocytic counterparts lack suppressive function. The loss of the granulocytic subset via conditional MCL-1 deletion did not alter tumor incidence implicating the monocytic compartment as the functionally immunosuppressive subset in vivo. Thus, death pathway modulation defines the development, survival, and function of myeloid suppressor cells. Copyright © 2014 Elsevier Inc. All rights reserved.

  17. Immunosuppressive activity enhances central carbon metabolism and bioenergetics in myeloid-derived suppressor cells in vitro models

    Directory of Open Access Journals (Sweden)

    Hammami Ines

    2012-07-01

    Full Text Available Abstract Background The tumor microenvironment contains a vast array of pro- and anti-inflammatory cytokines that alter myelopoiesis and lead to the maturation of immunosuppressive cells known as myeloid-derived suppressor cells (MDSCs. Incubating bone marrow (BM precursors with a combination of granulocyte-macrophage colony-stimulating factor (GM-CSF and interleukin-6 (IL-6 generated a tumor-infiltrating MDSC-like population that impaired anti-tumor specific T-cell functions. This in vitro experimental approach was used to simulate MDSC maturation, and the cellular metabolic response was then monitored. A complementary experimental model that inhibited L-arginine (L-Arg metabolizing enzymes in MSC-1 cells, an immortalized cell line derived from primary MDSCs, was used to study the metabolic events related to immunosuppression. Results Exposure of BM cells to GM-CSF and IL-6 activated, within 24 h, L-Arg metabolizing enzymes which are responsible for the MDSCs immunosuppressive potential. This was accompanied by an increased uptake of L-glutamine (L-Gln and glucose, the latter being metabolized by anaerobic glycolysis. The up-regulation of nutrient uptake lead to the accumulation of TCA cycle intermediates and lactate as well as the endogenous synthesis of L-Arg and the production of energy-rich nucleotides. Moreover, inhibition of L-Arg metabolism in MSC-1 cells down-regulated central carbon metabolism activity, including glycolysis, glutaminolysis and TCA cycle activity, and led to a deterioration of cell bioenergetic status. The simultaneous increase of cell specific concentrations of ATP and a decrease in ATP-to-ADP ratio in BM-derived MDSCs suggested cells were metabolically active during maturation. Moreover, AMP-activated protein kinase (AMPK was activated during MDSC maturation in GM-CSF and IL-6–treated cultures, as revealed by the continuous increase of AMP-to-ATP ratios and the phosphorylation of AMPK. Likewise, AMPK activity was

  18. Modulation of immune response by alloactivated suppressor T cells

    International Nuclear Information System (INIS)

    Bernstein, A.; Sopori, M.L.; Gose, J.E.; Sondel, P.M.

    1979-01-01

    These studies show that there may be several different kinds of suppressor cells, each activated by different pathways and able to suppress different parts of the immune response either specifically or nonspecifically. As such, the physiology of one type of suppressor cell need not necessarily apply to that of another type of suppressor. Thus we emphasize the trap that the suppressor cell option provides: that is, virtually any previously inexplicable in vitro and in vivo immune phenomenon can always be adequately accounted for by evoking a suppressor mechanism, either by suppressing the response or suppressing the suppressor

  19. Slit-Robo GTPase-Activating Protein 2 as a metastasis suppressor in osteosarcoma

    OpenAIRE

    Marko, Tracy A.; Shamsan, Ghaidan A.; Edwards, Elizabeth N.; Hazelton, Paige E.; Rathe, Susan K.; Cornax, Ingrid; Overn, Paula R.; Varshney, Jyotika; Diessner, Brandon J.; Moriarity, Branden S.; O?Sullivan, M. Gerard; Odde, David J.; Largaespada, David A.

    2016-01-01

    Osteosarcoma is the most common primary bone tumor, with metastatic disease responsible for most treatment failure and patient death. A forward genetic screen utilizing Sleeping Beauty mutagenesis in mice previously identified potential genetic drivers of osteosarcoma metastasis, including Slit-Robo GTPase-Activating Protein 2 (Srgap2). This study evaluates the potential role of SRGAP2 in metastases-associated properties of osteosarcoma cell lines through Srgap2 knockout via the CRISPR/Cas9 n...

  20. A novel tumor suppressor function of glycine N-methyltransferase is independent of its catalytic activity but requires nuclear localization.

    Directory of Open Access Journals (Sweden)

    Suchandra DebRoy

    Full Text Available Glycine N-methyltransferase (GNMT, an abundant cytosolic enzyme, catalyzes the transfer of a methyl group from S-adenosylmethionine (SAM to glycine generating S-adenosylhomocysteine and sarcosine (N-methylglycine. This reaction is regulated by 5-methyltetrahydrofolate, which inhibits the enzyme catalysis. In the present study, we observed that GNMT is strongly down regulated in human cancers and is undetectable in cancer cell lines while the transient expression of the protein in cancer cells induces apoptosis and results in the activation of ERK1/2 as an early pro-survival response. The antiproliferative effect of GNMT can be partially reversed by treatment with the pan-caspase inhibitor zVAD-fmk but not by supplementation with high folate or SAM. GNMT exerts the suppressor effect primarily in cells originated from malignant tumors: transformed cell line of non-cancer origin, HEK293, was insensitive to GNMT. Of note, high levels of GNMT, detected in regenerating liver and in NIH3T3 mouse fibroblasts, do not produce cytotoxic effects. Importantly, GNMT, a predominantly cytoplasmic protein, was translocated into nuclei upon transfection of cancer cells. The presence of GNMT in the nuclei was also observed in normal human tissues by immunohistochemical staining. We further demonstrated that the induction of apoptosis is associated with the GNMT nuclear localization but is independent of its catalytic activity or folate binding. GNMT targeted to nuclei, through the fusion with nuclear localization signal, still exerts strong antiproliferative effects while its restriction to cytoplasm, through the fusion with nuclear export signal, prevents these effects (in each case the protein was excluded from cytosol or nuclei, respectively. Overall, our study indicates that GNMT has a secondary function, as a regulator of cellular proliferation, which is independent of its catalytic role.

  1. RNA silencing of Mcl-1 enhances ABT-737-mediated apoptosis in melanoma: role for a caspase-8-dependent pathway.

    Science.gov (United States)

    Keuling, Angela M; Felton, Kathleen E A; Parker, Arabesque A M; Akbari, Majid; Andrew, Susan E; Tron, Victor A

    2009-08-17

    Malignant melanoma is resistant to almost all conventional forms of chemotherapy. Recent evidence suggests that anti-apoptotic proteins of the Bcl-2 family are overexpressed in melanoma and may contribute to melanoma's striking resistance to apoptosis. ABT-737, a small-molecule inhibitor of Bcl-2, Bcl-xl and Bcl-w, has demonstrated efficacy in several forms of leukemia, lymphoma as well as solid tumors. However, overexpression of Mcl-1, a frequent observance in melanoma, is known to confer ABT-737 resistance. Here we report that knockdown of Mcl-1 greatly reduces cell viability in combination with ABT-737 in six different melanoma cell lines. We demonstrate that the cytotoxic effect of this combination treatment is due to apoptotic cell death involving not only caspase-9 activation but also activation of caspase-8, caspase-10 and Bid, which are normally associated with the extrinsic pathway of apoptosis. Caspase-8 (and caspase-10) activation is abrogated by inhibition of caspase-9 but not by inhibitors of the death receptor pathways. Furthermore, while caspase-8/-10 activity is required for the full induction of cell death with treatment, the death receptor pathways are not. Finally, we demonstrate that basal levels of caspase-8 and Bid correlate with treatment sensitivity. Our findings suggest that the combination of ABT-737 and Mcl-1 knockdown represents a promising, new treatment strategy for malignant melanoma. We also report a death receptor-independent role for extrinsic pathway proteins in treatment response and suggest that caspase-8 and Bid may represent potential markers of treatment sensitivity.

  2. RNA silencing of Mcl-1 enhances ABT-737-mediated apoptosis in melanoma: role for a caspase-8-dependent pathway.

    Directory of Open Access Journals (Sweden)

    Angela M Keuling

    Full Text Available BACKGROUND: Malignant melanoma is resistant to almost all conventional forms of chemotherapy. Recent evidence suggests that anti-apoptotic proteins of the Bcl-2 family are overexpressed in melanoma and may contribute to melanoma's striking resistance to apoptosis. ABT-737, a small-molecule inhibitor of Bcl-2, Bcl-xl and Bcl-w, has demonstrated efficacy in several forms of leukemia, lymphoma as well as solid tumors. However, overexpression of Mcl-1, a frequent observance in melanoma, is known to confer ABT-737 resistance. METHODOLOGY/PRINCIPAL FINDINGS: Here we report that knockdown of Mcl-1 greatly reduces cell viability in combination with ABT-737 in six different melanoma cell lines. We demonstrate that the cytotoxic effect of this combination treatment is due to apoptotic cell death involving not only caspase-9 activation but also activation of caspase-8, caspase-10 and Bid, which are normally associated with the extrinsic pathway of apoptosis. Caspase-8 (and caspase-10 activation is abrogated by inhibition of caspase-9 but not by inhibitors of the death receptor pathways. Furthermore, while caspase-8/-10 activity is required for the full induction of cell death with treatment, the death receptor pathways are not. Finally, we demonstrate that basal levels of caspase-8 and Bid correlate with treatment sensitivity. CONCLUSIONS/SIGNIFICANCE: Our findings suggest that the combination of ABT-737 and Mcl-1 knockdown represents a promising, new treatment strategy for malignant melanoma. We also report a death receptor-independent role for extrinsic pathway proteins in treatment response and suggest that caspase-8 and Bid may represent potential markers of treatment sensitivity.

  3. Reexamining the P-Element Invasion of Drosophila melanogaster Through the Lens of piRNA Silencing

    Science.gov (United States)

    Kelleher, Erin S.

    2016-01-01

    Transposable elements (TEs) are both important drivers of genome evolution and genetic parasites with potentially dramatic consequences for host fitness. The recent explosion of research on regulatory RNAs reveals that small RNA-mediated silencing is a conserved genetic mechanism through which hosts repress TE activity. The invasion of the Drosophila melanogaster genome by P elements, which happened on a historical timescale, represents an incomparable opportunity to understand how small RNA-mediated silencing of TEs evolves. Repression of P-element transposition emerged almost concurrently with its invasion. Recent studies suggest that this repression is implemented in part, and perhaps predominantly, by the Piwi-interacting RNA (piRNA) pathway, a small RNA-mediated silencing pathway that regulates TE activity in many metazoan germlines. In this review, I consider the P-element invasion from both a molecular and evolutionary genetic perspective, reconciling classic studies of P-element regulation with the new mechanistic framework provided by the piRNA pathway. I further explore the utility of the P-element invasion as an exemplar of the evolution of piRNA-mediated silencing. In light of the highly-conserved role for piRNAs in regulating TEs, discoveries from this system have taxonomically broad implications for the evolution of repression. PMID:27516614

  4. Microbial Disruption of Autophagy Alters Expression of the RISC Component AGO2, a Critical Regulator of the miRNA Silencing Pathway.

    Science.gov (United States)

    Sibony, Michal; Abdullah, Majd; Greenfield, Laura; Raju, Deepa; Wu, Ted; Rodrigues, David M; Galindo-Mata, Esther; Mascarenhas, Heidi; Philpott, Dana J; Silverberg, Mark S; Jones, Nicola L

    2015-12-01

    Autophagy is implicated in Crohn's disease (CD) pathogenesis. Recent evidence suggests autophagy regulates the microRNA (miRNA)-induced silencing complex (miRISC). Therefore, autophagy may play a novel role in CD by regulating expression of miRISC, thereby altering miRNA silencing. As microbes associated with CD can alter autophagy, we hypothesized that microbial disruption of autophagy affects the critical miRISC component AGO2. AGO2 expression was assessed in epithelial and immune cells, and intestinal organoids with disrupted autophagy. Microarray technology was used to determine the expression of downstream miRNAs in cells with defective autophagy. Increased AGO2 was detected in autophagy-deficient ATG5-/- and ATG16-/- mouse embryonic fibroblast cells (MEFs) in comparison with wild-type MEFs. Chemical agents and VacA toxin, which disrupt autophagy, increased AGO2 expression in MEFs, epithelial cells lines, and human monocytes, respectively. Increased AGO2 was also detected in ATG7-/- intestinal organoids, in comparison with wild-type organoids. Five miRNAs were differentially expressed in autophagy-deficient MEFs. Pathway enrichment analysis of the differentially expressed miRNAs implicated signaling pathways previously associated with CD. Taken together, our results suggest that autophagy is involved in the regulation of the critical miRISC component AGO2 in epithelial and immune cells and primary intestinal epithelial cells. We propose a mechanism by which autophagy alters miRNA expression, which likely impacts the regulation of CD-associated pathways. Furthermore, as enteric microbial products can manipulate autophagy and AGO2, our findings suggest a novel mechanism by which enteric microbes could influence miRNA to promote disease.

  5. APC/C-mediated degradation of dsRNA-binding protein 4 (DRB4 involved in RNA silencing.

    Directory of Open Access Journals (Sweden)

    Katia Marrocco

    Full Text Available Selective protein degradation via the ubiquitin-26S proteasome is a major mechanism underlying DNA replication and cell division in all Eukaryotes. In particular, the APC/C (Anaphase Promoting Complex or Cyclosome is a master ubiquitin protein ligase (E3 that targets regulatory proteins for degradation allowing sister chromatid separation and exit from mitosis. Interestingly, recent work also indicates that the APC/C remains active in differentiated animal and plant cells. However, its role in post-mitotic cells remains elusive and only a few substrates have been characterized.In order to identify novel APC/C substrates, we performed a yeast two-hybrid screen using as the bait Arabidopsis APC10/DOC1, one core subunit of the APC/C, which is required for substrate recruitment. This screen identified DRB4, a double-stranded RNA binding protein involved in the biogenesis of different classes of small RNA (sRNA. This protein interaction was further confirmed in vitro and in plant cells. Moreover, APC10 interacts with DRB4 through the second dsRNA binding motif (dsRBD2 of DRB4, which is also required for its homodimerization and binding to its Dicer partner DCL4. We further showed that DRB4 protein accumulates when the proteasome is inactivated and, most importantly, we found that DRB4 stability depends on APC/C activity. Hence, depletion of Arabidopsis APC/C activity by RNAi leads to a strong accumulation of endogenous DRB4, far beyond its normal level of accumulation. However, we could not detect any defects in sRNA production in lines where DRB4 was overexpressed.Our work identified a first plant substrate of the APC/C, which is not a regulator of the cell cycle. Though we cannot exclude that APC/C-dependent degradation of DRB4 has some regulatory roles under specific growth conditions, our work rather points to a housekeeping function of APC/C in maintaining precise cellular-protein concentrations and homeostasis of DRB4.

  6. AAV-based shRNA silencing of NF-κB ameliorates muscle pathologies in mdx mice.

    Science.gov (United States)

    Yang, Q; Tang, Y; Imbrogno, K; Lu, A; Proto, J D; Chen, A; Guo, F; Fu, F H; Huard, J; Wang, B

    2012-12-01

    Chronic inflammation, promoted by an upregulated NF-kappa B (NF-κB) pathway, has a key role in Duchenne muscular dystrophy (DMD) patients' pathogenesis. Blocking the NF-κB pathway has been shown to be a viable approach to diminish chronic inflammation and necrosis in the dystrophin-defective mdx mouse, a murine DMD model. In this study, we used the recombinant adeno-associated virus serotype 9 (AAV9) carrying an short hairpin RNA (shRNA) specifically targeting the messenger RNA of NF-κB/p65 (p65-shRNA), the major subunit of NF-κB associated with chronic inflammation in mdx mice. We examined whether i.m. AAV9-mediated delivery of p65-shRNA could decrease NF-κB activation, allowing for amelioration of muscle pathologies in 1- and 4-month-old mdx mice. At 1 month after treatment, NF-κB/p65 levels were significantly decreased by AAV gene transfer of p65-shRNA in the two ages of treatment groups, with necrosis significantly decreased compared with controls. Quantitative analysis revealed that central nucleation (CN) of the myofibers of p65-shRNA-treated 1-month-old mdx muscles was reduced from 67 to 34%, but the level of CN was not significantly decreased in treated 4-month-old mdx mice. Moreover, delivery of the p65-shRNA enhanced the capacity of myofiber regeneration in old mdx mice treated at 4 months of age when the dystrophic myofibers were most exhausted; however, such p65 silencing diminished the myofiber regeneration in young mdx mice treated at 1 month of age. Taken together, these findings demonstrate that the AAV-mediated delivery of p65-shRNA has the capacity to ameliorate muscle pathologies in mdx mice by selectively reducing NF-κB/p65 activity.

  7. Downregulation of cinnamyl-alcohol dehydrogenase in switchgrass by RNA silencing results in enhanced glucose release after cellulase treatment.

    Directory of Open Access Journals (Sweden)

    Aaron J Saathoff

    Full Text Available Cinnamyl alcohol dehydrogenase (CAD catalyzes the last step in monolignol biosynthesis and genetic evidence indicates CAD deficiency in grasses both decreases overall lignin, alters lignin structure and increases enzymatic recovery of sugars. To ascertain the effect of CAD downregulation in switchgrass, RNA mediated silencing of CAD was induced through Agrobacterium mediated transformation of cv. "Alamo" with an inverted repeat construct containing a fragment derived from the coding sequence of PviCAD2. The resulting primary transformants accumulated less CAD RNA transcript and protein than control transformants and were demonstrated to be stably transformed with between 1 and 5 copies of the T-DNA. CAD activity against coniferaldehyde, and sinapaldehyde in stems of silenced lines was significantly reduced as was overall lignin and cutin. Glucose release from ground samples pretreated with ammonium hydroxide and digested with cellulases was greater than in control transformants. When stained with the lignin and cutin specific stain phloroglucinol-HCl the staining intensity of one line indicated greater incorporation of hydroxycinnamyl aldehydes in the lignin.

  8. Downregulation of cinnamyl-alcohol dehydrogenase in switchgrass by RNA silencing results in enhanced glucose release after cellulase treatment.

    Science.gov (United States)

    Saathoff, Aaron J; Sarath, Gautam; Chow, Elaine K; Dien, Bruce S; Tobias, Christian M

    2011-01-27

    Cinnamyl alcohol dehydrogenase (CAD) catalyzes the last step in monolignol biosynthesis and genetic evidence indicates CAD deficiency in grasses both decreases overall lignin, alters lignin structure and increases enzymatic recovery of sugars. To ascertain the effect of CAD downregulation in switchgrass, RNA mediated silencing of CAD was induced through Agrobacterium mediated transformation of cv. "Alamo" with an inverted repeat construct containing a fragment derived from the coding sequence of PviCAD2. The resulting primary transformants accumulated less CAD RNA transcript and protein than control transformants and were demonstrated to be stably transformed with between 1 and 5 copies of the T-DNA. CAD activity against coniferaldehyde, and sinapaldehyde in stems of silenced lines was significantly reduced as was overall lignin and cutin. Glucose release from ground samples pretreated with ammonium hydroxide and digested with cellulases was greater than in control transformants. When stained with the lignin and cutin specific stain phloroglucinol-HCl the staining intensity of one line indicated greater incorporation of hydroxycinnamyl aldehydes in the lignin.

  9. GW182-Free microRNA Silencing Complex Controls Post-transcriptional Gene Expression during Caenorhabditis elegans Embryogenesis.

    Directory of Open Access Journals (Sweden)

    Guillaume Jannot

    2016-12-01

    Full Text Available MicroRNAs and Argonaute form the microRNA induced silencing complex or miRISC that recruits GW182, causing mRNA degradation and/or translational repression. Despite the clear conservation and molecular significance, it is unknown if miRISC-GW182 interaction is essential for gene silencing during animal development. Using Caenorhabditis elegans to explore this question, we examined the relationship and effect on gene silencing between the GW182 orthologs, AIN-1 and AIN-2, and the microRNA-specific Argonaute, ALG-1. Homology modeling based on human Argonaute structures indicated that ALG-1 possesses conserved Tryptophan-binding Pockets required for GW182 binding. We show in vitro and in vivo that their mutations severely altered the association with AIN-1 and AIN-2. ALG-1 tryptophan-binding pockets mutant animals retained microRNA-binding and processing ability, but were deficient in reporter silencing activity. Interestingly, the ALG-1 tryptophan-binding pockets mutant phenocopied the loss of alg-1 in worms during larval stages, yet was sufficient to rescue embryonic lethality, indicating the dispensability of AINs association with the miRISC at this developmental stage. The dispensability of AINs in miRNA regulation is further demonstrated by the capacity of ALG-1 tryptophan-binding pockets mutant to regulate a target of the embryonic mir-35 microRNA family. Thus, our results demonstrate that the microRNA pathway can act independently of GW182 proteins during C. elegans embryogenesis.

  10. Impact of target mRNA structure on siRNA silencing efficiency: A large-scale study.

    Science.gov (United States)

    Gredell, Joseph A; Berger, Angela K; Walton, S Patrick

    2008-07-01

    The selection of active siRNAs is generally based on identifying siRNAs with certain sequence and structural properties. However, the efficiency of RNA interference has also been shown to depend on the structure of the target mRNA, primarily through studies using exogenous transcripts with well-defined secondary structures in the vicinity of the target sequence. While these studies provide a means for examining the impact of target sequence and structure independently, the predicted secondary structures for these transcripts are often not reflective of structures that form in full-length, native mRNAs where interactions can occur between relatively remote segments of the mRNAs. Here, using a combination of experimental results and analysis of a large dataset, we demonstrate that the accessibility of certain local target structures on the mRNA is an important determinant in the gene silencing ability of siRNAs. siRNAs targeting the enhanced green fluorescent protein were chosen using a minimal siRNA selection algorithm followed by classification based on the predicted minimum free energy structures of the target transcripts. Transfection into HeLa and HepG2 cells revealed that siRNAs targeting regions of the mRNA predicted to have unpaired 5'- and 3'-ends resulted in greater gene silencing than regions predicted to have other types of secondary structure. These results were confirmed by analysis of gene silencing data from previously published siRNAs, which showed that mRNA target regions unpaired at either the 5'-end or 3'-end were silenced, on average, approximately 10% more strongly than target regions unpaired in the center or primarily paired throughout. We found this effect to be independent of the structure of the siRNA guide strand. Taken together, these results suggest minimal requirements for nucleation of hybridization between the siRNA guide strand and mRNA and that both mRNA and guide strand structure should be considered when choosing candidate si

  11. Demonstration of helicase activity in the nonstructural protein, NSs, of the negative-sense RNA virus, groundnut bud necrosis virus.

    Science.gov (United States)

    Bhushan, Lokesh; Abraham, Ambily; Choudhury, Nirupam Roy; Rana, Vipin Singh; Mukherjee, Sunil Kumar; Savithri, Handanahal Subbarao

    2015-04-01

    The nonstructural protein NSs, encoded by the S RNA of groundnut bud necrosis virus (GBNV) (genus Tospovirus, family Bunyaviridae) has earlier been shown to possess nucleic-acid-stimulated NTPase and 5' α phosphatase activity. ATP hydrolysis is an essential function of a true helicase. Therefore, NSs was tested for DNA helicase activity. The results demonstrated that GBNV NSs possesses bidirectional DNA helicase activity. An alanine mutation in the Walker A motif (K189A rNSs) decreased DNA helicase activity substantially, whereas a mutation in the Walker B motif resulted in a marginal decrease in this activity. The parallel loss of the helicase and ATPase activity in the K189A mutant confirms that NSs acts as a non-canonical DNA helicase. Furthermore, both the wild-type and K189A NSs could function as RNA silencing suppressors, demonstrating that the suppressor activity of NSs is independent of its helicase or ATPase activity. This is the first report of a true helicase from a negative-sense RNA virus.

  12. Viral RNAi suppressor reversibly binds siRNA to outcompete Dicer and RISC via multiple-turnover

    Science.gov (United States)

    Rawlings, Renata A.; Krishnan, Vishalakshi; Walter, Nils G.

    2011-01-01

    RNA interference (RNAi) is a conserved gene regulatory mechanism employed by most eukaryotes as a key component of their innate immune response against viruses and retrotransposons. During viral infection, the RNase III-type endonuclease Dicer cleaves viral double-stranded RNA into small interfering RNAs (siRNAs), 21–24 nucleotides in length, and helps load them into the RNA-induced silencing complex (RISC) to guide cleavage of complementary viral RNA. As a countermeasure, many viruses have evolved viral RNA silencing suppressor (RSS) proteins that tightly, and presumably quantitatively, bind siRNAs to thwart RNAi-mediated degradation. Viral RSS proteins also act across kingdoms as potential immunosuppressors in gene therapeutic applications. Here we report fluorescence quenching and electrophoretic mobility shift assays that probe siRNA binding by the dimeric RSS p19 from Carnation Italian Ringspot Virus (CIRV), as well as by human Dicer and RISC assembly complexes. We find that the siRNA:p19 interaction is readily reversible, characterized by rapid binding ((1.69 ± 0.07)×108 M−1s−1) and marked dissociation (koff = 0.062 ± 0.002 s−1). We also observe that p19 efficiently competes with recombinant Dicer and inhibits formation of RISC-related assembly complexes found in human cell extract. Computational modeling based on these results provides evidence for the transient formation of a ternary complex between siRNA, human Dicer, and p19. An expanded model of RNA silencing indicates that multiple-turnover by reversible binding of siRNAs potentiates the efficiency of the suppressor protein. Our predictive model is expected to be applicable to the dosing of p19 as a silencing suppressor in viral gene therapy. PMID:21354178

  13. Complexes between the LKB1 tumor suppressor, STRADα/β and MO25α/β are upstream kinases in the AMP-activated protein kinase cascade

    Directory of Open Access Journals (Sweden)

    Alessi Dario R

    2003-09-01

    Full Text Available Abstract Background The AMP-activated protein kinase (AMPK cascade is a sensor of cellular energy charge that acts as a 'metabolic master switch' and inhibits cell proliferation. Activation requires phosphorylation of Thr172 of AMPK within the activation loop by upstream kinases (AMPKKs that have not been identified. Recently, we identified three related protein kinases acting upstream of the yeast homolog of AMPK. Although they do not have obvious mammalian homologs, they are related to LKB1, a tumor suppressor that is mutated in the human Peutz-Jeghers cancer syndrome. We recently showed that LKB1 exists as a complex with two accessory subunits, STRADα/β and MO25α/β. Results We report the following observations. First, two AMPKK activities purified from rat liver contain LKB1, STRADα and MO25α, and can be immunoprecipitated using anti-LKB1 antibodies. Second, both endogenous and recombinant complexes of LKB1, STRADα/β and MO25α/β activate AMPK via phosphorylation of Thr172. Third, catalytically active LKB1, STRADα or STRADβ and MO25α or MO25β are required for full activity. Fourth, the AMPK-activating drugs AICA riboside and phenformin do not activate AMPK in HeLa cells (which lack LKB1, but activation can be restored by stably expressing wild-type, but not catalytically inactive, LKB1. Fifth, AICA riboside and phenformin fail to activate AMPK in immortalized fibroblasts from LKB1-knockout mouse embryos. Conclusions These results provide the first description of a physiological substrate for the LKB1 tumor suppressor and suggest that it functions as an upstream regulator of AMPK. Our findings indicate that the tumors in Peutz-Jeghers syndrome could result from deficient activation of AMPK as a consequence of LKB1 inactivation.

  14. Myeloid derived suppressor cells (MDSCs are increased and exert immunosuppressive activity together with polymorphonuclear leukocytes (PMNs in chronic myeloid leukemia patients.

    Directory of Open Access Journals (Sweden)

    Cesarina Giallongo

    Full Text Available Tumor immune tolerance can derive from the recruitment of suppressor cell population, including myeloid derived suppressor cells (MDSCs, able to inhibit T cells activity. We identified a significantly expanded MDSCs population in chronic myeloid leukemia (CML patients at diagnosis that decreased to normal levels after imatinib therapy. In addition, expression of arginase 1 (Arg1 that depletes microenvironment of arginine, an essential aminoacid for T cell function, resulted in an increase in patients at diagnosis. Purified CML CD11b+CD33+CD14-HLADR- cells markedly suppressed normal donor T cell proliferation in vitro. Comparing CML Gr-MDSCs to autologous polymorphonuclear leukocytes (PMNs we observed a higher Arg1 expression and activity in PMNs, together with an inhibitory effect on T cells in vitro. Our data indicate that CML cells create an immuno-tolerant environment associated to MDSCs expansion with immunosuppressive capacity mediated by Arg1. In addition, we demonstrated for the first time also an immunosuppressive activity of CML PMNs, suggesting a strong potential immune escape mechanism created by CML cells, which control the anti-tumor reactive T cells. MDSCs should be monitored in imatinib discontinuation trials to understand their importance in relapsing patients.

  15. Retinoid-induced expression and activity of an immediate early tumor suppressor gene in vascular smooth muscle cells.

    Directory of Open Access Journals (Sweden)

    Jeffrey W Streb

    2011-04-01

    Full Text Available Retinoids are used clinically to treat a number of hyper-proliferative disorders and have been shown in experimental animals to attenuate vascular occlusive diseases, presumably through nuclear receptors bound to retinoic acid response elements (RARE located in target genes. Here, we show that natural or synthetic retinoids rapidly induce mRNA and protein expression of a specific isoform of A-Kinase Anchoring Protein 12 (AKAP12β in cultured smooth muscle cells (SMC as well as the intact vessel wall. Expression kinetics and actinomycin D studies indicate Akap12β is a retinoid-induced, immediate-early gene. Akap12β promoter analyses reveal a conserved RARE mildly induced with atRA in a region that exhibits hyper-acetylation. Immunofluorescence microscopy and protein kinase A (PKA regulatory subunit overlay assays in SMC suggest a physical association between AKAP12β and PKA following retinoid treatment. Consistent with its designation as a tumor suppressor, inducible expression of AKAP12β attenuates SMC growth in vitro. Further, immunohistochemistry studies establish marked decreases in AKAP12 expression in experimentally-injured vessels of mice as well as atheromatous lesions in humans. Collectively, these results demonstrate a novel role for retinoids in the induction of an AKAP tumor suppressor that blocks vascular SMC growth thus providing new molecular insight into how retiniods may exert their anti-proliferative effects in the injured vessel wall.

  16. Active suppression of host-vs-graft reaction in pregnant mice. VII. Spontaneous abortion of allogeneic CBA/J x DBA/2 fetuses in the uterus of CBA/J mice correlates with deficient non-T suppressor cell activity

    International Nuclear Information System (INIS)

    Clark, D.A.; Chaput, A.; Tutton, D.

    1986-01-01

    The mammalian fetus has been viewed as an unusually successful type of allograft and unexplained spontaneous abortion as a possible example of maternal rejection. Previous studies have shown the presence of small lymphocytic suppressor cells in the murine decidua which block the generation and reactivation of anti-paternal cytotoxic T lymphocytes (CTL) and lymphokine-activated killer cells (LAK) by elaborating a factor that inhibits the response to interleukin 2 (IL 2). A deficiency of these suppressor cells was associated with implants of xenogeneic Mus caroli embryos in the Mus musculus uterus which are infiltrated by maternal lymphoid cells and aborted. A deficiency of such suppressor cells in the lymph nodes draining the uterus of CBA/J females in the process of aborting their semi-allogeneic CBA x DBA/2 F 1 progeny has also been shown. CBA/J females possess significantly lower levels of decidua-associated non-T suppressor cells on day 8.5 to 10.5 of allopregnancy than do mothers that will produce large litters of live babies. The F 1 embryos are infiltrated by maternal lymphocytes prior to abortion, and the infiltration and abortion rate appears to be augmented by pre-immunization with paternal DBA/2 spleen cells. The CBA/J x DBA/2J mating combination provides a model of spontaneous abortion in which immunologic factors play an important role and demonstrates that the association between deficiency of decidua-associated suppressor cells and xenopregnancy failure also holds true for the failure of allopregnancies resulting from natural within-species mating

  17. Induction of suppressor cells in vitro by Candida albicans.

    Science.gov (United States)

    Cuff, C F; Rogers, C M; Lamb, B J; Rogers, T J

    1986-06-01

    Normal splenocytes cultured with Formalin-killed Candida albicans were shown to acquire significant suppressor cell activity in a period of 3 days. These cells were found to suppress both the phytohemagglutinin-induced mitogen response as well as the anti-sheep erythrocyte antibody response. Experiments were carried out to determine the nature of the suppressor cell population. Results showed that these cells were not susceptible to treatment with anti-Thy 1 antibody and complement. Panning experiments showed that the suppressor cells were not plastic-adherent or Mac-1 antigen-positive. The suppressor cells were, however, adherent to anti-mouse immunoglobulin (F(ab')2-fragment)-coated dishes. Additional experiments showed that the suppressor cell activity was susceptible to treatment with monoclonal anti-Lyb 2.1 antibody and complement. These results suggest that the suppressor cell induced in vitro by Candida is a member of the B-lymphocyte lineage.

  18. Long term effect of curcumin in restoration of tumour suppressor p53 and phase-II antioxidant enzymes via activation of Nrf2 signalling and modulation of inflammation in prevention of cancer.

    Directory of Open Access Journals (Sweden)

    Laxmidhar Das

    Full Text Available Inhibition of carcinogenesis may be a consequence of attenuation of oxidative stress via activation of antioxidant defence system, restoration and stabilization of tumour suppressor proteins along with modulation of inflammatory mediators. Previously we have delineated significant role of curcumin during its long term effect in regulation of glycolytic pathway and angiogenesis, which in turn results in prevention of cancer via modulation of stress activated genes. Present study was designed to investigate long term effect of curcumin in regulation of Nrf2 mediated phase-II antioxidant enzymes, tumour suppressor p53 and inflammation under oxidative tumour microenvironment in liver of T-cell lymphoma bearing mice. Inhibition of Nrf2 signalling observed during lymphoma progression, resulted in down regulation of phase II antioxidant enzymes, p53 as well as activation of inflammatory signals. Curcumin potentiated significant increase in Nrf2 activation. It restored activity of phase-II antioxidant enzymes like GST, GR, NQO1, and tumour suppressor p53 level. In addition, curcumin modulated inflammation via upregulation of TGF-β and reciprocal regulation of iNOS and COX2. The study suggests that during long term effect, curcumin leads to prevention of cancer by inducing phase-II antioxidant enzymes via activation of Nrf2 signalling, restoration of tumour suppressor p53 and modulation of inflammatory mediators like iNOS and COX2 in liver of lymphoma bearing mice.

  19. Lethal giant larvae 1 tumour suppressor activity is not conserved in models of mammalian T and B cell leukaemia.

    Directory of Open Access Journals (Sweden)

    Edwin D Hawkins

    Full Text Available In epithelial and stem cells, lethal giant larvae (Lgl is a potent tumour suppressor, a regulator of Notch signalling, and a mediator of cell fate via asymmetric cell division. Recent evidence suggests that the function of Lgl is conserved in mammalian haematopoietic stem cells and implies a contribution to haematological malignancies. To date, direct measurement of the effect of Lgl expression on malignancies of the haematopoietic lineage has not been tested. In Lgl1⁻/⁻ mice, we analysed the development of haematopoietic malignancies either alone, or in the presence of common oncogenic lesions. We show that in the absence of Lgl1, production of mature white blood cell lineages and long-term survival of mice are not affected. Additionally, loss of Lgl1 does not alter leukaemia driven by constitutive Notch, c-Myc or Jak2 signalling. These results suggest that the role of Lgl1 in the haematopoietic lineage might be restricted to specific co-operating mutations and a limited number of cellular contexts.

  20. Lethal Giant Larvae 1 Tumour Suppressor Activity Is Not Conserved in Models of Mammalian T and B Cell Leukaemia

    Science.gov (United States)

    Hawkins, Edwin D.; Oliaro, Jane; Ramsbottom, Kelly M.; Ting, Stephen B.; Sacirbegovic, Faruk; Harvey, Michael; Kinwell, Tanja; Ghysdael, Jacques; Johnstone, Ricky W.; Humbert, Patrick O.; Russell, Sarah M.

    2014-01-01

    In epithelial and stem cells, lethal giant larvae (Lgl) is a potent tumour suppressor, a regulator of Notch signalling, and a mediator of cell fate via asymmetric cell division. Recent evidence suggests that the function of Lgl is conserved in mammalian haematopoietic stem cells and implies a contribution to haematological malignancies. To date, direct measurement of the effect of Lgl expression on malignancies of the haematopoietic lineage has not been tested. In Lgl1−/− mice, we analysed the development of haematopoietic malignancies either alone, or in the presence of common oncogenic lesions. We show that in the absence of Lgl1, production of mature white blood cell lineages and long-term survival of mice are not affected. Additionally, loss of Lgl1 does not alter leukaemia driven by constitutive Notch, c-Myc or Jak2 signalling. These results suggest that the role of Lgl1 in the haematopoietic lineage might be restricted to specific co-operating mutations and a limited number of cellular contexts. PMID:24475281

  1. Analysis of the Mitogen-activated protein kinase kinase 4 (MAP2K4) tumor suppressor gene in ovarian cancer

    International Nuclear Information System (INIS)

    Davis, Sally J; Choong, David YH; Ramakrishna, Manasa; Ryland, Georgina L; Campbell, Ian G; Gorringe, Kylie L

    2011-01-01

    MAP2K4 is a putative tumor and metastasis suppressor gene frequently found to be deleted in various cancer types. We aimed to conduct a comprehensive analysis of this gene to assess its involvement in ovarian cancer. We screened for mutations in MAP2K4 using High Resolution Melt analysis of 149 primary ovarian tumors and methylation at the promoter using Methylation-Specific Single-Stranded Conformation Polymorphism analysis of 39 tumors. We also considered the clinical impact of changes in MAP2K4 using publicly available expression and copy number array data. Finally, we used siRNA to measure the effect of reducing MAP2K4 expression in cell lines. In addition to 4 previously detected homozygous deletions, we identified a homozygous 16 bp truncating deletion and a heterozygous 4 bp deletion, each in one ovarian tumor. No promoter methylation was detected. The frequency of MAP2K4 homozygous inactivation was 5.6% overall, and 9.8% in high-grade serous cases. Hemizygous deletion of MAP2K4 was observed in 38% of samples. There were significant correlations of copy number and expression in three microarray data sets. There was a significant correlation between MAP2K4 expression and overall survival in one expression array data set, but this was not confirmed in an independent set. Treatment of JAM and HOSE6.3 cell lines with MAP2K4 siRNA showed some reduction in proliferation. MAP2K4 is targeted by genetic inactivation in ovarian cancer and restricted to high grade serous and endometrioid carcinomas in our cohort

  2. ATP and MO25alpha regulate the conformational state of the STRADalpha pseudokinase and activation of the LKB1 tumour suppressor.

    Directory of Open Access Journals (Sweden)

    Elton Zeqiraj

    2009-06-01

    Full Text Available Pseudokinases lack essential residues for kinase activity, yet are emerging as important regulators of signal transduction networks. The pseudokinase STRAD activates the LKB1 tumour suppressor by forming a heterotrimeric complex with LKB1 and the scaffolding protein MO25. Here, we describe the structure of STRADalpha in complex with MO25alpha. The structure reveals an intricate web of interactions between STRADalpha and MO25alpha involving the alphaC-helix of STRADalpha, reminiscent of the mechanism by which CDK2 interacts with cyclin A. Surprisingly, STRADalpha binds ATP and displays a closed conformation and an ordered activation loop, typical of active protein kinases. Inactivity is accounted for by nonconservative substitution of almost all essential catalytic residues. We demonstrate that binding of ATP enhances the affinity of STRADalpha for MO25alpha, and conversely, binding of MO25alpha promotes interaction of STRADalpha with ATP. Mutagenesis studies reveal that association of STRADalpha with either ATP or MO25alpha is essential for LKB1 activation. We conclude that ATP and MO25alpha cooperate to maintain STRADalpha in an "active" closed conformation required for LKB1 activation. It has recently been demonstrated that a mutation in human STRADalpha that truncates a C-terminal region of the pseudokinase domain leads to the polyhydramnios, megalencephaly, symptomatic epilepsy (PMSE syndrome. We demonstrate this mutation destabilizes STRADalpha and prevents association with LKB1. In summary, our findings describe one of the first structures of a genuinely inactive pseudokinase. The ability of STRADalpha to activate LKB1 is dependent on a closed "active" conformation, aided by ATP and MO25alpha binding. Thus, the function of STRADalpha is mediated through an active kinase conformation rather than kinase activity. It is possible that other pseudokinases exert their function through nucleotide binding and active conformations.

  3. Genistein inhibits proliferation of colon cancer cells by attenuating a negative effect of epidermal growth factor on tumor suppressor FOXO3 activity

    International Nuclear Information System (INIS)

    Qi, Wentao; Weber, Christopher R; Wasland, Kaarin; Savkovic, Suzana D

    2011-01-01

    Soy consumption is associated with a lower incidence of colon cancer which is believed to be mediated by one of its of components, genistein. Genistein may inhibit cancer progression by inducing apoptosis or inhibiting proliferation, but mechanisms are not well understood. Epidermal growth factor (EGF)-induced proliferation of colon cancer cells plays an important role in colon cancer progression and is mediated by loss of tumor suppressor FOXO3 activity. The aim of this study was to assess if genistein exerts anti-proliferative properties by attenuating the negative effect of EGF on FOXO3 activity. The effect of genistein on proliferation stimulated by EGF-mediated loss of FOXO3 was examined in human colonic cancer HT-29 cells. EGF-induced FOXO3 phosphorylation and translocation were assessed in the presence of genistein. EGF-mediated loss of FOXO3 interactions with p53 (co-immunoprecipitation) and promoter of p27kip1 (ChIP assay) were examined in presence of genistein in cells with mutated p53 (HT-29) and wild type p53 (HCT116). Silencing of p53 determined activity of FOXO3 when it is bound to p53. Genistein inhibited EGF-induced proliferation, while favoring dephosphorylation and nuclear retention of FOXO3 (active state) in colon cancer cells. Upstream of FOXO3, genistein acts via the PI3K/Akt pathway to inhibit EGF-stimulated FOXO3 phosphorylation (i.e. favors active state). Downstream, EGF-induced disassociation of FOXO3 from mutated tumor suppressor p53, but not wild type p53, is inhibited by genistein favoring FOXO3-p53(mut) interactions with the promoter of the cell cycle inhibitor p27kip1 in colon cancer cells. Thus, the FOXO3-p53(mut) complex leads to elevated p27kip1 expression and promotes cell cycle arrest. These novel anti-proliferative mechanisms of genistein suggest a possible role of combining genistein with other chemoreceptive agents for the treatment of colon cancer

  4. Tumor suppressor PTEN affects tau phosphorylation: deficiency in the phosphatase activity of PTEN increases aggregation of an FTDP-17 mutant Tau

    Directory of Open Access Journals (Sweden)

    Zhang Xue

    2006-07-01

    Full Text Available Abstract Background Aberrant hyperphosphorylation of tau protein has been implicated in a variety of neurodegenerative disorders. Although a number of protein kinases have been shown to phosphorylate tau in vitro and in vivo, the molecular mechanisms by which tau phosphorylation is regulated pathophysiologically are largely unknown. Recently, a growing body of evidence suggests a link between tau phosphorylation and PI3K signaling. In this study, phosphorylation, aggregation and binding to the microtubule of a mutant frontal temporal dementia and parkinsonism linked to chromosome 17 (FTDP-17 tau in the presence of tumor suppressor PTEN, a major regulatory component in PI3K signaling, were investigated. Results Phosphorylation of the human mutant FTDP-17 tau, T40RW, was evaluated using different phospho-tau specific antibodies in the presence of human wild-type or phosphatase activity null mutant PTEN. Among the evaluated phosphorylation sites, the levels of Ser214 and Thr212 phospho-tau proteins were significantly decreased in the presence of wild-type PTEN, and significantly increased when the phosphatase activity null mutant PTEN was ectopically expressed. Fractionation of the mutant tau transfected cells revealed a significantly increased level of soluble tau in cytosol when wild-type PTEN was expressed, and an elevated level of SDS-soluble tau aggregates in the presence of the mutant PTEN. In addition, the filter/trap assays detected more SDS-insoluble mutant tau aggregates in the cells overexpressing the mutant PTEN compared to those in the cells overexpressing wild-type PTEN and control DNA. This notion was confirmed by the immunocytochemical experiment which demonstrated that the overexpression of the phosphatase activity null mutant PTEN caused the mutant tau to form aggregates in the COS-7 cells. Conclusion Tumor suppressor PTEN can alleviate the phosporylation of the mutant FTDP-17 tau at specific sites, and the phosphatase activity

  5. Mesenchymal Stem Cells (MSC Regulate Activation of Granulocyte-Like Myeloid Derived Suppressor Cells (G-MDSC in Chronic Myeloid Leukemia Patients.

    Directory of Open Access Journals (Sweden)

    Cesarina Giallongo

    Full Text Available It is well known that mesenchymal stem cells (MSC have a role in promotion of tumor growth, survival and drug-resistance in chronic myeloid leukemia (CML. Recent reports indicated that a subpopulation of myeloid cells, defined as granulocyte-like myeloid-derived suppressor cells (G-MDSC is increased in these patients. So far, the role of MSC in MDSC expansion and activation into the BM microenvironment remains unexplored. To address this question, here we use a specific experimental model in vitro, co-culturing MSC with peripheral blood mononucleated cells (PBMC from normal individuals, in order to generate MSC-educated G-MDSC. Although MSC of healthy donors (HD and CML patients were able to generate the same amount of MDSC, only CML-MSC-educated G-MDSC exhibited suppressive ability on autologous T lymphocytes. In addition, compared with HD-MSC, CML-MSC over-expressed some immunomodulatory factors including TGFβ, IL6 and IL10, that could be involved in MDSC activation. CML-MSC-educated G-MDSC expressed higher levels of ARG1, TNFα, IL1β, COX2 and IL6 than G-MDSC isolated from co-culture with HD-MSC. Our data provide evidence that CML-MSC may play a critical role in tumor microenvironment by orchestrating G-MDSC activation and regulating T lymphocytes-mediated leukemia surveillance, thus contributing to CML immune escape.

  6. Mesenchymal Stem Cells (MSC) Regulate Activation of Granulocyte-Like Myeloid Derived Suppressor Cells (G-MDSC) in Chronic Myeloid Leukemia Patients.

    Science.gov (United States)

    Giallongo, Cesarina; Romano, Alessandra; Parrinello, Nunziatina Laura; La Cava, Piera; Brundo, Maria Violetta; Bramanti, Vincenzo; Stagno, Fabio; Vigneri, Paolo; Chiarenza, Annalisa; Palumbo, Giuseppe Alberto; Tibullo, Daniele; Di Raimondo, Francesco

    2016-01-01

    It is well known that mesenchymal stem cells (MSC) have a role in promotion of tumor growth, survival and drug-resistance in chronic myeloid leukemia (CML). Recent reports indicated that a subpopulation of myeloid cells, defined as granulocyte-like myeloid-derived suppressor cells (G-MDSC) is increased in these patients. So far, the role of MSC in MDSC expansion and activation into the BM microenvironment remains unexplored. To address this question, here we use a specific experimental model in vitro, co-culturing MSC with peripheral blood mononucleated cells (PBMC) from normal individuals, in order to generate MSC-educated G-MDSC. Although MSC of healthy donors (HD) and CML patients were able to generate the same amount of MDSC, only CML-MSC-educated G-MDSC exhibited suppressive ability on autologous T lymphocytes. In addition, compared with HD-MSC, CML-MSC over-expressed some immunomodulatory factors including TGFβ, IL6 and IL10, that could be involved in MDSC activation. CML-MSC-educated G-MDSC expressed higher levels of ARG1, TNFα, IL1β, COX2 and IL6 than G-MDSC isolated from co-culture with HD-MSC. Our data provide evidence that CML-MSC may play a critical role in tumor microenvironment by orchestrating G-MDSC activation and regulating T lymphocytes-mediated leukemia surveillance, thus contributing to CML immune escape.

  7. Measurand transient signal suppressor

    Science.gov (United States)

    Bozeman, Richard J., Jr. (Inventor)

    1994-01-01

    A transient signal suppressor for use in a controls system which is adapted to respond to a change in a physical parameter whenever it crosses a predetermined threshold value in a selected direction of increasing or decreasing values with respect to the threshold value and is sustained for a selected discrete time interval is presented. The suppressor includes a sensor transducer for sensing the physical parameter and generating an electrical input signal whenever the sensed physical parameter crosses the threshold level in the selected direction. A manually operated switch is provided for adapting the suppressor to produce an output drive signal whenever the physical parameter crosses the threshold value in the selected direction of increasing or decreasing values. A time delay circuit is selectively adjustable for suppressing the transducer input signal for a preselected one of a plurality of available discrete suppression time and producing an output signal only if the input signal is sustained for a time greater than the selected suppression time. An electronic gate is coupled to receive the transducer input signal and the timer output signal and produce an output drive signal for energizing a control relay whenever the transducer input is a non-transient signal which is sustained beyond the selected time interval.

  8. A single mutation in the 15S rRNA gene confers nonsense suppressor activity and interacts with mRF1 the release factor in yeast mitochondria

    Directory of Open Access Journals (Sweden)

    Ali Gargouri

    2015-08-01

    Full Text Available We have determined the nucleotide sequence of the mim3-1 mitochondrial ribosomal suppressor, acting on ochre mitochondrial mutations and one frameshift mutation in Saccharomyces cerevisiae. The 15s rRNA suppressor gene contains a G633 to C transversion. Yeast mitochondrial G633 corresponds to G517 of the E.coli 15S rRNA, which is occupied by an invariant G in all known small rRNA sequences. Interestingly, this mutation has occurred at the same position as the known MSU1 mitochondrial suppressor which changes G633 to A. The suppressor mutation lies in a highly conserved region of the rRNA, known in E.coli as the 530-loop, interacting with the S4, S5 and S12 ribosomal proteins. We also show an interesting interaction between the mitochondrial mim3-1 and the nuclear nam3-1 suppressors, both of which have the same action spectrum on mitochondrial mutations: nam3-1 abolishes the suppressor effect when present with mim3-1 in the same haploid cell. We discuss these results in the light of the nature of Nam3, identified by [1] as the yeast mitochondrial translation release factor. A hypothetical mechanism of suppression by "ribosome shifting" is also discussed in view of the nature of mutations suppressed and not suppressed.

  9. Transmembrane Tumor Necrosis Factor Controls Myeloid-Derived Suppressor Cell Activity via TNF Receptor 2 and Protects from Excessive Inflammation during BCG-Induced Pleurisy

    Directory of Open Access Journals (Sweden)

    Leslie Chavez-Galan

    2017-08-01

    Full Text Available Pleural tuberculosis (TB is a form of extra-pulmonary TB observed in patients infected with Mycobacterium tuberculosis. Accumulation of myeloid-derived suppressor cells (MDSC has been observed in animal models of TB and in human patients but their role remains to be fully elucidated. In this study, we analyzed the role of transmembrane TNF (tmTNF in the accumulation and function of MDSC in the pleural cavity during an acute mycobacterial infection. Mycobacterium bovis BCG-induced pleurisy was resolved in mice expressing tmTNF, but lethal in the absence of tumor necrosis factor. Pleural infection induced MDSC accumulation in the pleural cavity and functional MDSC required tmTNF to suppress T cells as did pleural wild-type MDSC. Interaction of MDSC expressing tmTNF with CD4 T cells bearing TNF receptor 2 (TNFR2, but not TNFR1, was required for MDSC suppressive activity on CD4 T cells. Expression of tmTNF attenuated Th1 cell-mediated inflammatory responses generated by the acute pleural mycobacterial infection in association with effective MDSC expressing tmTNF and interacting with CD4 T cells expressing TNFR2. In conclusion, this study provides new insights into the crucial role played by the tmTNF/TNFR2 pathway in MDSC suppressive activity required during acute pleural infection to attenuate excessive inflammation generated by the infection.

  10. Alkylation of the tumor suppressor PTEN activates Akt and β-catenin signaling: a mechanism linking inflammation and oxidative stress with cancer.

    Directory of Open Access Journals (Sweden)

    Tracy M Covey

    2010-10-01

    Full Text Available PTEN, a phosphoinositide-3-phosphatase, serves dual roles as a tumor suppressor and regulator of cellular anabolic/catabolic metabolism. Adaptation of a redox-sensitive cysteinyl thiol in PTEN for signal transduction by hydrogen peroxide may have superimposed a vulnerability to other mediators of oxidative stress and inflammation, especially reactive carbonyl species, which are commonly occurring by-products of arachidonic acid peroxidation. Using MCF7 and HEK-293 cells, we report that several reactive aldehydes and ketones, e.g. electrophilic α,β-enals (acrolein, 4-hydroxy-2-nonenal and α,β-enones (prostaglandin A(2, Δ12-prostaglandin J(2 and 15-deoxy-Δ-12,14-prostaglandin J(2 covalently modify and inactivate cellular PTEN, with ensuing activation of PKB/Akt kinase; phosphorylation of Akt substrates; increased cell proliferation; and increased nuclear β-catenin signaling. Alkylation of PTEN by α,β-enals/enones and interference with its restraint of cellular PKB/Akt signaling may accentuate hyperplastic and neoplastic disorders associated with chronic inflammation, oxidative stress, or aging.

  11. Genomic loss of tumor suppressor miRNA-204 promotes cancer cell migration and invasion by activating AKT/mTOR/Rac1 signaling and actin reorganization.

    Directory of Open Access Journals (Sweden)

    J Saadi Imam

    Full Text Available Increasing evidence suggests that chromosomal regions containing microRNAs are functionally important in cancers. Here, we show that genomic loci encoding miR-204 are frequently lost in multiple cancers, including ovarian cancers, pediatric renal tumors, and breast cancers. MiR-204 shows drastically reduced expression in several cancers and acts as a potent tumor suppressor, inhibiting tumor metastasis in vivo when systemically delivered. We demonstrated that miR-204 exerts its function by targeting genes involved in tumorigenesis including brain-derived neurotrophic factor (BDNF, a neurotrophin family member which is known to promote tumor angiogenesis and invasiveness. Analysis of primary tumors shows that increased expression of BDNF or its receptor tropomyosin-related kinase B (TrkB parallel a markedly reduced expression of miR-204. Our results reveal that loss of miR-204 results in BDNF overexpression and subsequent activation of the small GTPase Rac1 and actin reorganization through the AKT/mTOR signaling pathway leading to cancer cell migration and invasion. These results suggest that microdeletion of genomic loci containing miR-204 is directly linked with the deregulation of key oncogenic pathways that provide crucial stimulus for tumor growth and metastasis. Our findings provide a strong rationale for manipulating miR-204 levels therapeutically to suppress tumor metastasis.

  12. Up-regulation of tumor suppressor genes by exogenous dhC16-Cer contributes to its anti-cancer activity in primary effusion lymphoma.

    Science.gov (United States)

    Cao, Yueyu; Qiao, Jing; Lin, Zhen; Zabaleta, Jovanny; Dai, Lu; Qin, Zhiqiang

    2017-02-28

    Primary effusion lymphoma (PEL) is a rare and highly aggressive B-cell malignancy with Kaposi's sarcoma-associated herpesvirus (KSHV) infection, while lack of effective therapies. Our recent data indicated that targeting the sphingolipid metabolism by either sphingosine kinase inhibitor or exogenous ceramide species induces PEL cell apoptosis and suppresses tumor progression in vivo. However, the underlying mechanisms for these exogenous ceramides "killing" PEL cells remain largely unknown. Based on the microarray analysis, we found that exogenous dhC16-Cer treatment affected the expression of many cellular genes with important functions within PEL cells such as regulation of cell cycle, cell survival/proliferation, and apoptosis/anti-apoptosis. Interestingly, we found that a subset of tumor suppressor genes (TSGs) was up-regulated from dhC16-Cer treated PEL cells. One of these elevated TSGs, Thrombospondin-1 (THBS1) was required for dhC16-Cer induced PEL cell cycle arrest. Moreover, dhC16-Cer up-regulation of THBS1 was through the suppression of multiple KSHV microRNAs expression. Our data demonstrate that exogenous ceramides display anti-cancer activities for PEL through regulation of both host and oncogenic virus factors.

  13. Human xenospecific T suppressor cells inhibit T helper cell proliferation to porcine aortic endothelial cells, and NF-kappaB activity in porcine APC.

    Science.gov (United States)

    Ciubotariu, R; Li, J; Colovai, A I; Platt, J L; Cortesini, R; Suciu Foca Cortesini, N

    2001-05-01

    Human T suppressor cells (Ts), capable of preventing autologous T helper cells (Th) from reacting against xenogeneic pig endothelial cells and pig APC can be generated in vitro. Ts derive from a population of CD3(+)CD8(+)CD28(-) T lymphocytes and specifically recognize the MHC class I antigens of the APC used for in vitro immunization. To study the mechanism that underlies suppression, we investigated whether Ts inhibit the expression of costimulatory molecules in xenogeneic professional and semiprofessional APC. We found that Ts down-regulate Th-induced expression of CD86 in pig APC, and that this effect occurs at the level of transcription, as indicated by nuclear run-on and Northern blot assays. EMSA results revealed that inhibition of CD86 expression is mediated by inactivation of transcription factor NF-kappaB. Furthermore, transfection of pig APC with a vector expressing NF-kappaB p65 partially rescued Th-induced expression of the CD86 molecule. These results strongly support the concept that xenospecific Ts inhibit the APC function of xenogeneic cells by preventing activation of NF-kappaB.

  14. Residues in the alternative reading frame tumor suppressor that influence its stability and p53-independent activities

    International Nuclear Information System (INIS)

    Tommaso, Anne di; Hagen, Jussara; Tompkins, Van; Muniz, Viviane; Dudakovic, Amel; Kitzis, Alain; Ladeveze, Veronique; Quelle, Dawn E.

    2009-01-01

    The Alternative Reading Frame (ARF) protein suppresses tumorigenesis through p53-dependent and p53-independent pathways. Most of ARF's anti-proliferative activity is conferred by sequences in its first exon. Previous work showed specific amino acid changes occurred in that region during primate evolution, so we programmed those changes into human p14ARF to assay their functional impact. Two human p14ARF residues (Ala 14 and Thr 31 ) were found to destabilize the protein while two others (Val 24 and Ala 41 ) promoted more efficient p53 stabilization and activation. Despite those effects, all modified p14ARF forms displayed robust p53-dependent anti-proliferative activity demonstrating there are no significant biological differences in p53-mediated growth suppression associated with simian versus human p14ARF residues. In contrast, p53-independent p14ARF function was considerably altered by several residue changes. Val 24 was required for p53-independent growth suppression whereas multiple residues (Val 24 , Thr 31 , Ala 41 and His 60 ) enabled p14ARF to block or reverse the inherent chromosomal instability of p53-null MEFs. Together, these data pinpoint specific residues outside of established p14ARF functional domains that influence its expression and signaling activities. Most intriguingly, this work reveals a novel and direct role for p14ARF in the p53-independent maintenance of genomic stability.

  15. Tomato Spotted Wilt Virus NSs Protein Supports Infection and Systemic Movement of a Potyvirus and Is a Symptom Determinant.

    Science.gov (United States)

    Garcia-Ruiz, Hernan; Gabriel Peralta, Sergio M; Harte-Maxwell, Patricia A

    2018-03-14

    Plant viruses are inducers and targets of antiviral RNA silencing. To condition susceptibility, most plant viruses encode silencing suppressor proteins that interfere with antiviral RNA silencing. The NSs protein is an RNA silencing suppressor in orthotospoviruses, such as the tomato spotted wilt virus (TSWV). The mechanism of RNA silencing suppression by NSs and its role in virus infection and movement are poorly understood. Here, we cloned and tagged TSWV NSs and expressed it from a GFP-tagged turnip mosaic virus (TuMV-GFP) carrying either a wild-type or suppressor-deficient (AS9) helper component proteinase (HC-Pro). When expressed in cis, NSs restored pathogenicity and promoted systemic infection of suppressor-deficient TuMV-AS9-GFP in Nicotiana benthamiana and Arabidopsis thaliana . Inactivating mutations were introduced in NSs RNA-binding domain one. A genetic analysis with active and suppressor-deficient NSs, in combination with wild-type and mutant plants lacking essential components of the RNA silencing machinery, showed that the NSs insert is stable when expressed from a potyvirus. NSs can functionally replace potyviral HC-Pro, condition virus susceptibility, and promote systemic infection and symptom development by suppressing antiviral RNA silencing through a mechanism that partially overlaps that of potyviral HC-Pro. The results presented provide new insight into the mechanism of silencing suppression by NSs and its effect on virus infection.

  16. Mechanism underlying the suppressor activity of retinoic acid on IL4-induced IgE synthesis and its physiological implication.

    Science.gov (United States)

    Seo, Goo-Young; Lee, Jeong-Min; Jang, Young-Saeng; Kang, Seung Goo; Yoon, Sung-Il; Ko, Hyun-Jeong; Lee, Geun-Shik; Park, Seok-Rae; Nagler, Cathryn R; Kim, Pyeung-Hyeun

    2017-12-01

    The present study extends an earlier report that retinoic acid (RA) down-regulates IgE Ab synthesis in vitro. Here, we show the suppressive activity of RA on IgE production in vivo and its underlying mechanisms. We found that RA down-regulated IgE class switching recombination (CSR) mainly through RA receptor α (RARα). Additionally, RA inhibited histone acetylation of germ-line ε (GL ε) promoter, leading to suppression of IgE CSR. Consistently, serum IgE levels were substantially elevated in vitamin A-deficient (VAD) mice and this was more dramatic in VAD-lecithin:retinol acyltransferase deficient (LRAT -/- ) mice. Further, serum mouse mast cell protease-1 (mMCP-1) level was elevated while frequency of intestinal regulatory T cells (Tregs) were diminished in VAD LRAT -/- mice, reflecting that deprivation of RA leads to allergic immune response. Taken together, our results reveal that RA has an IgE-repressive activity in vivo, which may ameliorate IgE-mediated allergic disease. Copyright © 2017 Elsevier Inc. All rights reserved.

  17. Tumour suppressor protein p53 regulates the stress activated bilirubin oxidase cytochrome P450 2A6

    Energy Technology Data Exchange (ETDEWEB)

    Hu, Hao, E-mail: hao.hu1@uqconnect.edu.au [The University of Queensland, National Research Centre for Environmental Toxicology (Entox), 4072 Brisbane, Queensland (Australia); Yu, Ting, E-mail: t.yu2@uq.edu.au [The University of Queensland, National Research Centre for Environmental Toxicology (Entox), 4072 Brisbane, Queensland (Australia); Arpiainen, Satu, E-mail: Satu.Juhila@orion.fi [Institute of Biomedicine, Department of Pharmacology and Toxicology and Medical Research Center Oulu, Oulu University Hospital and University of Oulu, Oulu (Finland); Lang, Matti A., E-mail: m.lang@uq.edu.au [The University of Queensland, National Research Centre for Environmental Toxicology (Entox), 4072 Brisbane, Queensland (Australia); Hakkola, Jukka, E-mail: Jukka.hakkola@oulu.fi [Institute of Biomedicine, Department of Pharmacology and Toxicology and Medical Research Center Oulu, Oulu University Hospital and University of Oulu, Oulu (Finland); Abu-Bakar, A' edah, E-mail: a.abubakar@uq.edu.au [The University of Queensland, National Research Centre for Environmental Toxicology (Entox), 4072 Brisbane, Queensland (Australia)

    2015-11-15

    Human cytochrome P450 (CYP) 2A6 enzyme has been proposed to play a role in cellular defence against chemical-induced oxidative stress. The encoding gene is regulated by various stress activated transcription factors. This paper demonstrates that p53 is a novel transcriptional regulator of the gene. Sequence analysis of the CYP2A6 promoter revealed six putative p53 binding sites in a 3 kb proximate promoter region. The site closest to transcription start site (TSS) is highly homologous with the p53 consensus sequence. Transfection with various stepwise deletions of CYP2A6-5′-Luc constructs – down to − 160 bp from the TSS – showed p53 responsiveness in p53 overexpressed C3A cells. However, a further deletion from − 160 to − 74 bp, including the putative p53 binding site, totally abolished the p53 responsiveness. Electrophoretic mobility shift assay with a probe containing the putative binding site showed specific binding of p53. A point mutation at the binding site abolished both the binding and responsiveness of the recombinant gene to p53. Up-regulation of the endogenous p53 with benzo[α]pyrene – a well-known p53 activator – increased the expression of the p53 responsive positive control and the CYP2A6-5′-Luc construct containing the intact p53 binding site but not the mutated CYP2A6-5′-Luc construct. Finally, inducibility of the native CYP2A6 gene by benzo[α]pyrene was demonstrated by dose-dependent increases in CYP2A6 mRNA and protein levels along with increased p53 levels in the nucleus. Collectively, the results indicate that p53 protein is a regulator of the CYP2A6 gene in C3A cells and further support the putative cytoprotective role of CYP2A6. - Highlights: • CYP2A6 is an immediate target gene of p53. • Six putative p53REs located on 3 kb proximate CYP2A6 promoter region. • The region − 160 bp from TSS is highly homologous with the p53 consensus sequence. • P53 specifically bind to the p53RE on the − 160 bp region. • HNF4

  18. Activating transcription factor-3 (ATF3) functions as a tumor suppressor in colon cancer and is up-regulated upon heat-shock protein 90 (Hsp90) inhibition

    International Nuclear Information System (INIS)

    Hackl, Christina; Stoeltzing, Oliver; Lang, Sven A; Moser, Christian; Mori, Akira; Fichtner-Feigl, Stefan; Hellerbrand, Claus; Dietmeier, Wolfgang; Schlitt, Hans J; Geissler, Edward K

    2010-01-01

    Activating transcription factor-3 (ATF3) is involved in the complex process of cellular stress response. However, its exact role in cancer is discussed controversially because both tumor suppressive and oncogenic effects have been described. Here we followed-up on our previous observation that inhibition of Hsp90 may increase ATF3 expression and sought to determine the role of ATF3 in colon cancer. Regulation of ATF3 was determined in cancer cells using signaling inhibitors and a heat-shock protein-90 (Hsp90) antagonist. Human HCT116 cancer cells were stably transfected with an ATF3-shRNA or a luciferase-shRNA expression plasmid and alterations in cell motility were assessed in migration assays. The impact of ATF3 down-regulation on cancer growth and metastasis were investigated in a subcutaneous tumor model, a model of hepatic tumor growth and in a model of peritoneal carcinomatosis. Human colon cancer tissues were analyzed for ATF3 expression. The results show that therapeutic Hsp90 inhibition substantially up-regulates the expression of ATF3 in various cancer cells, including colon, gastric and pancreatic cancer. This effect was evident both in vitro and in vivo. RNAi mediated knock-down of ATF3 in HCT116 colon cancer cells significantly increased cancer cell migration in vitro. Moreover, in xenogenic mouse models, ATF3 knock-down promoted subcutaneous tumor growth and hepatic metastasis, as well as peritoneal carcinomatosis. Importantly, ATF3 expression was lower in human colon cancer specimens, as compared to corresponding normal surrounding tissues, suggesting that ATF3 may represent a down-regulated tumor suppressor in colon cancer. In conclusion, ATF3 down-regulation in colon cancer promotes tumor growth and metastasis. Considering that blocking Hsp90 induces ATF3 expression, Hsp90 inhibition may represent a valid strategy to treat metastatic colon cancer by up-regulating this anti-metastatic transcription factor

  19. Malignant Trigeminal Nerve Sheath Tumor and Anaplastic Astrocytoma Collision Tumor with High Proliferative Activity and Tumor Suppressor P53 Expression

    Directory of Open Access Journals (Sweden)

    Maher Kurdi

    2014-01-01

    Full Text Available Background. The synchronous development of two primary brain tumors of distinct cell of origin in close proximity or in contact with each other is extremely rare. We present the first case of collision tumor with two histological distinct tumors. Case Presentation. A 54-year-old woman presented with progressive atypical left facial pain and numbness for 8 months. MRI of the brain showed left middle cranial fossa heterogeneous mass extending into the infratemporal fossa. At surgery, a distinct but intermingled intra- and extradural tumor was demonstrated which was completely removed through left orbitozygomatic-temporal craniotomy. Histopathological examination showed that the tumor had two distinct components: malignant nerve sheath tumor of the trigeminal nerve and temporal lobe anaplastic astrocytoma. Proliferative activity and expressed tumor protein 53 (TP53 gene mutations were demonstrated in both tumors. Conclusions. We describe the first case of malignant trigeminal nerve sheath tumor (MTNST and anaplastic astrocytoma in collision and discuss the possible hypothesis of this rare occurrence. We propose that MTNST, with TP53 mutation, have participated in the formation of anaplastic astrocytoma, or vice versa.

  20. Off and back-on again: a tumor suppressor's tale.

    Science.gov (United States)

    Acosta, Jonuelle; Wang, Walter; Feldser, David M

    2018-06-01

    Tumor suppressor genes play critical roles orchestrating anti-cancer programs that are both context dependent and mechanistically diverse. Beyond canonical tumor suppressive programs that control cell division, cell death, and genome stability, unexpected tumor suppressor gene activities that regulate metabolism, immune surveillance, the epigenetic landscape, and others have recently emerged. This diversity underscores the important roles these genes play in maintaining cellular homeostasis to suppress cancer initiation and progression, but also highlights a tremendous challenge in discerning precise context-specific programs of tumor suppression controlled by a given tumor suppressor. Fortunately, the rapid sophistication of genetically engineered mouse models of cancer has begun to shed light on these context-dependent tumor suppressor activities. By using techniques that not only toggle "off" tumor suppressor genes in nascent tumors, but also facilitate the timely restoration of gene function "back-on again" in disease specific contexts, precise mechanisms of tumor suppression can be revealed in an unbiased manner. This review discusses the development and implementation of genetic systems designed to toggle tumor suppressor genes off and back-on again and their potential to uncover the tumor suppressor's tale.

  1. Role of myeloid-derived suppressor cells in amelioration of experimental autoimmune hepatitis following activation of TRPV1 receptors by cannabidiol.

    Directory of Open Access Journals (Sweden)

    Venkatesh L Hegde

    2011-04-01

    Full Text Available Myeloid-derived suppressor cells (MDSCs are getting increased attention as one of the main regulatory cells of the immune system. They are induced at sites of inflammation and can potently suppress T cell functions. In the current study, we demonstrate how activation of TRPV1 vanilloid receptors can trigger MDSCs, which in turn, can inhibit inflammation and hepatitis.Polyclonal activation of T cells, following injection of concanavalin A (ConA, in C57BL/6 mice caused acute hepatitis, characterized by significant increase in aspartate transaminase (AST, induction of inflammatory cytokines, and infiltration of mononuclear cells in the liver, leading to severe liver injury. Administration of cannabidiol (CBD, a natural non-psychoactive cannabinoid, after ConA challenge, inhibited hepatitis in a dose-dependent manner, along with all of the associated inflammation markers. Phenotypic analysis of liver infiltrating cells showed that CBD-mediated suppression of hepatitis was associated with increased induction of arginase-expressing CD11b(+Gr-1(+ MDSCs. Purified CBD-induced MDSCs could effectively suppress T cell proliferation in vitro in arginase-dependent manner. Furthermore, adoptive transfer of purified MDSCs into naïve mice conferred significant protection from ConA-induced hepatitis. CBD failed to induce MDSCs and suppress hepatitis in the livers of vanilloid receptor-deficient mice (TRPV1(-/- thereby suggesting that CBD primarily acted via this receptor to induce MDSCs and suppress hepatitis. While MDSCs induced by CBD in liver consisted of granulocytic and monocytic subsets at a ratio of ∼2∶1, the monocytic MDSCs were more immunosuppressive compared to granulocytic MDSCs. The ability of CBD to induce MDSCs and suppress hepatitis was also demonstrable in Staphylococcal enterotoxin B-induced liver injury.This study demonstrates for the first time that MDSCs play a critical role in attenuating acute inflammation in the liver, and that agents

  2. Effects of siRNA Silencing of TUG1 and LCAL6 Long Non-coding RNAs on Patient-derived Xenograft of Non-small Cell Lung Cancer.

    Science.gov (United States)

    Fang, Tian; Huang, Hairong; Li, Xiaoyou; Liao, Jing; Yang, Zhijian; Hoffman, Robert M; Cheng, X I; Liang, Lei; Hu, Wenjuan; Yun, Shifeng

    2018-01-01

    The aim of the present study was to establish a patient-derived xenograft (PDX) mouse model of non-small cell lung cancer (NSCLC) and investigate the anti-tumor efficacy of silencing of TUG1 and LCAL6 long non-coding RNA in the PDX model. PDXs were established by subcutaneously implanting NSCLC surgical tumor fragments into immunodeficient mice. PDX characterization was performed by histopathological, immunohistochemical and real-time polymerase chain reaction (RT-PCR) analyses for NSCLC subtype-specific markers and expression of LCAL6 and TUG1. Anti-tumor efficacy of siRNA silencing of TUG1 and LCAL6 was also investigated in the PDX model. The effect of TUG1 and LCAL6 silencing on protein expression of proliferation marker Ki67 and HOX-gene family HOXB7 in the tumors was assessed by immunohistochemical staining and Western blotting. Establishment of NSCLC PDX models resulted in 9 of 26 cases (34.6%). Lung squamous cell carcinomas (SCC) had a higher engraftment rate (58.3%) than lung adenocarcinomas (ADC) (18.2%) (pTUG1. The tumor volume and weight were significantly reduced in the TUG1-silenced group as compared to the control group (p0.05). Expression of both TUG1and LCAL6 was reduced by siRNA treatment. Expression of Ki67 and HOXB7 was significantly suppressed in both the TUG1- and LCAL6-silenced groups compared to the control group (pTUG1-silenced group showed more reduced Ki67 expression than the LCAL6-silenced group (pTUG1 and LCAL6. Silencing of TUG1 inhibited both tumor growth and expression of the proliferation marker Ki67 and HOX-gene family HOXB7 in the PDX model of NSCLC. Copyright© 2018, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

  3. Modulation of allogeneic stimulation in man. I. Characterization of an in vitro induced suppressor macrophage population

    International Nuclear Information System (INIS)

    Stux, S.V.; Dubey, D.P.; Yunis, E.J.

    1981-01-01

    Cultured human peripheral blood mononuclear cells suppressed the allogeneic response of fresh autologous lymphocytes. This suppressor activity developed gradually over a period of one week. The cells primarily responsible for this effect were enriched by Ficoll density gradient centrifugation. It was found that the suppressor cell is a large, low density nylon wool adherent, radioresistant, phagocytic, and nonspecific esterase positive mononuclear cell. Moreover, these cells did not form E rosettes and were Fc positive. Electron microscopy confirmed that suppressor cells were macrophage like. Suppressor activity was not due to cytotoxicity, crowding, or steric hinderance by the cultured cells. The suppressor macrophage population did not appear to inhibit the allogeneic response via prostaglandin or arginase release, or interfere with the tritiated thymidine uptake by release of endogenous thymidine. The above system is viewed as an in vitro model of immune regulation by suppressor macrophages, in the context of allogeneic response

  4. RET is a potential tumor suppressor gene in colorectal cancer

    Science.gov (United States)

    Luo, Yanxin; Tsuchiya, Karen D.; Park, Dong Il; Fausel, Rebecca; Kanngurn, Samornmas; Welcsh, Piri; Dzieciatkowski, Slavomir; Wang, Jianping; Grady, William M.

    2012-01-01

    Cancer arises as the consequence of mutations and epigenetic alterations that activate oncogenes and inactivate tumor suppressor genes. Through a genome-wide screen for methylated genes in colon neoplasms, we identified aberrantly methylated RET in colorectal cancer. RET, a transmembrane receptor tyrosine kinase and a receptor for the GDNF-family ligands, was one of the first oncogenes to be identified and has been shown to be an oncogene in thyroid cancer and pheochromocytoma. However, unexpectedly, we found RET is methylated in 27% of colon adenomas and in 63% of colorectal cancers, and now provide evidence that RET has tumor suppressor activity in colon cancer. The aberrant methylation of RET correlates with decreased RET expression, whereas the restoration of RET in colorectal cancer cell lines results in apoptosis. Furthermore, in support of a tumor suppressor function of RET, mutant RET has also been found in primary colorectal cancer. We now show that these mutations inactivate RET, which is consistent with RET being a tumor suppressor gene in the colon. These findings suggest that the aberrant methylation of RET and the mutational inactivation of RET promote colorectal cancer formation and that RET can serve as a tumor suppressor gene in the colon. Moreover, the increased frequency of methylated RET in colon cancers compared to adenomas suggests RET inactivation is involved in the progression of colon adenomas to cancer. PMID:22751117

  5. Nonspecific suppressor T cells cause decreased mixed lymphocyte culture reactivity in bone marrow transplant patients

    International Nuclear Information System (INIS)

    Harada, M.; Ueda, M.; Nakao, S.; Kondo, K.; Odaka, K.; Shiobara, S.; Matsue, K.; Mori, T.; Matsuda, T.

    1986-01-01

    Decreased reactivity in mixed lymphocyte culture (MLC) was observed in patients within 1 yr after allogeneic and autologous bone marrow transplantation. Suppressor activity of peripheral blood mononuclear cells (PBMC) from transplant patients was studied by adding these cells as modulator cells to a bidirectional MLC with cells from normal individuals. PBMC from transplant patients markedly suppressed MLC reactivity in a dose-dependent manner. Suppressor activity was present in cells forming rosettes with sheep erythrocytes. Treatment of modulator cells with monoclonal antibodies against T cell differentiation antigens (OKT8, OKIa1) and complement completely abolished suppression of MLC. Suppressor activity was unaffected by 30 Gy irradiation. Suppressor activity declined gradually after transplantation and was inversely correlated with MLC reactivity of each patient at a significant level (p less than 0.01). These observations suggest that OKT8+ Ia+ radioresistant suppressor T cells play a role in the development of decreased MLC reactivity observed during the early post-transplant period

  6. Data on mass spectrometry-based proteomics for studying the involvement of CYLD in the ubiquitination events downstream of EGFR activation

    Directory of Open Access Journals (Sweden)

    Virginia Sanchez-Quiles

    2018-06-01

    Full Text Available The present data article corresponds to the proteomic data of the involvement of Cylindromatosis protein (CYLD in the ubiquitination signaling initiated by EGF stimulation. CYLD tumor suppressor protein has Lys63-chain deubiquitinase activity that has been proved essential for the negative regulation of crucial signaling mechanisms, namely the NFkB pathway. Previous results have suggested the involvement of CYLD in the EGF-dependent signal transduction as well, showing its engagement within the tyrosine-phosphorylated complexes formed following the addition of the growth factor. EGFR signaling participates in central cellular processes and its tight regulation, partly through ubiquitination cascades, is decisive for a balanced cellular homeostasis. We carried out the substitution of the endogenous pool of ubiquitin for a His-FLAG-tagged ubiquitin (Stable Ubiquitin Exchange, StUbEx, in combination with the shRNA silencing of CYLD and SILAC-labeling on HeLa cells. The subsequent tandem affinity purification of ubiquitinated proteins in control and CYLD-depleted cells was followed by mass spectrometric analysis. Therefore, we present an unbiased study investigating the impact of CYLD in the EGF-dependent ubiquitination. The data supplied herein is related to the research article entitled “Cylindromatosis tumor suppressor protein (CYLD deubiquitinase is necessary for proper ubiquitination and degradation of the epidermal growth factor receptor” (Sanchez-Quiles et al., 2017 [1]. We provide the associated mass spectrometry raw files, excel tables and gene ontology enrichments. The data have been deposited in the ProteomeXchange with the identifier PXD003423.

  7. The milk protein α-casein functions as a tumor suppressor via activation of STAT1 signaling, effectively preventing breast cancer tumor growth and metastasis

    Science.gov (United States)

    Bonuccelli, Gloria; Castello-Cros, Remedios; Capozza, Franco; Martinez-Outschoorn, Ubaldo E.; Lin, Zhao; Tsirigos, Aristotelis; Xuanmao, Jiao; Whitaker-Menezes, Diana; Howell, Anthony; Lisanti, Michael P.; Sotgia, Federica

    2012-01-01

    Here, we identified the milk protein α-casein as a novel suppressor of tumor growth and metastasis. Briefly, Met-1 mammary tumor cells expressing α-casein showed a ~5-fold reduction in tumor growth and a near 10-fold decrease in experimental metastasis. To identify the molecular mechanism(s), we performed genome-wide transcriptional profiling. Interestingly, our results show that α-casein upregulates gene transcripts associated with interferon/STAT1 signaling and downregulates genes associated with “stemness.” These findings were validated by immunoblot and FACS analysis, which showed the upregulation and hyperactivation of STAT1 and a decrease in the number of CD44(+) “cancer stem cells.” These gene signatures were also able to predict clinical outcome in human breast cancer patients. Thus, we conclude that a lactation-based therapeutic strategy using recombinant α-casein would provide a more natural and non-toxic approach to the development of novel anticancer therapies. PMID:23047602

  8. The Quest for the 1p36 Tumor Suppressor

    Science.gov (United States)

    Bagchi, Anindya; Mills, Alea A.

    2010-01-01

    Genomic analyses of late-stage human cancers have uncovered deletions encompassing 1p36, thereby providing an extensive body of literature supporting the idea that a potent tumor suppressor resides in this interval. Although a number of genes have been proposed as 1p36 candidate tumor suppressors, convincing evidence that their encoded products protect from cancer has been scanty. A recent functional study identified CHD5 as a novel tumor suppressor mapping to 1p36. Here we discuss evidence supporting CHD5’s tumor suppressive role. Together, these findings suggest that strategies designed to enhance CHD5 activity could provide novel approaches for treating a broad range of human malignancies. PMID:18413720

  9. Identification of a maize chlorotic dwarf virus silencing suppressor protein

    Science.gov (United States)

    Maize chlorotic dwarf virus (MCDV), a member of the genus Waikavirus, family Secoviridae, has a 11784 nt (+)ssRNA genome that encodes a 389 kDa proteolytically processed polyprotein. We show that an N-terminal 78kDa polyprotein (R78) has silencing suppressor activity, that it is cleaved by the viral...

  10. Microbial Regulation of p53 Tumor Suppressor.

    Directory of Open Access Journals (Sweden)

    Alexander I Zaika

    2015-09-01

    Full Text Available p53 tumor suppressor has been identified as a protein interacting with the large T antigen produced by simian vacuolating virus 40 (SV40. Subsequent research on p53 inhibition by SV40 and other tumor viruses has not only helped to gain a better understanding of viral biology, but also shaped our knowledge of human tumorigenesis. Recent studies have found, however, that inhibition of p53 is not strictly in the realm of viruses. Some bacterial pathogens also actively inhibit p53 protein and induce its degradation, resulting in alteration of cellular stress responses. This phenomenon was initially characterized in gastric epithelial cells infected with Helicobacter pylori, a bacterial pathogen that commonly infects the human stomach and is strongly linked to gastric cancer. Besides H. pylori, a number of other bacterial species were recently discovered to inhibit p53. These findings provide novel insights into host-bacteria interactions and tumorigenesis associated with bacterial infections.

  11. Human cord blood suppressor T lymphocytes. II. Characterization of inducer of suppressor cells

    International Nuclear Information System (INIS)

    Cheng, H.; Delespesse, G.

    1986-01-01

    Previously, we reported an antigen nonspecific inducer of T suppressor cell factor (TisF) produced by cord blood mononuclear cells (MNC) in 48-hr, two-way mixed lymphocyte cultures (MLC). The target of this factor was a radiosensitive, T4+ (T8-) adult suppressor T cell subset. The cellular origin of this TisF was examined in the present study. IgG production by pokeweed mitogen (PWM)-stimulated adult MNC was used as an assay for TisF activity. It was found that TisF-producing cells formed rosettes with sheep erythrocytes (E+) and were independent of adherent cells (AC) in the production of TisF. They were resistant to irradiation (2500 rads) and phenotypic characterization with T cell reactive monoclonal antibodies indicated that they resided in the T8- (T4+) population. Furthermore, both TQ1- and TQ1+ cells were required for the production of TisF activity and such activity could not be reconstituted by supernatants from TQ1- MLC and TQ1+ MLC. These results indicate that the production of TisF is dependent upon interactions between radioresistant E+, T8-, TQ1- and radioresistant E+, T8-, TQ1+ cells

  12. In vitro X-ray irradiation of human peripheral blood T lymphocytes enhances suppressor function

    International Nuclear Information System (INIS)

    Ogawa, H.; Tsunematsu, T.

    1983-01-01

    The effect of in vitro X-ray irradiation on human peripheral blood T lymphocytes was studied with regard to their suppressor activity related to the concanavalin A (Con A)-induced suppressor system. To generate suppressor T lymphocytes, purified human T lymphocytes were incubated for 3 days in the first culture, with or without Con A. These lymphocytes were irradiated with various doses of X-ray before, mid or after the culture. After doing a second culture for 6 days, the suppressive influence of these cells on T lymphocyte proliferation rates stimulated with allogeneic mononuclear cells, and B lymphocyte proliferation rates stimulated with pokeweed mitogen was measured. Irradiation of cultures to which Con A had not been added induced much the same level of suppressor activity as seen in the cultures with Con A. The suppressor activity gradually increased with time from the irradiation to the suppressor cell assay. Suppressor T lymphocytes were resistant to X-ray irradiation and independent of DNA synthesis. However, irradiation-induced enhancement was minimal in cultures incubated with con A, regardless of the irradiation time. (author)

  13. Suppressor Effects of Coping Strategies on Resilience

    Science.gov (United States)

    Yoon, Jae ho; Lee, Ji hae; Lee, Chae Yeon; Cho, Minhee; Lee, Sang Min

    2014-01-01

    The purpose of the current study is to demonstrate a significant suppressor effect among coping strategies on resilience. Two different samples were used to replicate the suppressor effect. Participants in the first example were 391 adolescents (middle school students) in Korea, and participants in the second example were 282 young adults…

  14. Avian Reovirus Protein p17 Functions as a Nucleoporin Tpr Suppressor Leading to Activation of p53, p21 and PTEN and Inactivation of PI3K/AKT/mTOR and ERK Signaling Pathways.

    Directory of Open Access Journals (Sweden)

    Wei-Ru Huang

    Full Text Available Avian reovirus (ARV protein p17 has been shown to regulate cell cycle and autophagy by activation of p53/PTEN pathway; nevertheless, it is still unclear how p53 and PTEN are activated by p17. Here, we report for the first time that p17 functions as a nucleoporin Tpr suppressor that leads to p53 nuclear accumulation and consequently activates p53, p21, and PTEN. The nuclear localization signal (119IAAKRGRQLD128 of p17 has been identified for Tpr binding. This study has shown that Tpr suppression occurs by p17 interacting with Tpr and by reducing the transcription level of Tpr, which together inhibit Tpr function. In addition to upregulation of PTEN by activation of p53 pathway, this study also suggests that ARV protein p17 acts as a positive regulator of PTEN. ARV p17 stabilizes PTEN by stimulating phosphorylation of cytoplasmic PTEN and by elevating Rak-PTEN association to prevent it from E3 ligase NEDD4-1 targeting. To activate PTEN, p17 is able to promote β-arrestin-mediated PTEN translocation from the cytoplasm to the plasma membrane via a Rock-1-dependent manner. The accumulation of p53 in the nucleus induces the PTEN- and p21-mediated downregulation of cyclin D1 and CDK4. Furthermore, Tpr and CDK4 knockdown increased virus production in contrast to depletion of p53, PTEN, and LC3 reducing virus yield. Taken together, our data suggest that p17-mediated Tpr suppression positively regulates p53, PTEN, and p21 and negatively regulates PI3K/AKT/mTOR and ERK signaling pathways, both of which are beneficial for virus replication.

  15. Analysis of Tospovirus NSs Proteins in Suppression of Systemic Silencing.

    Science.gov (United States)

    Hedil, Marcio; Sterken, Mark G; de Ronde, Dryas; Lohuis, Dick; Kormelink, Richard

    2015-01-01

    RNA silencing is a sequence-specific gene regulation mechanism that in plants also acts antiviral. In order to counteract antiviral RNA silencing, viruses have evolved RNA silencing suppressors (RSS). In the case of tospoviruses, the non-structural NSs protein has been identified as the RSS. Although the tomato spotted wilt virus (TSWV) tospovirus NSs protein has been shown to exhibit affinity to long and small dsRNA molecules, its ability to suppress the non-cell autonomous part of RNA silencing has only been studied to a limited extent. Here, the NSs proteins of TSWV, groundnut ringspot virus (GRSV) and tomato yellow ring virus (TYRV), representatives for three distinct tospovirus species, have been studied on their ability and strength to suppress local and systemic silencing. A system has been developed to quantify suppression of GFP silencing in Nicotiana benthamiana 16C lines, to allow a comparison of relative RNA silencing suppressor strength. It is shown that NSs of all three tospoviruses are suppressors of local and systemic silencing. Unexpectedly, suppression of systemic RNA silencing by NSsTYRV was just as strong as those by NSsTSWV and NSsGRSV, even though NSsTYRV was expressed in lower amounts. Using the system established, a set of selected NSsTSWV gene constructs mutated in predicted RNA binding domains, as well as NSs from TSWV isolates 160 and 171 (resistance breakers of the Tsw resistance gene), were analyzed for their ability to suppress systemic GFP silencing. The results indicate another mode of RNA silencing suppression by NSs that acts further downstream the biogenesis of siRNAs and their sequestration. The findings are discussed in light of the affinity of NSs for small and long dsRNA, and recent mutant screen of NSsTSWV to map domains required for RSS activity and triggering of Tsw-governed resistance.

  16. Suppressor cells in transplantation tolerance II. Maturation of suppressor cells in the bone marrow chimera

    International Nuclear Information System (INIS)

    Tutschka, P.J.; Ki, P.F.; Beschorner, W.E.; Hess, A.D.; Santos, G.W.

    1981-01-01

    Histoincompatible bone marrow allografts were established in lethally irradiated rats. At various times after transplantation, the spleen cells were harvested, subjected to mixed lymphocyte cultures, and assayed for suppressor cells in vitro and in vivo by adoptive transfer studies. Alloantigen-nonspecific suppressor cells appeared in the chimera at 40 days after grafting, coinciding with the resolution of graft-versus-host disease (GVHD). At 250 days the nonspecific suppressor cells were replaced by suppressor cells specifically suppressing donor-versus-host alloantigen responses. At 720 days suppressor cells could no longer be identified by in vitro methods but were identified by in vivo adoptive transfer of transplantation tolerance. After injection of host-type antigen into chimeras, the suppressor cells could be again demonstrated by in vitro methods

  17. Suppressor cells in transplantation tolerance. II. maturation of suppressor cells in the bone marrow chimera

    International Nuclear Information System (INIS)

    Tutschka, P.J.; Ki, P.F.; Beschorner, W.E.; Hess, A.D.; Santos, G.W.

    1981-01-01

    Histoincompatible bone marrow allografts were established in lethally irradiated rats. At various times after transplantation, the spleen cells were harvested, subjected to mixed lymphocyte cultures, and assayed for suppressor cells in vitro and in vivo by adoptive transfer studies. Alloantigen-nonspecific suppressor cells appeared in the chimera at 40 days after grafting, coinciding with the resolution of graft-versus-host disease (GVHD). At 250 days the nonspecific suppressor cells were replaced by suppressor cells specifically suppressing donor-versus-host alloantigen responses. At 720 days suppressor cells could no longer be identified by in vitro methods but were identified by in vivo adoptive transfer of transplantation tolerance. After injection of host-type antigen into chimeras, the suppressor cells could be again demonstrated by in vitro methods

  18. Stable shRNA Silencing of Lactate Dehydrogenase A (LDHA) in Human MDA-MB-231 Breast Cancer Cells Fails to Alter Lactic Acid Production, Glycolytic Activity, ATP or Survival.

    Science.gov (United States)

    Mack, Nzinga; Mazzio, Elizabeth A; Bauer, David; Flores-Rozas, Hernan; Soliman, Karam F A

    2017-03-01

    In the US, African Americans have a high death rate from triple-negative breast cancer (TNBC), characterized by lack of hormone receptors (ER, PR, HER2/ERRB2) which are otherwise valuable targets of chemotherapy. There is a need to identify novel targets that negatively impact TNBC tumorigenesis. TNBCs release an abundance of lactic acid, under normoxic, hypoxic and hyperoxic conditions; this referred to as the Warburg effect. Accumulated lactic acid sustains peri-cellular acidity which propels metastatic invasion and malignant aggressive transformation. The source of lactic acid is believed to be via conversion of pyruvate by lactate dehydrogenase (LDH) in the last step of glycolysis, with most studies focusing on the LDHA isoform. In this study, LDHA was silenced using long-term MISSION® shRNA lentivirus in human breast cancer MDA-MB-231 cells. Down-regulation of LDHA transcription and protein expression was confirmed by western blot, immunocytochemistry and qPCR. A number of parameters were measured in fully viable vector controls versus knock-down (KD) clones, including levels of lactic acid produced, glucose consumed, ATP and basic metabolic rates. The data show that lentivirus V-165 generated a knock-down clone most effective in reducing both gene and protein levels to less than 1% of vector controls. Stable KD showed absolutely no changes in cell viability, lactic acid production, ATP, glucose consumption or basic metabolic rate. Given the complete absence of impact on any observed parameter by LDH-A KD and this being somewhat contrary to findings in the literature, further analysis was required to determine why. Whole-transcriptome analytic profile on MDA-MB-231 for LDH subtypes using Agilent Human Genome 4×44k microarrays, where the data show the following component breakdown. Transcripts: 30.47 % LDHA, 69.36% LDHB, 0.12% LDHC and 0.05% LDHD. These findings underscore the importance of alternative isoforms of LDH in cancer cells to produce lactic acid, when LDHA is silenced or inhibited. LDHA silencing alone is not effective in hampering or inducing changes in survival, metabolism or lactic acid produced in a cell line with high concentrations of LDHB. Future research will be required to confirm effects of dual LDHA/B knockdown and further confirm that the sole source of lactic acid produced occurs through LDH (all isoforms) in breast cancer cells. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

  19. Gelsolin negatively regulates the activity of tumor suppressor p53 through their physical interaction in hepatocarcinoma HepG2 cells

    International Nuclear Information System (INIS)

    An, Joo-Hee; Kim, Jung-Woong; Jang, Sang-Min; Kim, Chul-Hong; Kang, Eun-Jin; Choi, Kyung-Hee

    2011-01-01

    Highlights: → The actin binding protein Gelsolin (GSN) interacts with transcription factor p53. → GSN interacts with transactivation- and DNA binding domains of p53. → GSN represses transactivity of p53 via inhibition of nuclear translocation of p53. → GSN inhibits the p53-mediated apoptosis in hepatocarcinoma HepG2 cells. -- Abstract: As a transcription factor, p53 modulates several cellular responses including cell-cycle control, apoptosis, and differentiation. In this study, we have shown that an actin regulatory protein, gelsolin (GSN), can physically interact with p53. The nuclear localization of p53 is inhibited by GSN overexpression in hepatocarcinoma HepG2 cells. Additionally, we demonstrate that GSN negatively regulates p53-dependent transcriptional activity of a reporter construct, driven by the p21-promoter. Furthermore, p53-mediated apoptosis was repressed in GSN-transfected HepG2 cells. Taken together, these results suggest that GSN binds to p53 and this interaction leads to the inhibition of p53-induced apoptosis by anchoring of p53 in the cytoplasm in HepG2 cells.

  20. INHIBITION OF THE DNA-BINDING ACTIVITY OF DROSOPHILA SUPPRESSOR OF HAIRLESS AND OF ITS HUMAN HOMOLOG, KBF2/RBP-J-KAPPA, BY DIRECT PROTEIN-PROTEIN INTERACTION WITH DROSOPHILA HAIRLESS

    NARCIS (Netherlands)

    BROU, C; LOGEAT, F; LECOURTOIS, M; VANDEKERCKHOVE, Joël; KOURILSKY, P; SCHWEISGUTH, F; ISRAEL, A

    1994-01-01

    We have purified the sequence-specific DNA-binding protein KBF2 and cloned the corresponding cDNA, which is derived from the previously described RBP-J kappa gene, the human homolog of the Drosophila Suppressor of Hairless [Su(H)] gene. Deletion studies of the RBP-J kappa and Su(H) proteins allowed

  1. Imprinted CDKN1C is a tumor suppressor in rhabdoid tumor and activated by restoration of SMARCB1 and histone deacetylase inhibitors.

    Directory of Open Access Journals (Sweden)

    Elizabeth M Algar

    Full Text Available SMARCB1 is deleted in rhabdoid tumor, an aggressive paediatric malignancy affecting the kidney and CNS. We hypothesized that the oncogenic pathway in rhabdoid tumors involved epigenetic silencing of key cell cycle regulators as a consequence of altered chromatin-remodelling, attributable to loss of SMARCB1, and that this hypothesis if proven could provide a biological rationale for testing epigenetic therapies in this disease. We used an inducible expression system to show that the imprinted cell cycle inhibitor CDKN1C is a downstream target for SMARCB1 and is transcriptionally activated by increased histone H3 and H4 acetylation at the promoter. We also show that CDKN1C expression induces cell cycle arrest, CDKN1C knockdown with siRNA is associated with increased proliferation, and is able to compete against the anti-proliferative effect of restored SMARCB1 expression. The histone deacetylase inhibitor (HDACi, Romidepsin, specifically restored CDKN1C expression in rhabdoid tumor cells through promoter histone H3 and H4 acetylation, recapitulating the effect of SMARCB1 on CDKNIC allelic expression, and induced cell cycle arrest in G401 and STM91-01 rhabdoid tumor cell lines. CDKN1C expression was also shown to be generally absent in clinical specimens of rhabdoid tumor, however CDKN1A and CDKN1B expression persisted. Our observations suggest that maintenance of CDKN1C expression plays a critical role in preventing rhabdoid tumor growth. Significantly, we report for the first time, parallels between the molecular pathways of SMARCB1 restoration and Romidepsin treatment, and demonstrate a biological basis for the further exploration of histone deacetylase inhibitors as relevant therapeutic reagents in the treatment of rhabdoid tumor.

  2. Regulatory role for the memory B cell as suppressor-inducer of feedback control

    International Nuclear Information System (INIS)

    Kennedy, M.W.; Thomas, D.B.

    1983-01-01

    A regulatory role is proposed for the antigen-responsive B cell, as suppressor-inducer of feedback control during the secondary response in vivo. In a double adoptive transfer of memory cells primed to a thymus-dependent antigen from one irradiated host to another, antigen-specific suppressors are generated after a critical time in the primary recipient, able to entirely ablate a secondary anti-hapten response. Positive cell selection in the fluorescence-activated cell sorter confirmed that suppression was mediated by an Lyt-2+ T cell; however, positively selected B cells were also inhibitory and able to induce suppressors in a carrier-specific manner: B hapten induced suppressors in a carrier-primed population, and B carrier induced suppressors in a hapten-carrier population. At the peak of the antibody response in the primary host, memory B cells and their progeny were unable to differentiate further to plasma cells due to their intrinsic suppressor-inducer activity, but this autoregulatory circuit could be severed by adoptive transfer to carrier-primed, X-irradiated recipients

  3. Differences in serum IgA responses to HIV-1 gp41 in elite controllers compared to viral suppressors on highly active antiretroviral therapy.

    Directory of Open Access Journals (Sweden)

    Rafiq Nabi

    Full Text Available Mechanisms responsible for natural control of human immunodeficiency type 1 (HIV replication in elite controllers (EC remain incompletely defined. To determine if EC generate high quality HIV-specific IgA responses, we used Western blotting to compare the specificities and frequencies of IgA to HIV antigens in serum of gender-, age- and race-matched EC and aviremic controllers (HC and viremic noncontrollers (HN on highly active antiretroviral therapy (HAART. Concentrations and avidity of IgA to HIV antigens were measured using ELISA or multiplex assays. Measurements for IgG were performed in parallel. EC were found to have stronger p24- and V1V2-specific IgG responses than HN, but there were no IgG differences for EC and HC. In contrast, IgA in EC serum bound more frequently to gp160 and gag proteins than IgA in HC or HN. The avidity of anti-gp41 IgA was also greater in EC, and these subjects had stronger IgA responses to the gp41 heptad repeat region 1 (HR1, a reported target of anti-bacterial RNA polymerase antibodies that cross react with gp41. However, EC did not demonstrate greater IgA responses to E. coli RNA polymerase or to peptides containing the shared LRAI sequence, suggesting that most of their HR1-specific IgA antibodies were not induced by intestinal microbiota. In both EC and HAART recipients, the concentrations of HIV-specific IgG were greater than HIV-specific IgA, but their avidities were comparable, implying that they could compete for antigen. Exceptions were C1 peptides and V1V2 loops. IgG and IgA responses to these antigens were discordant, with IgG reacting to V1V2, and IgA reacting to C1, especially in EC. Interestingly, EC with IgG hypergammaglobulinemia had greater HIV-specific IgA and IgG responses than EC with normal total IgG levels. Heterogeneity in EC antibody responses may therefore be due to a more focused HIV-specific B cell response in some of these individuals. Overall, these data suggest that development of

  4. Analysis of Tospovirus NSs Proteins in Suppression of Systemic Silencing

    NARCIS (Netherlands)

    Hedil, M.; Sterken, M.G.; Ronde, de D.; Lohuis, D.; Kormelink, R.

    2015-01-01

    RNA silencing is a sequence-specific gene regulation mechanism that in plants also acts antiviral. In order to counteract antiviral RNA silencing, viruses have evolved RNA silencing suppressors (RSS). In the case of tospoviruses, the non-structural NSs protein has been identified as the RSS.

  5. High-level HIV-1 Nef transient expression in Nicotiana benthamiana using the P19 gene silencing suppressor protein of Artichoke Mottled Crinckle Virus

    Directory of Open Access Journals (Sweden)

    Bianco Linda

    2009-11-01

    Full Text Available Abstract Background In recent years, different HIV antigens have been successfully expressed in plants by either stable transformation or transient expression systems. Among HIV proteins, Nef is considered a promising target for the formulation of a multi-component vaccine due to its implication in the first steps of viral infection. Attempts to express Nef as a single protein product (not fused to a stabilizing protein in transgenic plants resulted in disappointingly low yields (about 0.5% of total soluble protein. In this work we describe a transient expression system based on co-agroinfiltration of plant virus gene silencing suppressor proteins in Nicotiana benthamiana, followed by a two-step affinity purification protocol of plant-derived Nef. Results The effect of three gene silencing viral suppressor proteins (P25 of Potato Virus X, P19 of either Artichoke Mottled Crinckle virus and Tomato Bushy Stunt virus on Nef transient expression yield was evaluated. The P19 protein of Artichoke Mottled Crinckle virus (AMCV-P19 gave the highest expression yield in vacuum co-agroinfiltration experiments reaching 1.3% of total soluble protein, a level almost three times higher than that previously reported in stable transgenic plants. The high yield observed in the co-agroinfiltrated plants was correlated to a remarkable decrease of Nef-specific small interfering RNAs (siRNAs indicating an effective modulation of RNA silencing mechanisms by AMCV-P19. Interestingly, we also showed that expression levels in top leaves of vacuum co-agroinfiltrated plants were noticeably reduced compared to bottom leaves. Moreover, purification of Nef from agroinfiltrated tissue was achieved by a two-step immobilized metal ion affinity chromatography protocol with yields of 250 ng/g of fresh tissue. Conclusion We demonstrated that expression level of HIV-1 Nef in plant can be improved using a transient expression system enhanced by the AMCV-P19 gene silencing suppressor

  6. An unusual characteristic "flower-like" pattern: flash suppressor burns.

    Science.gov (United States)

    Gurcan, Altun

    2012-04-01

    The case on contact shots from firearms with a flash suppressor is rare. When a rifle fitted with a flash suppressor is fired, the emerging soot-laden gas in the barrel escapes from the slits of the flash suppressor. If the shot is contact or near contact, the flash suppressor will produce a characteristic "flower-like" pattern of seared, blackened zones around the entrance. This paper presents the injury pattern of the flash suppressor in a 29-year-old man who committed suicide with a G3 automatic infantry rifle.

  7. Multielement suppressor nozzles for thrust augmentation systems.

    Science.gov (United States)

    Lawrence, R. L.; O'Keefe, J. V.; Tate, R. B.

    1972-01-01

    The noise reduction and nozzle performance characteristics of large-scale, high-aspect-ratio multielement nozzle arrays operated at low velocities were determined by test. The nozzles are selected for application to high-aspect-ratio augmentor suppressors to be used for augmentor wing airplanes. Significant improvements in noise characteristics for multielement nozzles over those of round or high-aspect-ratio slot nozzles are obtained. Elliptical noise patterns typical of slot nozzles are presented for high-aspect-ratio multielement nozzle arrays. Additional advantages are available in OASPL noise reduction from the element size and spacing. Augmentor-suppressor systems can be designed for maximum beam pattern directivity and frequency spectrum shaping advantages. Measurements of the nozzle wakes show a correlation with noise level data and frequency spectrum peaks. The noise and jet wake results are compared with existing prediction procedures based on empirical jet flow equations, Lighthill relationships, Strouhal number, and empirical shock-induced screech noise effects.

  8. Suppressor Analysis of the Fusogenic Lambda Spanins.

    Science.gov (United States)

    Cahill, Jesse; Rajaure, Manoj; Holt, Ashley; Moreland, Russell; O'Leary, Chandler; Kulkarni, Aneesha; Sloan, Jordan; Young, Ry

    2017-07-15

    The final step of lysis in phage λ infections of Escherichia coli is mediated by the spanins Rz and Rz1. These proteins form a complex that bridges the cell envelope and that has been proposed to cause fusion of the inner and outer membranes. Accordingly, mutations that block spanin function are found within coiled-coil domains and the proline-rich region, motifs essential in other fusion systems. To gain insight into spanin function, pseudorevertant alleles that restored plaque formation for lysis-defective mutants of Rz and Rz1 were selected. Most second-site suppressors clustered within a coiled-coil domain of Rz near the outer leaflet of the cytoplasmic membrane and were not allele specific. Suppressors largely encoded polar insertions within the hydrophobic core of the coiled-coil interface. Such suppressor changes resulted in decreased proteolytic stability of the Rz double mutants in vivo Unlike the wild type, in which lysis occurs while the cells retain a rod shape, revertant alleles with second-site suppressor mutations supported lysis events that were preceded by spherical cell formation. This suggests that destabilization of the membrane-proximal coiled coil restores function for defective spanin alleles by increasing the conformational freedom of the complex at the cost of its normal, all-or-nothing functionality. IMPORTANCE Caudovirales encode cell envelope-spanning proteins called spanins, which are thought to fuse the inner and outer membranes during phage lysis. Recent genetic analysis identified the functional domains of the lambda spanins, which are similar to class I viral fusion proteins. While the pre- and postfusion structures of model fusion systems have been well characterized, the intermediate structure(s) formed during the fusion reaction remains elusive. Genetic analysis would be expected to identify functional connections between intermediates. Since most membrane fusion systems are not genetically tractable, only few such

  9. Subpopulation of human helper and suppressor T lymphocytes

    International Nuclear Information System (INIS)

    Venkataraman, M.; Levin, R.D.; Westerman, M.P.

    1983-01-01

    Mitogen driven differentiation of normal human mononuclear cells is a well-established model for the study of antibody synthesis in man. In certain rare individuals who are clinically normal, unfractionated mononuclear cells or a mixture of purified B plus T lymphocytes differentiate into immunoglobulin producing cells in response to purified protein derivative of tuberculin (PPD) but not in response to pokeweed mitogen (PWM). To evaluate this observation we have irradiated T cells from such individuals to eliminate naturally occurring suppressor T cell activity and then added the irradiated T cells back to autologous B cells before culture. The B cells then responded to PWM. The original PPD responses of cells from these individuals were now significantly reduced. Although, there was no difference between PWM nonresponders and responders in the number of OKT-8 positive cells, elimination of OKT-8 positive cells in the PWM nonresponders with OKT-8 monoclonal antibody and complement resulted in a significantly increased response to PWM. This study indicates that there are suppressor T cells which specifically inhibit B cell response to PWM without affecting the PPD response. These results also show that the helper T cells involved in the PWM response are radioresistant and those involved in the PPD response are radiosensitive

  10. History of myeloid-derived suppressor cells.

    Science.gov (United States)

    Talmadge, James E; Gabrilovich, Dmitry I

    2013-10-01

    Tumour-induced granulocytic hyperplasia is associated with tumour vasculogenesis and escape from immunity via T cell suppression. Initially, these myeloid cells were identified as granulocytes or monocytes; however, recent studies have revealed that this hyperplasia is associated with populations of multipotent progenitor cells that have been identified as myeloid-derived suppressor cells (MDSCs). The study of MDSCs has provided a wealth of information regarding tumour pathobiology, has extended our understanding of neoplastic progression and has modified our approaches to immune adjuvant therapy. In this Timeline article, we discuss the history of MDSCs, their influence on tumour progression and metastasis, and the crosstalk between tumour cells, MDSCs and the host macroenvironment.

  11. Anti-proliferative and pro-apoptotic activity of whole extract and isolated indicaxanthin from Opuntia ficus-indica associated with re-activation of the onco-suppressor p16(INK4a) gene in human colorectal carcinoma (Caco-2) cells.

    Science.gov (United States)

    Naselli, Flores; Tesoriere, Luisa; Caradonna, Fabio; Bellavia, Daniele; Attanzio, Alessandro; Gentile, Carla; Livrea, Maria A

    2014-07-18

    Phytochemicals may exert chemo-preventive effects on cells of the gastro-intestinal tract by modulating epigenome-regulated gene expression. The effect of the aqueous extract from the edible fruit of Opuntia ficus-indica (OFI extract), and of its betalain pigment indicaxanthin (Ind), on proliferation of human colon cancer Caco-2 cells has been investigated. Whole extract and Ind caused a dose-dependent apoptosis of proliferating cells at nutritionally relevant amounts, with IC50 400±25 mg fresh pulp equivalents/mL, and 115±15 μM (n=9), respectively, without toxicity for post-confluent differentiated cells. Ind accounted for ∼80% of the effect of the whole extract. Ind did not cause oxidative stress in proliferating Caco-2 cells. Epigenomic activity of Ind was evident as de-methylation of the tumor suppressor p16(INK4a) gene promoter, reactivation of the silenced mRNA expression and accumulation of p16(INK4a), a major controller of cell cycle. As a consequence, decrease of hyper-phosphorylated, in favor of the hypo-phosphorylated retinoblastoma was observed, with unaltered level of the cycline-dependent kinase CDK4. Cell cycle showed arrest in the G2/M-phase. Dietary cactus pear fruit and Ind may have chemo-preventive potential in intestinal cells. Copyright © 2014 Elsevier Inc. All rights reserved.

  12. Genetic analysis of suppressors of the PF10 mutation in Chlamydomonas reinhardtii

    International Nuclear Information System (INIS)

    Dutcher, S.K.; Gibbons, W.; Inwood, W.B.

    1988-01-01

    A mutation at the PF10 locus of the unicellular green alga Chlamydomonas reinhardtii leads to abnormal cell motility. The asymmetric form of the ciliary beat stroke characteristic of wild-type flagella is modified by this mutation to a nearly symmetric beat. We report here that this abnormal motility is a conditional phenotype that depends on light intensity. In the absence of light or under low light intensities, the motility is more severely impaired than at higher light intensities. By UV mutagenesis we obtained 11 intragenic and 70 extragenic strains that show reversion of the pf10 motility phenotype observed in low light. The intragenic events reverted the motility phenotype of the pf10 mutation completely. The extragenic events define at least seven suppressor loci; these map to linkage groups IV, VII, IX, XI, XII and XVII. Suppressor mutations at two of the seven loci (LIS1 and LIS2) require light for their suppressor activity. Forty-eight of the 70 extragenic suppressors were examined in heterozygous diploid cells; 47 of these mutants were recessive to the wild-type allele and one mutant (bop5-1) was dominant to the wild-type allele. Complementation analysis of the 47 recessive mutants showed unusual patterns. Most mutants within a recombinationally defined group failed to complement one another, although there were pairs that showed intra-allelic complementation. Additionally, some of the mutants at each recombinationally defined locus failed to complement mutants at other loci. They define dominant enhancers of one another

  13. Anti-proliferative and pro-apoptotic activity of whole extract and isolated indicaxanthin from Opuntia ficus-indica associated with re-activation of the onco-suppressor p16{sup INK4a} gene in human colorectal carcinoma (Caco-2) cells

    Energy Technology Data Exchange (ETDEWEB)

    Naselli, Flores; Tesoriere, Luisa; Caradonna, Fabio; Bellavia, Daniele; Attanzio, Alessandro; Gentile, Carla; Livrea, Maria A., E-mail: maria.livrea@unipa.it

    2014-07-18

    Highlights: • Cactus pear fruit extract and indicaxanthin cause apoptosis of colon cancer cells. • Indicaxanthin does not cause ROS formation, but affects epigenoma in Caco-2 cells. • Indicaxanthin reverses methylation of oncosuppressor p16{sup INK4a} gene in Caco-2 cells. • Indicaxanthin reactivates retinoblastoma in Caco-2 cells. • Bioavailable indicaxanthin may have chemopreventive activity in colon cancer. - Abstract: Phytochemicals may exert chemo-preventive effects on cells of the gastro-intestinal tract by modulating epigenome-regulated gene expression. The effect of the aqueous extract from the edible fruit of Opuntia ficus-indica (OFI extract), and of its betalain pigment indicaxanthin (Ind), on proliferation of human colon cancer Caco-2 cells has been investigated. Whole extract and Ind caused a dose-dependent apoptosis of proliferating cells at nutritionally relevant amounts, with IC{sub 50} 400 ± 25 mg fresh pulp equivalents/mL, and 115 ± 15 μM (n = 9), respectively, without toxicity for post-confluent differentiated cells. Ind accounted for ∼80% of the effect of the whole extract. Ind did not cause oxidative stress in proliferating Caco-2 cells. Epigenomic activity of Ind was evident as de-methylation of the tumor suppressor p16{sup INK4a} gene promoter, reactivation of the silenced mRNA expression and accumulation of p16{sup INK4a}, a major controller of cell cycle. As a consequence, decrease of hyper-phosphorylated, in favor of the hypo-phosphorylated retinoblastoma was observed, with unaltered level of the cycline-dependent kinase CDK4. Cell cycle showed arrest in the G2/M-phase. Dietary cactus pear fruit and Ind may have chemo-preventive potential in intestinal cells.

  14. Anti-proliferative and pro-apoptotic activity of whole extract and isolated indicaxanthin from Opuntia ficus-indica associated with re-activation of the onco-suppressor p16INK4a gene in human colorectal carcinoma (Caco-2) cells

    International Nuclear Information System (INIS)

    Naselli, Flores; Tesoriere, Luisa; Caradonna, Fabio; Bellavia, Daniele; Attanzio, Alessandro; Gentile, Carla; Livrea, Maria A.

    2014-01-01

    Highlights: • Cactus pear fruit extract and indicaxanthin cause apoptosis of colon cancer cells. • Indicaxanthin does not cause ROS formation, but affects epigenoma in Caco-2 cells. • Indicaxanthin reverses methylation of oncosuppressor p16 INK4a gene in Caco-2 cells. • Indicaxanthin reactivates retinoblastoma in Caco-2 cells. • Bioavailable indicaxanthin may have chemopreventive activity in colon cancer. - Abstract: Phytochemicals may exert chemo-preventive effects on cells of the gastro-intestinal tract by modulating epigenome-regulated gene expression. The effect of the aqueous extract from the edible fruit of Opuntia ficus-indica (OFI extract), and of its betalain pigment indicaxanthin (Ind), on proliferation of human colon cancer Caco-2 cells has been investigated. Whole extract and Ind caused a dose-dependent apoptosis of proliferating cells at nutritionally relevant amounts, with IC 50 400 ± 25 mg fresh pulp equivalents/mL, and 115 ± 15 μM (n = 9), respectively, without toxicity for post-confluent differentiated cells. Ind accounted for ∼80% of the effect of the whole extract. Ind did not cause oxidative stress in proliferating Caco-2 cells. Epigenomic activity of Ind was evident as de-methylation of the tumor suppressor p16 INK4a gene promoter, reactivation of the silenced mRNA expression and accumulation of p16 INK4a , a major controller of cell cycle. As a consequence, decrease of hyper-phosphorylated, in favor of the hypo-phosphorylated retinoblastoma was observed, with unaltered level of the cycline-dependent kinase CDK4. Cell cycle showed arrest in the G2/M-phase. Dietary cactus pear fruit and Ind may have chemo-preventive potential in intestinal cells

  15. Effect of hemopoietic microenvironment on splenic suppressor macrophages in congenitally anemic mice of genotype Sl/Sld

    International Nuclear Information System (INIS)

    Shibata, Y.; Volkman, A.

    1985-01-01

    Mechanisms underlying mononuclear phagocyte specialization are being probed by studying suppressor macrophages (M phi) as a reference population in mouse models with impaired blood monocyte formation. Splenic suppressor M phi, defined by PGE-mediated inhibition of Con A-induced T lymphocyte proliferation are induced by the i.p. administration of Corynebacterium parvum (CP). Mice severely depleted of bone marrow and blood monocytes by treatment with 89Sr fail to show this suppressor M phi response to CP, although M phi-forming stem cells, assessed as splenic M-CFC in vitro, are increased 20-fold. These observations suggest that radiosensitive bone marrow stem cells are necessary for the generation of both suppressor M phi and monocytes and that one such stem cell may be common to both types of mononuclear phagocytes. This notion was explored further by employing congenitally anemic mice of the genotype S1/S1d in which the hemopoietic microenvironment is genetically defective and thus unable to support the proliferation, differentiation, and function of stem cells. The congenital defect was found to be additionally expressed in the S1/S1d mouse by a monocytopenia of less than 10% of the values in normal congenic littermate controls and by the failure of splenic M-CFC to increase in response to CP. PGE-producing suppressor M phi expressing Fc gamma 2b receptors, however, were induced by CP in S1/S1d mice with no significant diminution of suppressor activity. These data establish the fact that significant impairment of the formation of monocytes is part of the overall hemopoietic defect in S1/S1d mice. PGE-producing suppressor M phi, however, were inducible at normal functional levels in the presence of a profound monocytopenia, and therefore appear to be independent of the mechanisms that regulate blood monocyte formation

  16. Infrared suppressor effect on T63 turboshaft engine performance

    Science.gov (United States)

    Bailey, E. E.; Civinskas, K. C.; Walker, C. L.

    1978-01-01

    Tests were conducted to determine if there are performance penalties associated with the installation of infrared (IR) suppressors on the T63-A-700 turboshaft engine. The testing was done in a sea-level, static test cell. The same engine (A-E402808 B) was run with the standard OH-58 aircraft exhaust stacks and with the ejector-type IR suppressors in order to make a valid comparison. Repeatability of the test results for the two configurations was verified by rerunning the conditions over a period of days. Test results showed no measurable difference in performance between the standard exhaust stacks and the IR suppressors.

  17. Identification and Functional Analysis of Gene Regulatory Sequences Interacting with Colorectal Tumor Suppressors

    DEFF Research Database (Denmark)

    Dahlgaard, Katja; Troelsen, Jesper

    2018-01-01

    Several tumor suppressors possess gene regulatory activity. Here, we describe how promoter and promoter/enhancer reporter assays can be used to characterize a colorectal tumor suppressor proteins’ gene regulatory activity of possible target genes. In the first part, a bioinformatic approach...... of the quick and efficient In-Fusion cloning method, and how to carry out transient transfections of Caco-2 colon cancer cells with the produced luciferase reporter plasmids using polyethyleneimine (PEI). A plan describing how to set up and carry out the luciferase expression assay is presented. The luciferase...... to identify relevant gene regulatory regions of potential target genes is presented. In the second part, it is demonstrated how to prepare and carry out the functional assay. We explain how to clone the bioinformatically identified gene regulatory regions into luciferase reporter plasmids by the use...

  18. The Ras effector RASSF2 is a novel tumor-suppressor gene in human colorectal cancer.

    Science.gov (United States)

    Akino, Kimishige; Toyota, Minoru; Suzuki, Hiromu; Mita, Hiroaki; Sasaki, Yasushi; Ohe-Toyota, Mutsumi; Issa, Jean-Pierre J; Hinoda, Yuji; Imai, Kohzoh; Tokino, Takashi

    2005-07-01

    Activation of Ras signaling is a hallmark of colorectal cancer (CRC), but the roles of negative regulators of Ras are not fully understood. Our aim was to address that question by surveying genetic and epigenetic alterations of Ras-Ras effector genes in CRC cells. The expression and methylation status of 6 RASSF family genes were examined using RT-PCR and bisulfite PCR in CRC cell lines and in primary CRCs and colorectal adenomas. Colony formation assays and flow cytometry were used to assess the tumor suppressor activities of RASSF1 and RASSF2. Immunofluorescence microscopy was used to determine the effect of altered RASSF2 expression on cell morphology. Mutations of K- ras , BRAF, and p53 were identified using single-strand conformation analysis and direct sequencing. Aberrant methylation and histone deacetylation of RASSF2 was associated with the gene's silencing in CRC. The activities of RASSF2, which were distinct from those of RASSF1, included induction of morphologic changes and apoptosis; moreover, its ability to prevent cell transformation suggests that RASSF2 acts as a tumor suppressor in CRC. Primary CRCs that showed K- ras /BRAF mutations also frequently showed RASSF2 methylation, and inactivation of RASSF2 enhanced K- ras -induced oncogenic transformation. RASSF2 methylation was also frequently identified in colorectal adenomas. RASSF2 is a novel tumor suppressor gene that regulates Ras signaling and plays a pivotal role in the early stages of colorectal tumorigenesis.

  19. Transducer of ERBB2.1 (TOB1) as a Tumor Suppressor: A Mechanistic Perspective.

    Science.gov (United States)

    Lee, Hun Seok; Kundu, Juthika; Kim, Ryong Nam; Shin, Young Kee

    2015-12-15

    Transducer of ERBB2.1 (TOB1) is a tumor-suppressor protein, which functions as a negative regulator of the receptor tyrosine-kinase ERBB2. As most of the other tumor suppressor proteins, TOB1 is inactivated in many human cancers. Homozygous deletion of TOB1 in mice is reported to be responsible for cancer development in the lung, liver, and lymph node, whereas the ectopic overexpression of TOB1 shows anti-proliferation, and a decrease in the migration and invasion abilities on cancer cells. Biochemical studies revealed that the anti-proliferative activity of TOB1 involves mRNA deadenylation and is associated with the reduction of both cyclin D1 and cyclin-dependent kinase (CDK) expressions and the induction of CDK inhibitors. Moreover, TOB1 interacts with an oncogenic signaling mediator, β-catenin, and inhibits β-catenin-regulated gene transcription. TOB1 antagonizes the v-akt murine thymoma viral oncogene (AKT) signaling and induces cancer cell apoptosis by activating BCL2-associated X (BAX) protein and inhibiting the BCL-2 and BCL-XL expressions. The tumor-specific overexpression of TOB1 results in the activation of other tumor suppressor proteins, such as mothers against decapentaplegic homolog 4 (SMAD4) and phosphatase and tensin homolog-10 (PTEN), and blocks tumor progression. TOB1-overexpressing cancer cells have limited potential of growing as xenograft tumors in nude mice upon subcutaneous implantation. This review addresses the molecular basis of TOB1 tumor suppressor function with special emphasis on its regulation of intracellular signaling pathways.

  20. Transducer of ERBB2.1 (TOB1 as a Tumor Suppressor: A Mechanistic Perspective

    Directory of Open Access Journals (Sweden)

    Hun Seok Lee

    2015-12-01

    Full Text Available Transducer of ERBB2.1 (TOB1 is a tumor-suppressor protein, which functions as a negative regulator of the receptor tyrosine-kinase ERBB2. As most of the other tumor suppressor proteins, TOB1 is inactivated in many human cancers. Homozygous deletion of TOB1 in mice is reported to be responsible for cancer development in the lung, liver, and lymph node, whereas the ectopic overexpression of TOB1 shows anti-proliferation, and a decrease in the migration and invasion abilities on cancer cells. Biochemical studies revealed that the anti-proliferative activity of TOB1 involves mRNA deadenylation and is associated with the reduction of both cyclin D1 and cyclin-dependent kinase (CDK expressions and the induction of CDK inhibitors. Moreover, TOB1 interacts with an oncogenic signaling mediator, β-catenin, and inhibits β-catenin-regulated gene transcription. TOB1 antagonizes the v-akt murine thymoma viral oncogene (AKT signaling and induces cancer cell apoptosis by activating BCL2-associated X (BAX protein and inhibiting the BCL-2 and BCL-XL expressions. The tumor-specific overexpression of TOB1 results in the activation of other tumor suppressor proteins, such as mothers against decapentaplegic homolog 4 (SMAD4 and phosphatase and tensin homolog-10 (PTEN, and blocks tumor progression. TOB1-overexpressing cancer cells have limited potential of growing as xenograft tumors in nude mice upon subcutaneous implantation. This review addresses the molecular basis of TOB1 tumor suppressor function with special emphasis on its regulation of intracellular signaling pathways.

  1. About hidden influence of predictor variables: Suppressor and mediator variables

    Directory of Open Access Journals (Sweden)

    Milovanović Boško

    2013-01-01

    Full Text Available In this paper procedure for researching hidden influence of predictor variables in regression models and depicting suppressor variables and mediator variables is shown. It is also shown that detection of suppressor variables and mediator variables could provide refined information about the research problem. As an example for applying this procedure, relation between Atlantic atmospheric centers and air temperature and precipitation amount in Serbia is chosen. [Projekat Ministarstva nauke Republike Srbije, br. 47007

  2. Activation of Akt is essential for the propagation of mitochondrial respiratory stress signaling and activation of the transcriptional coactivator heterogeneous ribonucleoprotein A2.

    Science.gov (United States)

    Guha, Manti; Fang, Ji-Kang; Monks, Robert; Birnbaum, Morris J; Avadhani, Narayan G

    2010-10-15

    Mitochondrial respiratory stress (also called mitochondrial retrograde signaling) activates a Ca(2+)/calcineurin-mediated signal that culminates in transcription activation/repression of a large number of nuclear genes. This signal is propagated through activation of the regulatory proteins NFκB c-Rel/p50, C/EBPδ, CREB, and NFAT. Additionally, the heterogeneous ribonucleoprotein A2 (hnRNPA2) functions as a coactivator in up-regulating the transcription of Cathepsin L, RyR1, and Glut-4, the target genes of stress signaling. Activation of IGF1R, which causes a metabolic switch to glycolysis, cell invasiveness, and resistance to apoptosis, is a phenotypic hallmark of C2C12 myoblasts subjected to mitochondrial stress. In this study, we report that mitochondrial stress leads to increased expression, activation, and nuclear localization of Akt1. Mitochondrial respiratory stress also activates Akt1-gene expression, which involves hnRNPA2 as a coactivator, indicating a complex interdependency of these two factors. Using Akt1(-/-) mouse embryonic fibroblasts and Akt1 mRNA-silenced C2C12 cells, we show that Akt1-mediated phosphorylation is crucial for the activation and recruitment of hnRNPA2 to the enhanceosome complex. Akt1 mRNA silencing in mtDNA-depleted cells resulted in reversal of the invasive phenotype, accompanied by sensitivity to apoptotic stimuli. These results show that Akt1 is an important regulator of the nuclear transcriptional response to mitochondrial stress.

  3. SIRT3: Oncogene and Tumor Suppressor in Cancer

    Directory of Open Access Journals (Sweden)

    Margalida Torrens-Mas

    2017-07-01

    Full Text Available Sirtuin 3 (SIRT3, the major deacetylase in mitochondria, plays a crucial role in modulating oxygen reactive species (ROS and limiting the oxidative damage in cellular components. SIRT3 targets different enzymes which regulate mitochondrial metabolism and participate in ROS detoxification, such as the complexes of the respiratory chain, the isocitrate dehydrogenase, or the manganese superoxide dismutase. Thus, SIRT3 activity is essential in maintaining mitochondria homeostasis and has recently received great attention, as it is considered a fidelity protein for mitochondrial function. In some types of cancer, SIRT3 functions as a tumoral promoter, since it keeps ROS levels under a certain threshold compatible with cell viability and proliferation. On the contrary, other studies describe SIRT3 as a tumoral suppressor, as SIRT3 could trigger cell death under stress conditions. Thus, SIRT3 could have a dual role in cancer. In this regard, modulation of SIRT3 activity could be a new target to develop more personalized therapies against cancer.

  4. Myeloid-derived suppressor cells in breast cancer.

    Science.gov (United States)

    Markowitz, Joseph; Wesolowski, Robert; Papenfuss, Tracey; Brooks, Taylor R; Carson, William E

    2013-07-01

    Myeloid-derived suppressor cells (MDSCs) are a population of immature myeloid cells defined by their suppressive actions on immune cells such as T cells, dendritic cells, and natural killer cells. MDSCs typically are positive for the markers CD33 and CD11b but express low levels of HLADR in humans. In mice, MDSCs are typically positive for both CD11b and Gr1. These cells exert their suppressive activity on the immune system via the production of reactive oxygen species, arginase, and cytokines. These factors subsequently inhibit the activity of multiple protein targets such as the T cell receptor, STAT1, and indoleamine-pyrrole 2,3-dioxygenase. The numbers of MDSCs tend to increase with cancer burden while inhibiting MDSCs improves disease outcome in murine models. MDSCs also inhibit immune cancer therapeutics. In light of the poor prognosis of metastatic breast cancer in women and the correlation of increasing levels of MDSCs with increasing disease burden, the purposes of this review are to (1) discuss why MDSCs may be important in breast cancer, (2) describe model systems used to study MDSCs in vitro and in vivo, (3) discuss mechanisms involved in MDSC induction/function in breast cancer, and (4) present pre-clinical and clinical studies that explore modulation of the MDSC-immune system interaction in breast cancer. MDSCs inhibit the host immune response in breast cancer patients and diminishing MDSC actions may improve therapeutic outcomes.

  5. Inhibition of tumor growth in syngenetic chimeric mice mediated by a depletion of suppressor T cells

    International Nuclear Information System (INIS)

    Rotter, V.; Trainin, N.

    1975-01-01

    Syngeneic chimeric (lethally irradiated and reconstituted with syngeneic bone marrow cells) mice manifested an increased resistance to the development of Lewis lung carcinoma. In addition, these mice had a higher response to polyvinylpyrrolidone and a reduced reactivity to T mitogens. The present findings suggest that syngeneic chimeric mice lack suppressor T cells shown to regulate the development of Lewis lung tumor and the response to polyvinylpyrrolidone. Other components of the T cell population, such as helper cells responding to sheep red blood cells or cells involved in allograft rejection, assayed in these syngeneic chimeras were found unaffected. The fact that chimeric mice are deficient in a certain suppressor T cell population whereas other T activities are normal suggests the existence of different cell lines within the T cell population. (U.S.)

  6. Tumor suppressor WWOX and p53 alterations and drug resistance in glioblastomas

    Directory of Open Access Journals (Sweden)

    Ming-Fu eChiang

    2013-03-01

    Full Text Available Tumor suppressor p53 are frequently mutated in glioblastomas (GBMs and appears to contribute, in part, to resistance to temozolomide and therapeutic drugs. WW domain-containing oxidoreductase WWOX (FOR or WOX1 is a proapoptotic protein and is considered as a tumor suppressor. Loss of WWOX gene expression is frequently seen in malignant cancer cells due to promoter hypermethylation, genetic alterations, and translational blockade. Intriguingly, ectopic expression of wild type WWOX preferentially induces apoptosis in human glioblastoma cells harboring mutant p53. WWOX is known to physically bind and stabilize wild type p53. Here, we provide an overview for the updated knowledge in p53 and WWOX, and postulate a potential scenarios that wild type and mutant p53, or isoforms, modulate the apoptotic function of WWOX. We propose that triggering WWOX activation by therapeutic drugs under p53 functional deficiency is needed to overcome TMZ resistance and induce GBM cell death.

  7. KLF10, transforming growth factor-{beta}-inducible early gene 1, acts as a tumor suppressor

    Energy Technology Data Exchange (ETDEWEB)

    Song, Ki-Duk [Center for Agricultural Biomaterials, Seoul National University, Seoul 151-921 (Korea, Republic of); Laboratory of Protein Engineering and Comparative Immunology, School of Agricultural Biotechnology, Seoul National University, Seoul 151-921 (Korea, Republic of); Kim, Duk-Jung [The Institute of Hankook Life Science, 7-9 Myungryun-dong, Jongno-gu, Seoul 110-521 (Korea, Republic of); Lee, Jong Eun [Department of Anatomy, College of Medicine, Yonsei University, Seoul 120-752 (Korea, Republic of); Yun, Cheol-Heui [Center for Agricultural Biomaterials, Seoul National University, Seoul 151-921 (Korea, Republic of); Laboratory of Protein Engineering and Comparative Immunology, School of Agricultural Biotechnology, Seoul National University, Seoul 151-921 (Korea, Republic of); Lee, Woon Kyu, E-mail: wklee@inha.ac.kr [Laboratory of Developmental Genetics, School of Medicine, Inha University, Incheon 400-712 (Korea, Republic of); Brain Korea 21 Center for Advanced Medical Education, School of Medicine, Inha University, Incheon 400-712 (Korea, Republic of)

    2012-03-09

    Highlights: Black-Right-Pointing-Pointer KLF10{sup -/-} mice exhibited accelerated papilloma development after DMBA/TPA treatment. Black-Right-Pointing-Pointer KLF10{sup -/-} keratinocytes showed increased proliferation and apoptosis. Black-Right-Pointing-Pointer KLF10{sup -/-} MEFs yielded more colonies than wild-type one with H-Ras transfection. Black-Right-Pointing-Pointer KLF10 dose-dependently activated p21{sup WAF1/CIP1} transcription. Black-Right-Pointing-Pointer KLF10 is a tumor suppressor and that it targets p21{sup WAF1/CIP1} transcription. -- Abstract: Krueppel-like factor 10 (KLF10) has been suggested to be a putative tumor suppressor. In the present study, we generated KLF10 deficient mice to explore this hypothesis in vivo. KLF10 deficient mice exhibited increased predisposition to skin tumorigenesis and markedly accelerated papilloma development after DMBA/TPA treatment. On the other hand, KLF10 deficient keratinocytes showed increased proliferation and apoptosis. In colony formation assays after oncogenic H-Ras transfection, KLF10 deficient mouse embryonic fibroblasts (MEFs) yielded more colonies than wild-type MEFs. Furthermore, KLF10 dose-dependently activated p21{sup WAF1/CIP1} transcription, which was independent of p53 and Sp1 binding sites in p21{sup WAF1/CIP1} promoter. This study demonstrates that KLF10 is a tumor suppressor and that it targets p21{sup WAF1/CIP1} transcription.

  8. Tumor Suppressor Function of CYLD in Nonmelanoma Skin Cancer

    Directory of Open Access Journals (Sweden)

    K. C. Masoumi

    2011-01-01

    Full Text Available Ubiquitin and ubiquitin-related proteins posttranslationally modify substrates, and thereby alter the functions of their targets. The ubiquitination process is involved in various physiological responses, and dysregulation of components of the ubiquitin system has been linked to many diseases including skin cancer. The ubiquitin pathways activated among skin cancers are highly diverse and may reflect the various characteristics of the cancer type. Basal cell carcinoma and squamous cell carcinoma, the most common types of human skin cancer, are instances where the involvement of the deubiquitination enzyme CYLD has been recently highlighted. In basal cell carcinoma, the tumor suppressor protein CYLD is repressed at the transcriptional levels through hedgehog signaling pathway. Downregulation of CYLD in basal cell carcinoma was also shown to interfere with TrkC expression and signaling, thereby promoting cancer progression. By contrast, the level of CYLD is unchanged in squamous cell carcinoma, instead, catalytic inactivation of CYLD in the skin has been linked to the development of squamous cell carcinoma. This paper will focus on the current knowledge that links CYLD to nonmelanoma skin cancers and will explore recent insights regarding CYLD regulation of NF-κB and hedgehog signaling during the development and progression of these types of human tumors.

  9. Melanoma Suppressor Functions of the Carcinoma Oncogene FOXQ1

    Directory of Open Access Journals (Sweden)

    Archis Bagati

    2017-09-01

    Full Text Available Lineage-specific regulation of tumor progression by the same transcription factor is understudied. We find that levels of the FOXQ1 transcription factor, an oncogene in carcinomas, are decreased during melanoma progression. Moreover, in contrast to carcinomas, FOXQ1 suppresses epithelial-to-mesenchymal transition, invasion, and metastasis in melanoma cells. We find that these lineage-specific functions of FOXQ1 largely depend on its ability to activate (in carcinomas or repress (in melanoma transcription of the N-cadherin gene (CDH2. We demonstrate that FOXQ1 interacts with nuclear β-catenin and TLE proteins, and the β-catenin/TLE ratio, which is higher in carcinoma than melanoma cells, determines the effect of FOXQ1 on CDH2 transcription. Accordingly, other FOXQ1-dependent phenotypes can be manipulated by altering nuclear β-catenin or TLE proteins levels. Our data identify FOXQ1 as a melanoma suppressor and establish a mechanism underlying its inverse lineage-specific transcriptional regulation of transformed phenotypes.

  10. Tumor Suppressor Function of CYLD in Non melanoma Skin Cancer

    International Nuclear Information System (INIS)

    Masoumi, K. C.; Hallgren, G. S.; Massoumi, R.

    2011-01-01

    Ubiquitin and ubiquitin-related proteins post translationally modify substrates, and thereby alter the functions of their targets. The ubiquitination process is involved in various physiological responses, and dysregulation of components of the ubiquitin system has been linked to many diseases including skin cancer. The ubiquitin pathways activated among skin cancers are highly diverse and may reflect the various characteristics of the cancer type. Basal cell carcinoma and squamous cell carcinoma, the most common types of human skin cancer, are instances where the involvement of the deubiquitination enzyme CYLD has been recently highlighted. In basal cell carcinoma, the tumor suppressor protein CYLD is repressed at the transcriptional levels through hedgehog signaling pathway. Downregulation of CYLD in basal cell carcinoma was also shown to interfere with TrkC expression and signaling, thereby promoting cancer progression. By contrast, the level of CYLD is unchanged in squamous cell carcinoma, instead, catalytic inactivation of CYLD in the skin has been linked to the development of squamous cell carcinoma. This paper will focus on the current knowledge that links CYLD to non melanoma skin cancers and will explore recent insights regarding CYLD regulation of NF-κB and hedgehog signaling during the development and progression of these types of human tumors.

  11. The LKB1 tumor suppressor differentially affects anchorage independent growth of HPV positive cervical cancer cell lines

    International Nuclear Information System (INIS)

    Mack, Hildegard I.D.; Munger, Karl

    2013-01-01

    Infection with high-risk human papillomaviruses is causally linked to cervical carcinogenesis. However, most lesions caused by high-risk HPV infections do not progress to cancer. Host cell mutations contribute to malignant progression but the molecular nature of such mutations is unknown. Based on a previous study that reported an association between liver kinase B1 (LKB1) tumor suppressor loss and poor outcome in cervical cancer, we sought to determine the molecular basis for this observation. LKB1-negative cervical and lung cancer cells were reconstituted with wild type or kinase defective LKB1 mutants and we examined the importance of LKB1 catalytic activity in known LKB1-regulated processes including inhibition of cell proliferation and elevated resistance to energy stress. Our studies revealed marked differences in the biological activities of two kinase defective LKB1 mutants in the various cell lines. Thus, our results suggest that LKB1 may be a cell-type specific tumor suppressor. - Highlights: • LKB1 is a tumor suppressor that is linked to Peutz-Jeghers syndrome. • Peutz-Jeghers syndrome patients have a high incidence of cervical cancer. • Cervical cancer is caused by HPV infections. • This study investigates LKB1 tumor suppressor activity in cervical cancer

  12. Effect of bone marrow depletion on prostaglandin E-producing suppressor macrophages in mouse spleen

    International Nuclear Information System (INIS)

    Shibata, Y.; Volkman, A.

    1985-01-01

    The i.p. injection of Corynebacterium parvum (CP) into CBA/J mice effected increases in macrophage colony-forming cells (M-CFC) when spleen cells were cultured with L cell culture filtrate as a source of colony-stimulating factor. Significant increases in phagocytic macrophages (M phi) with Fc receptors for IgG2a and IgG2b immune complexes were additionally noted among the spleen cells in these mice. These M phi effectively inhibited Con A-induced lymphocyte proliferation, probably reflecting a 10-fold increase above normal controls in prostaglandin E to 47 ng/3 X 10(6) spleen cells/ml. To determine whether the suppressor M phi are immediate derivatives of splenic M-CFC, we tried to induce suppressor M phi by the injection of CP into mice depleted of bone marrow M-CFC by the earlier administration of the bone-seeking isotope, 89Sr. This procedure reduced M-CFC in the bone marrow to less than 1% of normal for more than 30 days. Monocytes in the blood fell to 5% of normal by day 10 and were 30% on day 30. Levels of resident peritoneal M phi showed relatively little change in this period. By contrast, splenic M-CFC increased to 20-fold higher than the cold 88Sr controls. CP-induced suppressor M phi activity, however, was sharply reduced in 89Sr marrow-depleted mice on day 10, despite the striking increase in M-CFC. There was a threefold increase in the number of phagocytic M phi binding IgG2a immune complexes, with no significant increase in IgG2b binding M phi. The kinetics of recovery of suppressor M phi activity showed that on days 20, 30, and 50 after 89Sr injection the activities reached 20%, 30%, and 70% of the cold control, respectively, and correlated with the recovery of significant levels of M-CFC in the bone marrow. Taken together, these observations suggest that splenic M-CFC are not an immediate source of PGE-suppressor M phi in vivo

  13. Control of polyclonal immunoglobulin production from human lymphocytes by leukotrienes; leukotriene B4 induces an OKT8(+), radiosensitive suppressor cell from resting, human OKT8(-) T cells

    International Nuclear Information System (INIS)

    Atluru, D.; Goodwin, J.S.

    1984-01-01

    We report that leukotriene B4 (LTB4), a 5-lipoxygenase metabolite of arachidonic acid, is a potent suppressor of polyclonal Ig production in pokeweed mitogen (PWM)-stimulated cultures of human peripheral blood lymphocytes, while LTC4 and LTD4 have little activity in this system. Preincubation of T cells with LTB4 in nanomolar to picomolar concentrations rendered these cells suppressive of Ig production in subsequent PWM-stimulated cultures of fresh, autologous B + T cells. This LTB4-induced suppressor cell was radiosensitive, and its generation could be blocked by cyclohexamide but not by mitomycin C. The LTB4-induced suppressor cell was OKT8(+), while the precursor for the cell could be OKT8(-). The incubation of OKT8(-) T cells with LTB4 for 18 h resulted in the appearance of the OKT8(+) on 10-20% of the cells, and this could be blocked by cyclohexamide but not by mitomycin C. Thus, LTB4 in very low concentrations induces a radiosensitive OKT8(+) suppressor cell from OKT8(-) cells. In this regard, LTB4 is three to six orders of magnitude more potent than any endogenous hormonal inducer of suppressor cells previously described. Glucocorticosteroids, which block suppressor cell induction in many systems, may act by inhibiting endogenous production of LTB4

  14. Effect of irradiation on human T-cell proliferation: low dose irradiation stimulates mitogen-induced proliferation and function of the suppressor/cytotoxic T-cell subset

    International Nuclear Information System (INIS)

    Gualde, N.; Goodwin, J.S.

    1984-01-01

    Unfractionated human T cells exposed to 10-50 rad of X irradiation incorporated less [ 3 H]thymidine than nonirradiated T cells when subsequently cultured with PHA or Con A. The cytotoxic/suppressor T-cell subset, isolated as either OKT8(+) or OKT4(-) cells, demonstrated significantly enhanced [ 3 H]thymidine incorporation in PHA- or Con A-stimulated cultures after exposure to 10-50 rad, compared to unirradiated cells, while the proliferation of the OKT4(+) helper/inducer subset was inhibited by low dose irradiation. It has been previously reported that approximately 30% of the cytotoxic/suppressor subset also stains with OKM1. When the cytotoxic/suppressor subset was further subdivided into OKT4(-), OKM1(+), and OKT4(-), OKM1(-) cells, proliferation of the OKT4(-), OKM1(+) population was inhibited by exposure to 25 rad while proliferation of the OKT4(-), OKM1(-) population was stimulated. The increase in proliferation of the cytotoxic/suppressor T-cell subset after low dose irradiation is paralleled by an increase in suppressor activity of these cells. T cells exposed to 25 rad and then cultured with Con A for 48 hr caused greater inhibition of IgG production when added to fresh autologous lymphocytes stimulated by pokeweed mitogen than did unirradiated cells. Thus, low dose irradiation enhances both the proliferation and function of the human suppressor T-cell subset

  15. BASP1 is a transcriptional cosuppressor for the Wilms' tumor suppressor protein WT1

    DEFF Research Database (Denmark)

    Carpenter, Brian; Hill, Kathryn J; Charalambous, Marika

    2004-01-01

    The Wilms' tumor suppressor protein WT1 is a transcriptional regulator that plays a key role in the development of the kidneys. The transcriptional activation domain of WT1 is subject to regulation by a suppression region within the N terminus of WT1. Using a functional assay, we provide direct...... evidence that this requires a transcriptional cosuppressor, which we identify as brain acid soluble protein 1 (BASP1). WT1 and BASP1 associate within the nuclei of cells that naturally express both proteins. BASP1 can confer WT1 cosuppressor activity in transfection assays, and elimination of endogenous...

  16. Adenovirus E4 open reading frame 4-induced dephosphorylation inhibits E1A activation of the E2 promoter and E2F-1-mediated transactivation independently of the retinoblastoma tumor suppressor protein

    DEFF Research Database (Denmark)

    Mannervik, M; Fan, S; Ström, A C

    1999-01-01

    of the viral E4 open reading frame 4 (E4-ORF4) protein. This effect does not to require the retinoblastoma protein that previously has been shown to regulate E2F activity. The inhibitory activity of E4-ORF4 appears to be specific because E4-ORF4 had little effect on, for example, E4-ORF6/7 transactivation......Previous studies have shown that the cell cycle-regulated E2F transcription factor is subjected to both positive and negative control by phosphorylation. Here we show that in transient transfection experiments, adenovirus E1A activation of the viral E2 promoter is abrogated by coexpression...... of the E2 promoter. We further show that the repressive effect of E4-ORF4 on E2 transcription works mainly through the E2F DNA-binding sites in the E2 promoter. In agreement with this, we find that E4-ORF4 inhibits E2F-1/DP-1-mediated transactivation. We also show that E4-ORF4 inhibits E2 mRNA expression...

  17. Nonspecific suppressor elements in murine allogeneic radiation chimeras

    International Nuclear Information System (INIS)

    Urso, P.; Gengozian, N.

    1979-01-01

    Spleen cells from long-term mouse allogeneic radiation chimeras were tested for their ability to modulate the graft-versus-host (GVH) or plaque-forming cell (PFC) response of normal lymphocytes transplanted in lethally x-irradiated recipients. In vivo GVH proliferation of normal lymphocytes (syngeneic to donor cells of the chimera) against antigens of host-type in which the chimeric state had been established was reduced by chimera cells. Inhibition varied, some chimeras suppressing GVH more than others and a few not suppressing at all. The suppressive effect was abrogated if the chimera cells were treated with anti-THETA; treatment with anti-IgM did not eliminate this activity. When mixtures of normal donor lymphocytes and chimera cells were given to irradiated recipients genetically different from host or donor, reduction of donor cell GVH also occurred. Further, chimera cells reduced the GVH activity of normal host cells in irradiated recipients differing from the host at one H-2 locus and from the donor at minor histocompatibility loci. The modulating effect of spleen cells from chimeras on the PFC response by normal lymphocytes also varied. Six chimeras induced a 25 to 90% suppression, two enhanced the response, and one showed no effect. Where suppression occurred, treatment of chimera cells with anti-THETA most often, but not always, restored PFC production. Our results show that the suppressive action of splenic lymphoid cells by chimeras is highly nonspecific and variable in expression. We suggest that tolerance in chimeras may be mediated by nonspecific suppressor elements leading to unresponsiveness to a variety of antigens including SRBC

  18. [Contact shot from infantry weapons with a flash-suppressor].

    Science.gov (United States)

    Perdekamp, Markus Grosse; Braunwarth, Roland; Schmidt, Ulrike; Schmidt, Wolfgang; Pollak, Stefan

    2003-01-01

    The number of reports on contact shots from firearms with a flash suppressor attached to the muzzle is small. On the basis of a case report (suicidal shot to the forehead with a Kalschnikow AKMS 47 assault rifle) the morphological peculiarities (characteristics soot pattern, relatively small powder cavity and only minor skin tears in the presence of a bony support) are presented and the conclusions to be drawn from the findings regarding the flash-suppressor, the shot distance, the angle of the shot and the way of holding the weapon are discussed.

  19. Suppressors of DnaAATP imposed overinitiation in Escherichia coli

    DEFF Research Database (Denmark)

    Charbon, Godefroid; Riber, Leise; Cohen, Malene

    2011-01-01

    Chromosome replication in Escherichia coli is limited by the supply of DnaA associated with ATP. Cells deficient in RIDA (Regulatory Inactivation of DnaA) due to a deletion of the hda gene accumulate suppressor mutations (hsm) to counteract the overinitiation caused by an elevated DnaAATP level....... Eight spontaneous hda suppressor mutations were identified by whole-genome sequencing, and three of these were analysed further. Two mutations (hsm-2 and hsm-4) mapped in the dnaA gene and led to a reduced ability to initiate replication from oriC. One mutation (hsm-1) mapped to the seqA promoter...

  20. Cervical Cancer Cell Supernatants Induce a Phenotypic Switch from U937-Derived Macrophage-Activated M1 State into M2-Like Suppressor Phenotype with Change in Toll-Like Receptor Profile

    Directory of Open Access Journals (Sweden)

    Karina Sánchez-Reyes

    2014-01-01

    Full Text Available Cervical cancer (CC is the second most common cancer among women worldwide. Infection with human papillomavirus (HPV is the main risk factor for developing CC. Macrophages are important immune effector cells; they can be differentiated into two phenotypes, identified as M1 (classically activated and M2 (alternatively activated. Macrophage polarization exerts profound effects on the Toll-like receptor (TLR profile. In this study, we evaluated whether the supernatant of human CC cells HeLa, SiHa, and C-33A induces a shift of M1 macrophage toward M2 macrophage in U937-derived macrophages. Results. The results showed that soluble factors secreted by CC cells induce a change in the immunophenotype of macrophages from macrophage M1 into macrophage M2. U937-derived macrophages M1 released proinflammatory cytokines and nitric oxide; however, when these cells were treated with the supernatant of CC cell lines, we observed a turnover of M1 toward M2. These cells increased CD163 and IL-10 expression. The expression of TLR-3, -7, and -9 is increased when the macrophages were treated with the supernatant of CC cells. Conclusions. Our result strongly suggests that CC cells may, through the secretion of soluble factors, induce a change of immunophenotype M1 into M2 macrophages.

  1. Δ9-Tetrahydrocannabinol attenuates allogeneic host-versus-graft response and delays skin graft rejection through activation of cannabinoid receptor 1 and induction of myeloid-derived suppressor cells

    Science.gov (United States)

    Sido, Jessica M.; Nagarkatti, Prakash S.; Nagarkatti, Mitzi

    2015-01-01

    Immune cells have been shown to express cannabinoid receptors and to produce endogenous ligands. Moreover, activation of cannabinoid receptors on immune cells has been shown to trigger potent immunosuppression. Despite such studies, the role of cannabinoids in transplantation, specifically to prevent allograft rejection, has not, to our knowledge, been investigated previously. In the current study, we tested the effect of THC on the suppression of HvGD as well as rejection of skin allografts. To this end, we studied HvGD by injecting H-2k splenocytes into H-2b mice and analyzing the immune response in the draining ingLNs. THC treatment significantly reduced T cell proliferation and activation in draining LNs of the recipient mice and decreased early stage rejection-indicator cytokines, including IL-2 and IFN-γ. THC treatment also increased the allogeneic skin graft survival. THC treatment in HvGD mice led to induction of MDSCs. Using MDSC depletion studies as well as adoptive transfer experiments, we found that THC-induced MDSCs were necessary for attenuation of HvGD. Additionally, using pharmacological inhibitors of CB1 and CB2 receptors and CB1 and CB2 knockout mice, we found that THC was working preferentially through CB1. Together, our research shows, for the first time to our knowledge, that targeting cannabinoid receptors may provide a novel treatment modality to attenuate HvGD and prevent allograft rejection. PMID:26034207

  2. Induction of specific suppressor T cells in vitro

    International Nuclear Information System (INIS)

    Eardley, D.D.; Gershon, R.K.

    1976-01-01

    We describe conditions for generating sheep red blood cell-specific suppressor T cells in Mishell-Dutton cultures. The production of specific suppressor cells is favored by increasing antigen dose in the initial culture but can be produced by transferring more cells when lower doses of antigen are used. Transfer of small numbers of cells cultured with low doses of antigen leads to a specific helper effect. Transfer of large numbers of educated cells leads to nonspecific suppression. Suppression can be effected by the effluent cells from nylon wool columns which do not make detectable PFC. A fraction of these cells become resistant to treatment with anti-T cell sera and complement after culture. The suppressor cells are radiation sensitive and must be able to synthesize protein to suppress. They take 2 to 3 days of education to reach maximum suppressive efficiency and will not suppress cultures if added 2 to 3 days after culture initiation. Their production is favored by the absence of mercaptoethanol, suggesting that the observed suppression is not ''too much help.'' The ability to generate specific suppressor cells in vitro should be of great benefit in determining the factors that regulate their appearance in vivo

  3. The tumor suppressor Rb and its related Rbl2 genes are regulated by Utx histone demethylase

    Energy Technology Data Exchange (ETDEWEB)

    Terashima, Minoru; Ishimura, Akihiko; Yoshida, Masakazu [Division of Functional Genomics, Cancer Research Institute, Kanazawa University, Kakuma-machi, Kanazawa 920-1192, Ishikawa (Japan); Suzuki, Yutaka; Sugano, Sumio [Graduate School of Frontier Sciences, The University of Tokyo, Kashiwa 277-8561, Chiba (Japan); Suzuki, Takeshi, E-mail: suzuki-t@staff.kanazawa-u.ac.jp [Division of Functional Genomics, Cancer Research Institute, Kanazawa University, Kakuma-machi, Kanazawa 920-1192, Ishikawa (Japan)

    2010-08-20

    Research highlights: {yields} Utx increases expression of Rb and Rbl2 genes through its demethylase activity. {yields} Utx changes histone H3 methylation on the Rb and Rbl2 promoters. {yields} Utx induces decreased cell proliferation of mammalian primary cells. -- Abstract: Utx is a candidate tumor suppressor gene that encodes histone H3 lysine 27 (H3K27) demethylase. In this study, we found that ectopic expression of Utx enhanced the expression of retinoblastoma tumor suppressor gene Rb and its related gene Rbl2. This activation was dependent on the demethylase activity of Utx, and was suggested to contribute to the decreased cell proliferation induced by Utx. A chromatin immunoprecipitation assay showed that over-expressed Utx was associated with the promoter regions of Rb and Rbl2 resulting in the removal of repressive H3K27 tri-methylation and the increase in active H3K4 tri-methylation. Furthermore, siRNA-mediated knockdown of Utx revealed the recruitment of endogenous Utx protein on the promoters of Rb and Rbl2 genes. These results indicate that Rb and Rbl2 are downstream target genes of Utx and may play important roles in Utx-mediated cell growth control.

  4. The PTPN14 Tumor Suppressor Is a Degradation Target of Human Papillomavirus E7.

    Science.gov (United States)

    Szalmás, Anita; Tomaić, Vjekoslav; Basukala, Om; Massimi, Paola; Mittal, Suruchi; Kónya, József; Banks, Lawrence

    2017-04-01

    Activation of signaling pathways ensuring cell growth is essential for the proliferative competence of human papillomavirus (HPV)-infected cells. Tyrosine kinases and phosphatases are key regulators of cellular growth control pathways. A recently identified potential cellular target of HPV E7 is the cytoplasmic protein tyrosine phosphatase PTPN14, which is a potential tumor suppressor and is linked to the control of the Hippo and Wnt/beta-catenin signaling pathways. In this study, we show that the E7 proteins of both high-risk and low-risk mucosal HPV types can interact with PTPN14. This interaction is independent of retinoblastoma protein (pRb) and involves residues in the carboxy-terminal region of E7. We also show that high-risk E7 induces proteasome-mediated degradation of PTPN14 in cells derived from cervical tumors. This degradation appears to be independent of cullin-1 or cullin-2 but most likely involves the UBR4/p600 ubiquitin ligase. The degree to which E7 downregulates PTPN14 would suggest that this interaction is important for the viral life cycle and potentially also for the development of malignancy. In support of this we find that overexpression of PTPN14 decreases the ability of HPV-16 E7 to cooperate with activated EJ-ras in primary cell transformation assays. IMPORTANCE This study links HPV E7 to the deregulation of protein tyrosine phosphatase signaling pathways. PTPN14 is classified as a potential tumor suppressor protein, and here we show that it is very susceptible to HPV E7-induced proteasome-mediated degradation. Intriguingly, this appears to use a mechanism that is different from that employed by E7 to target pRb. Therefore, this study has important implications for our understanding of the molecular basis for E7 function and also sheds important light on the potential role of PTPN14 as a tumor suppressor. Copyright © 2017 American Society for Microbiology.

  5. Murine neonatal spleen contains natural T and non-T suppressor cells capable of inhibiting adult alloreactive and newborn autoreactive T-cell proliferation.

    Science.gov (United States)

    Hooper, D C; Hoskin, D W; Gronvik, K O; Murgita, R A

    1986-05-01

    The spleen of neonatal mice is known to be a rich source of cells capable of suppressing a variety of immune functions of adult lymphocytes in vitro. From such observations has emerged the concept that the gradual development in ability to express immune functions after birth is due in part to the parallel normal physiological decay of naturally occurring regulatory suppressor cells. There is, however, some confusion in the literature as to the exact nature of the newborn of the newborn inhibitory cell type(s). In contrast to most previous reports which detect only a single type of neonatal suppressor cell, usually a T cell, we show here that newborn spleen harbors both T and non-T inhibitory cells. Both types of suppressor cells could be shown to suppress the proliferative response of adult spleen to alloantigens as well as newborn T cells reacting against self-Ia antigen in the autologous mixed lymphocyte reaction (AMLR). Newborn suppressor T cells were characterized as being non-adherent to Ig-anti-Ig affinity columns, soybean agglutinin receptor negative (SBA-), and susceptible to lysis by anti-T-cell specific antiserum plus complement. Non-T suppressor cells were identified as non-phagocytic, SBA receptor positive (SBA+), and resistant to cytotoxic treatment with anti-T-cell antibodies and complement. The apparent controversy surrounding previous reports as to the T versus non-T nature of newborn suppressor cells can be reconciled by the present observation that both types of inhibitory cells coexist in the spleen. Furthermore, the demonstration that newborn suppressor cells can effectively regulate T-cell proliferative activity mediated by other newborn cells provides more direct support for the contention that such inhibitory cells play a physiological role in controlling immune responsiveness during early ontogeny.

  6. SFRP Tumour Suppressor Genes Are Potential Plasma-Based Epigenetic Biomarkers for Malignant Pleural Mesothelioma

    Directory of Open Access Journals (Sweden)

    Yuen Yee Cheng

    2017-01-01

    Full Text Available Malignant pleural mesothelioma (MPM is associated with asbestos exposure. Asbestos can induce chronic inflammation which in turn can lead to silencing of tumour suppressor genes. Wnt signaling pathway can be affected by chronic inflammation and is aberrantly activated in many cancers including colon and MPM. SFRP genes are antagonists of Wnt pathway, and SFRPs are potential tumour suppressors in colon, gastric, breast, ovarian, and lung cancers and mesothelioma. This study investigated the expression and DNA methylation of SFRP genes in MPM cells lines with and without demethylation treatment. Sixty-six patient FFPE samples were analysed and have showed methylation of SFRP2 (56% and SFRP5 (70% in MPM. SFRP2 and SFRP5 tumour-suppressive activity in eleven MPM lines was confirmed, and long-term asbestos exposure led to reduced expression of the SFRP1 and SFRP2 genes in the mesothelium (MeT-5A via epigenetic alterations. Finally, DNA methylation of SFRPs is detectable in MPM patient plasma samples, with methylated SFRP2 and SFRP5 showing a tendency towards greater abundance in patients. These data suggested that SFRP genes have tumour-suppresive activity in MPM and that methylated DNA from SFRP gene promoters has the potential to serve as a biomarker for MPM patient plasma.

  7. An unusual characteristic “flower-like” pattern: flash suppressor burns

    OpenAIRE

    Gurcan, Altun

    2012-01-01

    The case on contact shots from firearms with a flash suppressor is rare. When a rifle fitted with a flash suppressor is fired, the emerging soot-laden gas in the barrel escapes from the slits of the flash suppressor. If the shot is contact or near contact, the flash suppressor will produce a characteristic “flower-like” pattern of seared, blackened zones around the entrance. This paper presents the injury pattern of the flash suppressor in a 29-year-old man who committed suicide with a G3 aut...

  8. An unusual characteristic “flower-like” pattern: flash suppressor burns

    Science.gov (United States)

    Gurcan, Altun

    2012-01-01

    The case on contact shots from firearms with a flash suppressor is rare. When a rifle fitted with a flash suppressor is fired, the emerging soot-laden gas in the barrel escapes from the slits of the flash suppressor. If the shot is contact or near contact, the flash suppressor will produce a characteristic “flower-like” pattern of seared, blackened zones around the entrance. This paper presents the injury pattern of the flash suppressor in a 29-year-old man who committed suicide with a G3 automatic infantry rifle. PMID:23935280

  9. Immunoregulatory T cells in man. Histamine-induced suppressor T cells are derived from a Leu 2+ (T8+) subpopulation distinct from that which gives rise to cytotoxic T cells

    International Nuclear Information System (INIS)

    Sansoni, P.; Silverman, E.D.; Khan, M.M.; Melmon, K.L.; Engleman, E.G.

    1985-01-01

    One mechanism of histamine-mediated inhibition of the immune response in man is to activate T suppressor cells that bear the Leu 2 (OKT8) marker. The current study was undertaken to characterize the histamine-induced suppressor cell using a monoclonal antibody (9.3) shown previously to distinguish cytotoxic T cells from antigen-specific suppressor T cells. Leu 2+ cells isolated from peripheral blood were further separated with antibody 9.3 into Leu 2+, 9.3+, and Leu 2+, 9.3- subsets and each subset was incubated with different concentrations of histamine before determining their ability to suppress immune responses in vitro. The results indicate that the Leu 2+, 9.3- subpopulation includes all histamine-induced suppressor cells, that 10(-4) M histamine is the optimal concentration for suppressor cell induction, and that exposure of Leu 2+, 9.3- cells to histamine for 30 s is sufficient to initiate the induction process. After treatment with histamine these cells inhibit both phytohemagglutinin-induced T cell proliferation and pokeweed mitogen-induced B cell differentiation. The suppression of phytohemagglutinin-induced proliferation was resistant to x-irradiation with 1,200 rad, either before or after histamine exposure, suggesting that Leu 2+, 9.3- cells need not proliferate to become suppressor cells or exert suppression. Moreover, suppression by these cells was not due to altered kinetics of the response. Finally, a histamine type 2 receptor antagonist (cimetidine) but not a type 1 receptor antagonist (mepyramine) blocked the induction of suppressor cells. On the basis of these results and our previous studies of antigen specific suppressor cells, we conclude that Leu 2+ suppressor cells in man are derived from a precursor pool that is phenotypically distinct from cells that can differentiate into cytotoxic T cells

  10. BDNF: An Oncogene or Tumor Suppressor?

    Science.gov (United States)

    Radin, Daniel P; Patel, Parth

    2017-08-01

    Neurotrophins are a family of growth factors that are vital to the proper development of the central nervous system. Their effects on cells are governed by the expression and activation of the tyrosine kinase receptors TrkA, TrkB and TrkC. TrkB has been immensely implicated in mediating neuronal migration, development and differentiation. It has also been shown to protect several neuronal cell types from an array of cytotoxic stressors after activation by its conjugate ligand brain-derived neurotrophic factor (BDNF). Over the past two decades, it has been shown that TrkB and BDNF are up-regulated in many types of cancers, conferring aggressive phenotypes underpinned by their resistance to several standard chemotherapeutic agents. This resistance to chemotherapy is modulated by the downstream targets of the TrkB receptor which include the well-characterized PI3K /Akt growth pathway, a hallmark of uncontrolled cancer cell growth and proliferation. Pre-clinical efforts to develop inhibitors of this receptor are promising, and such inhibitors also seem to sensitize cancer cells to standard chemotherapies. However, new evidence suggests that BDNF overexpression in the hypothalamus has immunoaugmenting properties, eliciting an increased anti-tumor immune response and reducing the activity of several proteins that would normally confer resistance to chemotherapeutic agents. In the current work, we provide a global analysis of the physiological consequences of TrkB receptor activation in vitro and discuss the dynamic consequences of TrkB activation in vivo. Finally, we propose a clinically-feasible option for increasing BDNF expression in the hypothalamus to more readily utilize the oncolytic effects of BDNF. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

  11. Suppressor of cytokine signaling 1 interacts with oncogenic lymphocyte-specific protein tyrosine kinase.

    Science.gov (United States)

    Venkitachalam, Srividya; Chueh, Fu-Yu; Leong, King-Fu; Pabich, Samantha; Yu, Chao-Lan

    2011-03-01

    Lymphocyte-specific protein tyrosine kinase (Lck) plays a key role in T cell signal transduction and is tightly regulated by phosphorylation and dephosphorylation. Lck can function as an oncoprotein when overexpressed or constantly activated by mutations. Our previous studies showed that Lck-induced cellular transformation could be suppressed by enforced expression of suppressor of cytokine signaling 1 (SOCS1), a SOCS family member involved in the negative feedback control of cytokine signaling. We observed attenuated Lck kinase activity in SOCS1-expressing cells, suggesting an important role of SOCS in regulating Lck functions. It remains largely unknown whether and how SOCS proteins interact with the oncogenic Lck kinase. Here, we report that among four SOCS family proteins, SOCS1, SOCS2, SOCS3 and CIS (cytokine-inducible SH2 domain containing protein), SOCS1 has the highest affinity in binding to the oncogenic Lck kinase. We identified the positive regulatory phosphotyrosine 394 residue in the kinase domain as the key interacting determinant in Lck. Additionally, the Lck kinase domain alone is sufficient to bind SOCS1. While the SH2 domain in SOCS1 is important in its association with the oncogenic Lck kinase, other functional domains may also contribute to overall binding affinity. These findings provide important mechanistic insights into the role of SOCS proteins as tumor suppressors in cells transformed by oncogenic protein tyrosine kinases.

  12. Molecular biology III - Oncogenes and tumor suppressor genes

    International Nuclear Information System (INIS)

    Giaccia, Amato J.

    1996-01-01

    Purpose: The purpose of this course is to introduce to radiation oncologists the basic concepts of tumorigenesis, building on the information that will be presented in the first and second part of this series of lectures. Objective: Our objective is to increase the current understanding of radiation oncologists with the process of tumorigenesis, especially focusing on genes that are altered in many tumor types that are potential candidates for novel molecular strategies. As strategies to treat cancer of cancer are becoming more sophisticated, it will be important for both the practitioner and academician to develop a basic understanding of the function of cancer 'genes'. This will be the third in a series of refresher courses that are meant to address recent advances in Cancer Biology in a way that both clinicians without previous knowledge of molecular biology or experienced researchers will find interesting. The lecture will begin with a basic overview of tumorigenesis; methods of detecting chromosome/DNA alterations, approaches used to isolate oncogenes and tumor suppressor genes, and their role in cell killing by apoptosis. Special attention will be given to oncogenes and tumor suppressor genes that are modulated by ionizing radiation and the tumor microenvironment. We will relate the biology of oncogenes and tumor suppressor genes to basic aspects of radiation biology that would be important in clinical practice. Finally, we will review recent studies on the prognostic significance of p53 mutations and apoptosis in tumor specimens. The main point of this lecture is to relate both researcher and clinician what are the therapeutic ramifications of oncogene and tumor suppressor gene mutations found in human neoptasia

  13. Targeting of the tumor suppressor GRHL3 by a miR-21-dependent proto-oncogenic network results in PTEN loss and tumorigenesis.

    Science.gov (United States)

    Darido, Charbel; Georgy, Smitha R; Wilanowski, Tomasz; Dworkin, Sebastian; Auden, Alana; Zhao, Quan; Rank, Gerhard; Srivastava, Seema; Finlay, Moira J; Papenfuss, Anthony T; Pandolfi, Pier Paolo; Pearson, Richard B; Jane, Stephen M

    2011-11-15

    Despite its prevalence, the molecular basis of squamous cell carcinoma (SCC) remains poorly understood. Here, we identify the developmental transcription factor Grhl3 as a potent tumor suppressor of SCC in mice, and demonstrate that targeting of Grhl3 by a miR-21-dependent proto-oncogenic network underpins SCC in humans. Deletion of Grhl3 in adult epidermis evokes loss of expression of PTEN, a direct GRHL3 target, resulting in aggressive SCC induced by activation of PI3K/AKT/mTOR signaling. Restoration of Pten expression completely abrogates SCC formation. Reduced levels of GRHL3 and PTEN are evident in human skin, and head and neck SCC, associated with increased expression of miR-21, which targets both tumor suppressors. Our data define the GRHL3-PTEN axis as a critical tumor suppressor pathway in SCC. 2011 Elsevier Inc. All rights reserved.

  14. Classification of suppressor additives based on synergistic and antagonistic ensemble effects

    Energy Technology Data Exchange (ETDEWEB)

    Broekmann, P., E-mail: peter.broekmann@iac.unibe.ch [BASF SE, Global Business Unit Electronic Materials, 67056 Ludwigshafen (Germany); Department of Chemistry and Biochemistry, University of Bern, Bern (Switzerland); Fluegel, A.; Emnet, C.; Arnold, M.; Roeger-Goepfert, C.; Wagner, A. [BASF SE, Global Business Unit Electronic Materials, 67056 Ludwigshafen (Germany); Hai, N.T.M. [Department of Chemistry and Biochemistry, University of Bern, Bern (Switzerland); Mayer, D. [BASF SE, Global Business Unit Electronic Materials, 67056 Ludwigshafen (Germany)

    2011-05-01

    Highlights: > Three fundamental types of suppressor additives for copper electroplating could be identified by means of potential transient measurements. > These suppressor additives differ in their synergistic and antagonistic interplay with anions that are chemisorbed on the metallic copper surface during electrodeposition. > In addition these suppressor chemistries reveal different barrier properties with respect to cupric ions and plating additives (Cl, SPS). - Abstract: Three fundamental types of suppressor additives for copper electroplating could be identified by means of potential transient measurements. These suppressor additives differ in their synergistic and antagonistic interplay with anions that are chemisorbed on the metallic copper surface during electrodeposition. In addition these suppressor chemistries reveal different barrier properties with respect to cupric ions and plating additives (Cl, SPS). While the type-I suppressor selectively forms efficient barriers for copper inter-diffusion on chloride-terminated electrode surfaces we identified a type-II suppressor that interacts non-selectively with any kind of anions chemisorbed on copper (chloride, sulfate, sulfonate). Type-I suppressors are vital for the superconformal copper growth mode in Damascene processing and show an antagonistic interaction with SPS (Bis-Sodium-Sulfopropyl-Disulfide) which involves the deactivation of this suppressor chemistry. This suppressor deactivation is rationalized in terms of compositional changes in the layer of the chemisorbed anions due to the competition of chloride and MPS (Mercaptopropane Sulfonic Acid) for adsorption sites on the metallic copper surface. MPS is the product of the dissociative SPS adsorption within the preexisting chloride matrix on the copper surface. The non-selectivity in the adsorption behavior of the type-II suppressor is rationalized in terms of anion/cation pairing effects of the poly-cationic suppressor and the anion-modified copper

  15. Paracrine Apoptotic Effect of p53 Mediated by Tumor Suppressor Par-4

    Directory of Open Access Journals (Sweden)

    Ravshan Burikhanov

    2014-01-01

    Full Text Available The guardian of the genome, p53, is often mutated in cancer and may contribute to therapeutic resistance. Given that p53 is intact and functional in normal tissues, we harnessed its potential to inhibit the growth of p53-deficient cancer cells. Specific activation of p53 in normal fibroblasts selectively induced apoptosis in p53-deficient cancer cells. This paracrine effect was mediated by p53-dependent secretion of the tumor suppressor Par-4. Accordingly, the activation of p53 in normal mice, but not p53−/− or Par-4−/− mice, caused systemic elevation of Par-4, which induced apoptosis of p53-deficient tumor cells. Mechanistically, p53 induced Par-4 secretion by suppressing the expression of its binding partner, UACA, which sequesters Par-4. Thus, normal cells can be empowered by p53 activation to induce Par-4 secretion for the inhibition of therapy-resistant tumors.

  16. A novel proapoptotic gene PANO encodes a post-translational modulator of the tumor suppressor p14ARF

    Energy Technology Data Exchange (ETDEWEB)

    Watari, Akihiro; Li, Yang; Higashiyama, Shinji; Yutsudo, Masuo, E-mail: yutsudo@biken.osaka-u.ac.jp

    2012-02-01

    The protein p14ARF is a known tumor suppressor protein controlling cell proliferation and survival, which mainly localizes in nucleoli. However, the regulatory mechanisms that govern its activity or expression remain unclear. Here, we report that a novel proapoptotic nucleolar protein, PANO, modulates the expression and activity of p14ARF in HeLa cells. Overexpression of PANO enhances the stability of p14ARF protein by protecting it from degradation, resulting in an increase in p14ARF expression levels. Overexpression of PANO also induces apoptosis under low serum conditions. This effect is dependent on the nucleolar localization of PANO and inhibited by knocking-down p14ARF. Alternatively, PANO siRNA treated cells exhibit a reduction in p14ARF protein levels. In addition, ectopic expression of PANO suppresses the tumorigenicity of HeLa cells in nude mice. These results indicate that PANO is a new apoptosis-inducing gene by modulating the tumor suppressor protein, p14ARF, and may itself be a new candidate tumor suppressor gene.

  17. The Regulation of Tumor Suppressor p63 by the Ubiquitin-Proteasome System

    Directory of Open Access Journals (Sweden)

    Stephen R. Armstrong

    2016-12-01

    Full Text Available The protein p63 has been identified as a homolog of the tumor suppressor protein p53 and is capable of inducing apoptosis, cell cycle arrest, or senescence. p63 has at least six isoforms, which can be divided into two major groups: the TAp63 variants that contain the N-terminal transactivation domain and the ΔNp63 variants that lack the N-terminal transactivation domain. The TAp63 variants are generally considered to be tumor suppressors involved in activating apoptosis and suppressing metastasis. ΔNp63 variants cannot induce apoptosis but can act as dominant negative inhibitors to block the function of TAp53, TAp73, and TAp63. p63 is rarely mutated in human tumors and is predominately regulated at the post-translational level by phosphorylation and ubiquitination. This review focuses primarily on regulation of p63 by the ubiquitin E-3 ligase family of enzymes via ubiquitination and proteasome-mediated degradation, and introduces a new key regulator of the p63 protein.

  18. Interference in plant defense and development by non-structural protein NSs of Groundnut bud necrosis virus.

    Science.gov (United States)

    Goswami, Suneha; Sahana, Nandita; Pandey, Vanita; Doblas, Paula; Jain, R K; Palukaitis, Peter; Canto, Tomas; Praveen, Shelly

    2012-01-01

    Groundnut bud necrosis virus (GBNV) infects a large number of leguminous and solanaceous plants. To elucidate the biological function of the non-structural protein encoded by the S RNA of GBNV (NSs), we studied its role in RNA silencing suppression and in viral pathogenesis. Our results demonstrated that GBNV NSs functions as a suppressor of RNA silencing using the agroinfiltration patch assay. An in silico analysis suggested the presence of pro-apoptotic protein Reaper-like sequences in the GBNV NSs, which were known to be present in animal infecting bunyaviruses. Utilizing NSs mutants, we demonstrated that a Leu-rich domain was required for RNA silencing suppression activity, but not the non-overlapping Trp/GH3 motif of the Reaper-like sequence. To investigate the role of NSs in symptom development we generated transgenic tomato expressing the GBNV NSs and showed that the expression of NSs in tomato mimics symptoms induced by infection with GBNV, such as leaf senescence and necrosis. As leaf senescence is controlled by miR319 regulation of the transcription factor TCP1, we assessed the accumulation of both RNAs in transgenic NSs-expressing and GBNV-infected tomato plants. In both types of plants the levels of miR319 decreased, while the levels of TCP1 transcripts increased. We propose that GBNV-NSs affects miRNA biogenesis through its RNA silencing suppressor activity and interferes with TCP1-regulated leaf developmental pathways. Copyright © 2011 Elsevier B.V. All rights reserved.

  19. The tumor suppressor gene Trp53 protects the mouse lens against posterior subcapsular cataracts and the BMP receptor Acvr1 acts as a tumor suppressor in the lens

    Directory of Open Access Journals (Sweden)

    Luke A. Wiley

    2011-07-01

    We previously found that lenses lacking the Acvr1 gene, which encodes a bone morphogenetic protein (BMP receptor, had abnormal proliferation and cell death in epithelial and cortical fiber cells. We tested whether the tumor suppressor protein p53 (encoded by Trp53 affected this phenotype. Acvr1 conditional knockout (Acvr1CKO mouse fiber cells had increased numbers of nuclei that stained for p53 phosphorylated on serine 15, an indicator of p53 stabilization and activation. Deletion of Trp53 rescued the Acvr1CKO cell death phenotype in embryos and reduced Acvr1-dependent apoptosis in postnatal lenses. However, deletion of Trp53 alone increased the number of fiber cells that failed to withdraw from the cell cycle. Trp53CKO and Acvr1;Trp53DCKO (double conditional knockout, but not Acvr1CKO, lenses developed abnormal collections of cells at the posterior of the lens that resembled posterior subcapsular cataracts. Cells from human posterior subcapsular cataracts had morphological and molecular characteristics similar to the cells at the posterior of mouse lenses lacking Trp53. In Trp53CKO lenses, cells in the posterior plaques did not proliferate but, in Acvr1;Trp53DCKO lenses, many cells in the posterior plaques continued to proliferate, eventually forming vascularized tumor-like masses at the posterior of the lens. We conclude that p53 protects the lens against posterior subcapsular cataract formation by suppressing the proliferation of fiber cells and promoting the death of any fiber cells that enter the cell cycle. Acvr1 acts as a tumor suppressor in the lens. Enhancing p53 function in the lens could contribute to the prevention of steroid- and radiation-induced posterior subcapsular cataracts.

  20. PHTS, a novel putative tumor suppressor, is involved in the transformation reversion of HeLaHF cells independently of the p53 pathway

    International Nuclear Information System (INIS)

    Yu Dehua; Fan, Wufang; Liu, Guohong; Nguy, Vivian; Chatterton, Jon E.; Long Shilong; Ke, Ning; Meyhack, Bernd; Bruengger, Adrian; Brachat, Arndt; Wong-Staal, Flossie; Li, Qi-Xiang

    2006-01-01

    HeLaHF is a non-transformed revertant of HeLa cells, likely resulting from the activation of a putative tumor suppressor(s). p53 protein was stabilized in this revertant and reactivated for certain transactivation functions. Although p53 stabilization has not conclusively been linked to the reversion, it is clear that the genes in p53 pathway are involved. The present study confirms the direct role of p53 in HeLaHF reversion by demonstrating that RNAi-mediated p53 silencing partially restores anchorage-independent growth potential of the revertant through the suppression of anoikis. In addition, we identified a novel gene, named PHTS, with putative tumor suppressor properties, and showed that this gene is also involved in HeLaHF reversion independently of the p53 pathway. Expression profiling revealed that PHTS is one of the genes that is up-regulated in HeLaHF but not in HeLa. It encodes a putative protein with CD59-like domains. RNAi-mediated PHTS silencing resulted in the partial restoration of transformation (anchorage-independent growth) in HeLaHF cells, similar to that of p53 gene silencing, implying its tumor suppressor effect. However, the observed increased transformation potential by PHTS silencing appears to be due to an increased anchorage-independent proliferation rate rather than suppression of anoikis, unlike the effect of p53 silencing. p53 silencing did not affect PHTS gene expression, and vice versa, suggesting PHTS may function in a new and p53-independent tumor suppressor pathway. Furthermore, over-expression of PHTS in different cancer cell lines, in addition to HeLa, reduces cell growth likely via induced apoptosis, confirming the broad PHTS tumor suppressor properties

  1. Role of Ubiquitylation in Controlling Suppressor of Cytokine Signalling 3 (SOCS3 Function and Expression

    Directory of Open Access Journals (Sweden)

    Jamie J. L. Williams

    2014-05-01

    Full Text Available The realisation that unregulated activation of the Janus kinase–signal transducer and activator of transcription (JAK–STAT pathway is a key driver of a wide range of diseases has identified its components as targets for therapeutic intervention by small molecule inhibitors and biologicals. In this review, we discuss JAK-STAT signalling pathway inhibition by the inducible inhibitor “suppressor of cytokine signaling 3 (SOCS3, its role in diseases such as myeloproliferative disorders, and its function as part of a multi-subunit E3 ubiquitin ligase complex. In addition, we highlight potential applications of these insights into SOCS3-based therapeutic strategies for management of conditions such as vascular re-stenosis associated with acute vascular injury, where there is strong evidence that multiple processes involved in disease progression could be attenuated by localized potentiation of SOCS3 expression levels.

  2. Nature of the suppressor cells mediating prolonged graft survival after administration of extracted histocompatibility antigen and cyclosporine

    International Nuclear Information System (INIS)

    Yoshimura, N.; Kahan, B.D.

    1985-01-01

    Antigen-specific suppressor T cells are induced by donor histocompatibility antigen extracted from spleen cells with 3M KCl combined with cyclosporine (Ag-CsA). A single i.v. injection of 5 mg 3M-KCl-extracted donor Buffalo (Buf, RT1b) antigen (Ag) combined with a three day course of CsA prolonged renal allograft survival in Wistar-Furth (WFu, RT1u) hosts to a greater extent (MST 26.5 days) than CsA alone (MST 11.8 days). Peripheral blood lymphocytes (PBL) or spleen cells harvested from Ag-CsA-treated recipients ten days after transplantation inhibited the mixed lymphocyte reaction (MLR) between normal responder WFu cells and irradiated Buf cells (55.6% and 64.4% suppression, respectively, P less than 0.025), but not third-party Brown-Norway (BN, RT1n) stimulator cells (13.6% and -18.3% suppression, respectively, NS). The suppressor effect was not mediated by cytolytic cells; there was neither primary nor secondary cytolytic activity against 51 Cr-labeled Con-A blastoid Buf cells. The suppressor cells were neither adherent to plastic dishes nor to nylon-wool columns. PBL irradiated with 800 rads, but not 1500 rads, suppressed the MLR. A single injection of cyclophosphamide (CY, 25 mg/kg) seven days after transplantation abrogated the suppression induced by Ag-CsA treatment. Moreover, PBL from Ag-CsA recipients failed to suppress the MLR, if depleted either of all T cells by treatment with monoclonal antibody (Mab) W3/13 HLK (pan T cells; % suppression -15.8), or of cytotoxic/suppressor cells with Mab OX-8 (-19.3% suppression) together with rabbit antimouse immunoglobulin and complement

  3. Leucine insertion caused by a yeast amber suppressor

    Energy Technology Data Exchange (ETDEWEB)

    Liebman, S W [Univ. of Rochester School of Medicine and Dentistry, NY; Stewart, J W; Parker, J H; Sherman, F

    1977-01-01

    The amber suppressor SUP52 can cause the production of approximately 15 to 20% of the normal amount of iso-l-cytochrome c when coupled to the amber (UAG) mutant cyc1-76. The suppressed iso-l-cytochrome c contains a residue of leucine at the position corresponding to the site of the amber codon. SUP52 also supresses another amber allele cyc1-179, but only with a low efficiency of approximately 2%. It does not appear to act at all on ochre (UAA) mutants. SUP52 was found to be on the left arm of chromosome X closely linked to the centromere.

  4. The Tumor Suppressor Hace1 Is a Critical Regulator of TNFR1-Mediated Cell Fate

    Directory of Open Access Journals (Sweden)

    Luigi Tortola

    2016-05-01

    Full Text Available Summary: The HECT domain E3 ligase HACE1 has been identified as a tumor suppressor in multiple cancers. Here, we report that HACE1 is a central gatekeeper of TNFR1-induced cell fate. Genetic inactivation of HACE1 inhibits TNF-stimulated NF-κB activation and TNFR1-NF-κB-dependent pathogen clearance in vivo. Moreover, TNF-induced apoptosis was impaired in hace1 mutant cells and knockout mice in vivo. Mechanistically, HACE1 is essential for the ubiquitylation of the adaptor protein TRAF2 and formation of the apoptotic caspase-8 effector complex. Intriguingly, loss of HACE1 does not impair TNFR1-mediated necroptotic cell fate via RIP1 and RIP3 kinases. Loss of HACE1 predisposes animals to colonic inflammation and carcinogenesis in vivo, which is markedly alleviated by genetic inactivation of RIP3 kinase and TNFR1. Thus, HACE1 controls TNF-elicited cell fate decisions and exerts tumor suppressor and anti-inflammatory activities via a TNFR1-RIP3 kinase-necroptosis pathway. : Tortola et al. report that the E3 ubiquitin ligase HACE1 is a gatekeeper of TNFR1-mediated cell fate. Hace1 deficiency impairs TNF-driven NF-κB activation and apoptosis and predisposes cells to necroptosis. Consequently, hace1–/– mice show enhanced colitis and colon cancer, which can be reverted by inactivation of pro-necroptotic kinase RIP3 and TNFR1.

  5. Transonic Performance Characteristics of Several Jet Noise Suppressors

    Science.gov (United States)

    Schmeer, James W.; Salters, Leland B., Jr.; Cassetti, Marlowe D.

    1960-01-01

    An investigation of the transonic performance characteristics of several noise-suppressor configurations has been conducted in the Langley 16-foot transonic tunnel. The models were tested statically and over a Mach number range from 0.70 to 1.05 at an angle of attack of 0 deg. The primary jet total-pressure ratio was varied from 1.0 (jet off) to about 4.5. The effect of secondary air flow on the performance of two of the configurations was investigated. A hydrogen peroxide turbojet-engine simulator was used to supply the hot-jet exhaust. An 8-lobe afterbody with centerbody, short shroud, and secondary air had the highest thrust-minus-drag coefficients of the six noise-suppressor configurations tested. The 12-tube and 12-lobe afterbodies had the lowest internal losses. The presence of an ejector shroud partially shields the external pressure distribution of the 8-lobe after-body from the influence of the primary jet. A ring-airfoil shroud increased the static thrust of the annular nozzle but generally decreased the thrust minus drag at transonic Mach numbers.

  6. The Ras suppressor-1 (RSU-1 in cancer

    Directory of Open Access Journals (Sweden)

    Lefteris C Zacharia

    2017-04-01

    Full Text Available Primary tumors are seldom the cause of death for cancer patients as most patients die from metastatic disease. Thus, deciphering metastatic mechanisms and key molecules involved is of utmost importance for the improved survival of cancer patients. Metastasis is a complex process in which cancer cells dissociate from the original tumor and spread to distant sites of the body. During the metastatic process, cancer cells lose contact both with the extracellular matrix (ECM and the neighboring cells within the primary tumor, thus invading though surrounding tissues. Therefore, ECM, and ECM-related adhesion proteins play a critical role in the metastatic process. Ras suppressor-1 (RSU-1 was first identified as a suppressor of Ras-dependent oncogenic transformation and is localized to cell-ECM adhesions where it is known to interact with the pro-survival adhesion protein PINCH-1. Although the connection to cancer is obvious, little is known regarding its expression in various cancer types. This opinion piece is focusing on recent literature regarding the expression of RSU-1 in various cancer types and the possible molecular mechanism of its action, pointing towards questions that need still to be addressed in this research field.

  7. Free silanols and ionic liquids as their suppressors in liquid chromatography.

    Science.gov (United States)

    Buszewska-Forajta, Magdalena; Markuszewski, Michał J; Kaliszan, Roman

    2018-07-20

    In this review, we will firstly discuss the types and the general properties of silica, focusing on the silica support used in chromatography and capillary electrophoresis. Additionally, the characterization of functional groups (silanols and siloxanes) will be considered in terms of activity of the stationary phases. We will then discuss physical chemistry of the stationary phases applied in liquid chromatography and capillary electrophoresis. The use of ionic liquids as a silanols' suppressors will be presented in the next parts of the study, along with the examples of specific applications. The review is completed with conclusions and an outlook for the future developments in the area of analytical applications of ionic liquids. Copyright © 2018 Elsevier B.V. All rights reserved.

  8. Enhancer-Mediated Oncogenic Function of the Menin Tumor Suppressor in Breast Cancer

    Directory of Open Access Journals (Sweden)

    Koen M.A. Dreijerink

    2017-03-01

    Full Text Available While the multiple endocrine neoplasia type 1 (MEN1 gene functions as a tumor suppressor in a variety of cancer types, we explored its oncogenic role in breast tumorigenesis. The MEN1 gene product menin is involved in H3K4 trimethylation and co-activates transcription. We integrated ChIP-seq and RNA-seq data to identify menin target genes. Our analysis revealed that menin-dependent target gene promoters display looping to distal enhancers that are bound by menin, FOXA1 and GATA3. In this fashion, MEN1 co-regulates a proliferative breast cancer-specific gene expression program in ER+ cells. In primary mammary cells, MEN1 exerts an anti-proliferative function by regulating a distinct expression signature. Our findings clarify the cell-type-specific functions of MEN1 and inform the development of menin-directed treatments for breast cancer.

  9. The Role and Potential Therapeutic Application of Myeloid-Derived Suppressor Cells in Allo- and Autoimmunity

    Directory of Open Access Journals (Sweden)

    Qi Zhang

    2015-01-01

    Full Text Available Myeloid-derived suppressor cells (MDSCs are a heterogeneous population of cells that consists of myeloid progenitor cells and immature myeloid cells. They have been identified as a cell population that may affect the activation of CD4+ and CD8+ T-cells to regulate the immune response negatively, which makes them attractive targets for the treatment of transplantation and autoimmune diseases. Several studies have suggested the potential suppressive effect of MDSCs on allo- and autoimmune responses. Conversely, MDSCs have also been found at various stages of differentiation, accumulating during pathological situations, not only during tumor development but also in a variety of inflammatory immune responses, bone marrow transplantation, and some autoimmune diseases. These findings appear to be contradictory. In this review, we summarize the roles of MDSCs in different transplantation and autoimmune diseases models as well as the potential to target these cells for therapeutic benefit.

  10. DA-Raf, a dominant-negative antagonist of the Ras-ERK pathway, is a putative tumor suppressor.

    Science.gov (United States)

    Kanno, Emiri; Kawasaki, Osamu; Takahashi, Kazuya; Takano, Kazunori; Endo, Takeshi

    2018-01-01

    Activating mutations of RAS genes, particularly KRAS, are detected with high frequency in human tumors. Mutated Ras proteins constitutively activate the ERK pathway (Raf-MEK-ERK phosphorylation cascade), leading to cellular transformation and tumorigenesis. DA-Raf1 (DA-Raf) is a splicing variant of A-Raf and contains the Ras-binding domain (RBD) but lacks the kinase domain. Accordingly, DA-Raf antagonizes the Ras-ERK pathway in a dominant-negative fashion and suppresses constitutively activated K-Ras-induced cellular transformation. Thus, we have addressed whether DA-Raf serves as a tumor suppressor of Ras-induced tumorigenesis. DA-Raf(R52Q), which is generated from a single nucleotide polymorphism (SNP) in the RBD, and DA-Raf(R52W), a mutant detected in a lung cancer, neither bound to active K-Ras nor interfered with the activation of the ERK pathway. They were incapable of suppressing activated K-Ras-induced cellular transformation and tumorigenesis in mice, in which K-Ras-transformed cells were transplanted. Furthermore, although DA-Raf was highly expressed in lung alveolar epithelial type 2 (AE2) cells, its expression was silenced in AE2-derived lung adenocarcinoma cell lines with oncogenic KRAS mutations. These results suggest that DA-Raf represents a tumor suppressor protein against Ras-induced tumorigenesis. Copyright © 2017 Elsevier Inc. All rights reserved.

  11. Comparison of the suppressor cells found in the spleens of 89Sr-treated mice and in normal murine bone marrow

    International Nuclear Information System (INIS)

    Levy, E.M.; Corvese, J.S.; Bennett, M.

    1981-01-01

    Normal murine bone marrow cells and spleen cells of mice treated with 89 Sr both have suppressive activity. These nonspecific suppressor cells inhibit the ability of normal spleen cells to undergo antibody responses in vitro. After being precultured for 24 hr, these cells will also suppress antibody responses in vivo and the responses of normal spleen cells to T and B cell mitogens in vitro. These cells have previously been shown not to be mature T or B lymphocytes or macrophages. Velocity sedimentation and cell-size analysis indicated that both suppressor cells are large (approx. =206 μ 3 ). Mitomycin C treatment eliminated the ability of both suppressor cells to inhibit an in vitro antibody response. In contrast, this treatment did not reduce the ability of the cells to inhibit an in vitro antibody response. In contrast, this treatment did not reduce the ability of the cells to suppress a mitogenic response. Irradiation (1000 R) was also ineffective in eliminating the ability of either cell to suppress a mitogenic response. We conclude that the 2 suppressor cells are closely related if not identical, and we speculate that these cells may function in vivo to suppress immune reactivity in areas of intense hematopoiesis

  12. Determination of Heritage SSME Pogo Suppressor Resistance and Inertance from Waterflow Pulse Testing

    Science.gov (United States)

    McDougal, Chris; Eberhart, Chad; Lee, Erik

    2016-01-01

    Waterflow tests of a heritage Space Shuttle Main Engine pogo suppressor were performed to experimentally quantify the resistance and inertance provided by the suppressor. Measurements of dynamic pressure and flow rate in response to pulsing flow were made throughout the test loop. A unique system identification methodology combined all sensor measurements with a one-dimensional perturbational flow model of the complete water flow loop to spatially translate physical measurements to the device under test. Multiple techniques were then employed to extract the effective resistance and inertance for the pogo suppressor. Parameters such as steady flow rate, perturbational flow rate magnitude, and pulse frequency were investigated to assess their influence on the behavior of the pogo suppressor dynamic response. These results support validation of the RS-25 pogo suppressor performance for use on the Space Launch System Core Stage.

  13. Structural characterization of suppressor lipids by high-resolution mass spectrometry

    DEFF Research Database (Denmark)

    Rovillos, Mary Joy; Pauling, Josch Konstantin; Hannibal-Bach, Hans Kristian

    2016-01-01

    RATIONALE: Suppressor lipids were originally identified in 1993 and reported to encompass six lipid classes that enable Saccharomyces cerevisiae to live without sphingolipids. Structural characterization, using non-mass spectrometric approaches, revealed that these suppressor lipids are very long...... chain fatty acid (VLCFA)-containing glycerophospholipids with polar head groups that are typically incorporated into sphingolipids. Here we report, for the first time, the structural characterization of the yeast suppressor lipids using high-resolution mass spectrometry. METHODS: Suppressor lipids were...... isolated by preparative chromatography and subjected to structural characterization using hybrid quadrupole time-of-flight and ion trap-orbitrap mass spectrometry. RESULTS: Our investigation recapitulates the overall structural features of the suppressor lipids and provides an in-depth characterization...

  14. NDRG2 is a candidate tumor-suppressor for oral squamous-cell carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Furuta, Hiroshi; Kondo, Yuudai [Division of Oral and Maxillofacial Surgery, Medicine of Sensory and Motor Organs, University of Miyazaki, 5200 Kihara, Kiyotake, Miyazaki-gun, Miyazaki 889-1692 (Japan); Division of Tumor and Cellular Biochemistry, Department of Medical Sciences, Faculty of Medicine, University of Miyazaki, 5200 Kihara, Kiyotake, Miyazaki-gun, Miyazaki 889-1692 (Japan); Nakahata, Shingo; Hamasaki, Makoto [Division of Tumor and Cellular Biochemistry, Department of Medical Sciences, Faculty of Medicine, University of Miyazaki, 5200 Kihara, Kiyotake, Miyazaki-gun, Miyazaki 889-1692 (Japan); Sakoda, Sumio [Division of Oral and Maxillofacial Surgery, Medicine of Sensory and Motor Organs, University of Miyazaki, 5200 Kihara, Kiyotake, Miyazaki-gun, Miyazaki 889-1692 (Japan); Morishita, Kazuhiro, E-mail: kmorishi@med.miyazaki-u.ac.jp [Division of Tumor and Cellular Biochemistry, Department of Medical Sciences, Faculty of Medicine, University of Miyazaki, 5200 Kihara, Kiyotake, Miyazaki-gun, Miyazaki 889-1692 (Japan)

    2010-01-22

    Oral cancer is one of the most common cancers worldwide, and squamous-cell carcinoma (OSCC) is the most common phenotype of oral cancer. Although patients with OSCC have poor survival rates and a high incidence of metastasis, the molecular mechanisms of OSCC development have not yet been elucidated. This study investigated whether N-myc downstream-regulated gene 2 (NDRG2) contributes to the carcinogenesis of OSCC, as NDRG2 is reported to be a candidate tumor-suppressor gene in a wide variety of cancers. The down-regulation of NDRG2 mRNA, which was dependent on promoter methylation, was seen in the majority of OSCC cases and in several cases of precancerous leukoplakia with dysplasia. Induction of NDRG2 expression in an HSC-3/OSCC cell line significantly inhibited cell proliferation and decreased colony formation ability on soft agar. The majority of OSCC cell lines showed an activation of PI3K/Akt signaling, and enforced expression of NDRG2 in HSC-3 cells decreased the level of phosphorylated Akt at Serine 473 (p-Akt). Immunohistochemical p-Akt staining was detected in 56.5% of the OSCC tumors, and 80.4% of the tumors were negative for NDRG2 staining. Moreover, positive p-Akt staining was inversely correlated with decreased NDRG2 expression in OSCC tumors with moderate to poor differentiation (p < 0.005). Therefore, NDRG2 is a candidate tumor-suppressor gene for OSCC development and probably contributes to the tumorigenesis of OSCC partly via the modulation of Akt signaling.

  15. Insight into the tumor suppressor function of CBP through the viral oncoprotein tax.

    Science.gov (United States)

    Van Orden, K; Nyborg, J K

    2000-01-01

    CREB binding protein (CBP) is a cellular coactivator protein that regulates essentially all known pathways of gene expression. The transcriptional coactivator properties of CBP are utilized by at least 25 different transcription factors representing nearly all known classes of DNA binding proteins. Once bound to their target genes, these transcription factors are believed to tether CBP to the promoter, leading to activated transcription. CBP functions to stimulate transcription through direct recruitment of the general transcription machinery as well as acetylation of both histone and transcription factor substrates. Recent observations indicate that a critical dosage of CBP is required for normal development and tumor suppression, and that perturbations in CBP concentrations may disrupt cellular homeostasis. Furthermore, there is accumulating evidence that CBP deregulation plays a direct role in hematopoietic malignancies. However, the molecular events linking CBP deregulation and malignant transformation are unclear. Further insight into the function of CBP, and its role as a tumor suppressor, can be gained through recent studies of the human T-cell leukemia virus, type I (HTLV-I) Tax oncoprotein. Tax is known to utilize CBP to stimulate transcription from the viral promoter. However, recent data suggest that as a consequence of the Tax-CBP interaction, many cellular transcription factor pathways may be deregulated. Tax disruption of CBP function may play a key role in transformation of the HTLV-I-infected cell. Thus, Tax derailment of CBP may lend important information about the tumor suppressor properties of CBP and serve as a model for the role of CBP in hematopoietic malignancies.

  16. Intragenic suppressor of Osiaa23 revealed a conserved tryptophan residue crucial for protein-protein interactions.

    Directory of Open Access Journals (Sweden)

    Jun Ni

    Full Text Available The Auxin/Indole-3-Acetic Acid (Aux/IAA and Auxin Response Factor (ARF are two important families that play key roles in auxin signal transduction. Both of the families contain a similar carboxyl-terminal domain (Domain III/IV that facilitates interactions between these two families. In spite of the importance of protein-protein interactions among these transcription factors, the mechanisms involved in these interactions are largely unknown. In this study, we isolated six intragenic suppressors of an auxin insensitive mutant, Osiaa23. Among these suppressors, Osiaa23-R5 successfully rescued all the defects of the mutant. Sequence analysis revealed that an amino acid substitution occurred in the Tryptophan (W residue in Domain IV of Osiaa23. Yeast two-hybrid experiments showed that the mutation in Domain IV prevents the protein-protein interactions between Osiaa23 and OsARFs. Phylogenetic analysis revealed that the W residue is conserved in both OsIAAs and OsARFs. Next, we performed site-specific amino acid substitutions within Domain IV of OsARFs, and the conserved W in Domain IV was exchanged by Serine (S. The mutated OsARF(WSs can be released from the inhibition of Osiaa23 and maintain the transcriptional activities. Expression of OsARF(WSs in Osiaa23 mutant rescued different defects of the mutant. Our results suggest a previously unknown importance of Domain IV in both families and provide an indirect way to investigate functions of OsARFs.

  17. Evidence against suppressor cell involvement in naturally acquired tolerance of a minor histocompatibility antigen

    International Nuclear Information System (INIS)

    Johnson, L.L.

    1991-01-01

    The hypothesis was investigated that suppressor cells may be responsible for maintenance of immunologic tolerance of a minor H3 antigen in mice that express the antigen naturally. Lymphoid cell populations from B6.C-H-24c (HW54) mice, a congenic-resistant strain histoincompatible with H-24b-expressing C57BL/6 (B6) mice only with respect to the H-24 locus, were examined in cell-transfer experiments to see if they contained naturally arising H-24c-specific suppressor cells. The H-24 antigen was chosen for these studies because, unlike most other minor and major histocompatibility (H) antigens, it is not detectable on mature lymphoid cells by any of several functional criteria. Thus transfer of HW54 lymphoid cells to B6 hosts could be done without the complication of inducing hyporesponsiveness de novo in the host, as occurs with other minor H antigens that are expressed on lymphocytes. B6 hosts were given HW54 skin grafts along with HW54 lymphoid cells to assess their tolerance of the H-24c-encoded antigen. The hosts were either (1) normal, nonimmune B6 mice; (2) B6 mice rendered immunodeficient by thymectomy and irradiation (TxB) and repopulated with H-24c-immune B6 lymphocytes; or (3) TxB B6 hosts repopulated with nonimmune B6 lymphocytes. In each case it was found that the additionally infused HW54 lymphoid cells did not suppress the ability of these hosts to reject HW54 skin grafts. In other words, HW54 lymphoid cells appear not to possess suppressive activity specific for the H-24c antigen that might maintain antigen-specific natural tolerance. Additional experiments were performed to determine whether HW54 lymphoid cells can inhibit the ability of sublethally irradiated B6 mice to regain the capacity to reject HW54 skin

  18. Myeloid derived suppressor cells as therapeutic target in hematological malignancies

    Directory of Open Access Journals (Sweden)

    Kim eDe Veirman

    2014-12-01

    Full Text Available Myeloid derived suppressor cells (MDSC are a heterogeneous population of immature myeloid cells that accumulate during pathological conditions such as cancer and are associated with a poor clinical outcome. MDSC expansion hampers the host anti-tumor immune response by inhibition of T cell proliferation, cytokine secretion and recruitment of regulatory T cells. In addition, MDSC exert non-immunological functions including the promotion of angiogenesis, tumor invasion and metastasis. Recent years, MDSC are considered as a potential target in solid tumors and hematological malignancies to enhance the effects of currently used immune modulating agents. This review focuses on the characteristics, distribution, functions, cell-cell interactions and targeting of MDSC in hematological malignancies including multiple myeloma, lymphoma and leukemia.

  19. Identification of an MLC suppressor cell population in acute leukemia

    International Nuclear Information System (INIS)

    Bryan, C.F.; Broxmeyer, H.E.; Hansen, J.; Pollack, M.; Dupont, B.

    1978-01-01

    The MLC data from the 20 nonsuppressing patients and the 10 suppressing leukemia patients were analyzed with regard to HLA-A, -B, and -C antigens in the leukemia patients and compared with the presence or absence of suppression. These results demonstrate a significant increase (p < 0.02, Mann-Whitney U test) of HLA antigens Al, A3, and A11 in the leukemia suppressor group. Seven of the 10 leukemia patients showing suppression were A1, A3, or A11, while only 4 of the 20 nonsuppressing leukemia patients carried any of these three HLA-A antigens. The studies demonstrate that a nonspecific suppression of MLC responses is observed in 33% of the patients with acute leukemia

  20. The protein histidine phosphatase LHPP is a tumour suppressor.

    Science.gov (United States)

    Hindupur, Sravanth K; Colombi, Marco; Fuhs, Stephen R; Matter, Matthias S; Guri, Yakir; Adam, Kevin; Cornu, Marion; Piscuoglio, Salvatore; Ng, Charlotte K Y; Betz, Charles; Liko, Dritan; Quagliata, Luca; Moes, Suzette; Jenoe, Paul; Terracciano, Luigi M; Heim, Markus H; Hunter, Tony; Hall, Michael N

    2018-03-29

    Histidine phosphorylation, the so-called hidden phosphoproteome, is a poorly characterized post-translational modification of proteins. Here we describe a role of histidine phosphorylation in tumorigenesis. Proteomic analysis of 12 tumours from an mTOR-driven hepatocellular carcinoma mouse model revealed that NME1 and NME2, the only known mammalian histidine kinases, were upregulated. Conversely, expression of the putative histidine phosphatase LHPP was downregulated specifically in the tumours. We demonstrate that LHPP is indeed a protein histidine phosphatase. Consistent with these observations, global histidine phosphorylation was significantly upregulated in the liver tumours. Sustained, hepatic expression of LHPP in the hepatocellular carcinoma mouse model reduced tumour burden and prevented the loss of liver function. Finally, in patients with hepatocellular carcinoma, low expression of LHPP correlated with increased tumour severity and reduced overall survival. Thus, LHPP is a protein histidine phosphatase and tumour suppressor, suggesting that deregulated histidine phosphorylation is oncogenic.

  1. p53 tumor suppressor gene: significance in neoplasia - a review

    International Nuclear Information System (INIS)

    Alam, J.M.

    2000-01-01

    p53 is a tumor suppressor gene located on chromosome 17p13.1. Its function includes cell cycle control and apoptosis. Loss of p53 function, either due to decreased level or genetic transformation, is associated with loss of cell cycle control, decrease, apoptosis and genomic modification, such mutation of p53 gene is now assessed and the indicator of neoplasia of cancer of several organs and cell types, p53 has demonstrated to have critical role in defining various progressive stages of neoplasia, therapeutic strategies and clinical application. The present review briefly describes function of p53 in addition to its diagnostic and prognostic significance in detecting several types of neoplasia. (author)

  2. PML tumor suppressor protein is required for HCV production

    International Nuclear Information System (INIS)

    Kuroki, Misao; Ariumi, Yasuo; Hijikata, Makoto; Ikeda, Masanori; Dansako, Hiromichi; Wakita, Takaji; Shimotohno, Kunitada; Kato, Nobuyuki

    2013-01-01

    Highlights: ► PML tumor suppressor protein is required for HCV production. ► PML is dispensable for HCV RNA replication. ► HCV could not alter formation of PML-NBs. ► INI1 and DDX5, PML-related proteins, are involved in HCV life cycle. -- Abstract: PML tumor suppressor protein, which forms discrete nuclear structures termed PML-nuclear bodies, has been associated with several cellular functions, including cell proliferation, apoptosis and antiviral defense. Recently, it was reported that the HCV core protein colocalizes with PML in PML-NBs and abrogates the PML function through interaction with PML. However, role(s) of PML in HCV life cycle is unknown. To test whether or not PML affects HCV life cycle, we examined the level of secreted HCV core and the infectivity of HCV in the culture supernatants as well as the level of HCV RNA in HuH-7-derived RSc cells, in which HCV-JFH1 can infect and efficiently replicate, stably expressing short hairpin RNA targeted to PML. In this context, the level of secreted HCV core and the infectivity in the supernatants from PML knockdown cells was remarkably reduced, whereas the level of HCV RNA in the PML knockdown cells was not significantly affected in spite of very effective knockdown of PML. In fact, we showed that PML is unrelated to HCV RNA replication using the subgenomic HCV-JFH1 replicon RNA, JRN/3-5B. Furthermore, the infectivity of HCV-like particle in the culture supernatants was significantly reduced in PML knockdown JRN/3-5B cells expressing core to NS2 coding region of HCV-JFH1 genome using the trans-packaging system. Finally, we also demonstrated that INI1 and DDX5, the PML-related proteins, are involved in HCV production. Taken together, these findings suggest that PML is required for HCV production.

  3. Cell size checkpoint control by the retinoblastoma tumor suppressor pathway.

    Science.gov (United States)

    Fang, Su-Chiung; de los Reyes, Chris; Umen, James G

    2006-10-13

    Size control is essential for all proliferating cells, and is thought to be regulated by checkpoints that couple cell size to cell cycle progression. The aberrant cell-size phenotypes caused by mutations in the retinoblastoma (RB) tumor suppressor pathway are consistent with a role in size checkpoint control, but indirect effects on size caused by altered cell cycle kinetics are difficult to rule out. The multiple fission cell cycle of the unicellular alga Chlamydomonas reinhardtii uncouples growth from division, allowing direct assessment of the relationship between size phenotypes and checkpoint function. Mutations in the C. reinhardtii RB homolog encoded by MAT3 cause supernumerous cell divisions and small cells, suggesting a role for MAT3 in size control. We identified suppressors of an mat3 null allele that had recessive mutations in DP1 or dominant mutations in E2F1, loci encoding homologs of a heterodimeric transcription factor that is targeted by RB-related proteins. Significantly, we determined that the dp1 and e2f1 phenotypes were caused by defects in size checkpoint control and were not due to a lengthened cell cycle. Despite their cell division defects, mat3, dp1, and e2f1 mutants showed almost no changes in periodic transcription of genes induced during S phase and mitosis, many of which are conserved targets of the RB pathway. Conversely, we found that regulation of cell size was unaffected when S phase and mitotic transcription were inhibited. Our data provide direct evidence that the RB pathway mediates cell size checkpoint control and suggest that such control is not directly coupled to the magnitude of periodic cell cycle transcription.

  4. Grape seed proanthocyanidins reactivate silenced tumor suppressor genes in human skin cancer cells by targeting epigenetic regulators

    International Nuclear Information System (INIS)

    Vaid, Mudit; Prasad, Ram; Singh, Tripti; Jones, Virginia; Katiyar, Santosh K.

    2012-01-01

    Grape seed proanthocyanidins (GSPs) have been shown to have anti-skin carcinogenic effects in in vitro and in vivo models. However, the precise epigenetic molecular mechanisms remain unexplored. This study was designed to investigate whether GSPs reactivate silenced tumor suppressor genes following epigenetic modifications in skin cancer cells. For this purpose, A431 and SCC13 human squamous cell carcinoma cell lines were used as in vitro models. The effects of GSPs on DNA methylation, histone modifications and tumor suppressor gene expressions were studied in these cell lines using enzyme activity assays, western blotting, dot-blot analysis and real-time polymerase chain reaction (RT-PCR). We found that treatment of A431 and SCC13 cells with GSPs decreased the levels of: (i) global DNA methylation, (ii) 5-methylcytosine, (iii) DNA methyltransferase (DNMT) activity and (iv) messenger RNA (mRNA) and protein levels of DNMT1, DNMT3a and DNMT3b in these cells. Similar effects were noted when these cancer cells were treated identically with 5-aza-2′-deoxycytidine, an inhibitor of DNA methylation. GSPs decreased histone deacetylase activity, increased levels of acetylated lysines 9 and 14 on histone H3 (H3-Lys 9 and 14) and acetylated lysines 5, 12 and 16 on histone H4, and reduced the levels of methylated H3-Lys 9. Further, GSP treatment resulted in re-expression of the mRNA and proteins of silenced tumor suppressor genes, RASSF1A, p16 INK4a and Cip1/p21. Together, this study provides a new insight into the epigenetic mechanisms of GSPs and may have significant implications for epigenetic therapy in the treatment/prevention of skin cancers in humans. -- Highlights: ►Epigenetic modulations have been shown to have a role in cancer risk. ►Proanthocyanidins decrease the levels of DNA methylation and histone deacetylation. ►Proanthocyanidins inhibit histone deacetylase activity in skin cancer cells. ►Proanthocyanidins reactivate tumor suppressor genes in skin

  5. DMPD: Regulation of innate immunity by suppressor of cytokine signaling (SOCS)proteins. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 18406369 Regulation of innate immunity by suppressor of cytokine signaling (SOCS)proteins...svg) (.html) (.csml) Show Regulation of innate immunity by suppressor of cytokine signaling (SOCS)proteins. ...PubmedID 18406369 Title Regulation of innate immunity by suppressor of cytokine signaling (SOCS)proteins

  6. An identity crisis for fps/fes: oncogene or tumor suppressor?

    Science.gov (United States)

    Sangrar, Waheed; Zirgnibl, Ralph A; Gao, Yan; Muller, William J; Jia, Zongchao; Greer, Peter A

    2005-05-01

    Fps/Fes proteins were among the first members of the protein tyrosine kinase family to be characterized as dominant-acting oncoproteins. Addition of retroviral GAG sequences or other experimentally induced mutations activated the latent transforming potential of Fps/Fes. However, activating mutations in fps/fes had not been found in human tumors until recently, when mutational analysis of a panel of colorectal cancers identified four somatic mutations in sequences encoding the Fps/Fes kinase domain. Here, we report biochemical and theoretical structural analysis demonstrating that three of these mutations result in inactivation, not activation, of Fps/Fes, whereas the fourth mutation compromised in vivo activity. These results did not concur with a classic dominant-acting oncogenic role for fps/fes involving activating somatic mutations but instead raised the possibility that inactivating fps/fes mutations might promote tumor progression in vivo. Consistent with this, we observed that tumor onset in a mouse model of breast epithelial cancer occurred earlier in mice targeted with either null or kinase-inactivating fps/fes mutations. Furthermore, a fps/fes transgene restored normal tumor onset kinetics in targeted fps/fes null mice. These data suggest a novel and unexpected tumor suppressor role for Fps/Fes in epithelial cells.

  7. The role of baculovirus apoptotic suppressors in AcMNPV-mediated translation arrest in Ld652Y cells

    International Nuclear Information System (INIS)

    Thiem, Suzanne M.; Chejanovsky, Nor

    2004-01-01

    Infecting the insect cell line IPLB-Ld652Y with the baculovirus Autographa californica multinucleocapsid nucleopolyhedrovirus (AcMNPV) results in global translation arrest, which correlates with the presence of the AcMNPV apoptotic suppressor, p35. In this study, we investigated the role of apoptotic suppression on AcMNPV-induced translation arrest. Infecting cells with AcMNPV bearing nonfunctional mutant p35 did not result in global translation arrest. In contrast, global translation arrest was observed in cells infected with AcMNPV in which p35 was replaced with Opiap, Cpiap, or p49, baculovirus apoptotic suppressors that block apoptosis by different mechanisms than p35. These results indicated that suppressing apoptosis triggered translation arrest in AcMNPV-infected Ld652Y cells. Experiments using the DNA synthesis inhibitor aphidicolin and temperature shift experiments, using the AcMNPV replication mutants ts8 and ts8Δp35, indicated that translation arrest initiated during the early phase of infection, but events during the late phase were required for global translation arrest. Peptide caspase inhibitors could not substitute for baculovirus apoptotic suppressors to induce translation arrest in Ld652Y cells infected with a p35-null virus. However, if the p35-null-AcMNPV also carried hrf-1, a novel baculovirus host range gene, progeny virus was produced and treatment with peptide caspase inhibitors enhanced translation of a late viral gene transcript. Together, these results indicate that translation arrest in AcMNPV-infected Ld652Y cells is due to the anti-apoptotic function of p35, but suggests that rather than simply preventing caspase activation, its activity enhances signaling to a separate translation arrest pathway, possibly by stimulating the late stages of the baculovirus infection cycle

  8. The chromatin remodelling factor BRG1 is a novel binding partner of the tumor suppressor p16INK4a

    Directory of Open Access Journals (Sweden)

    Mann Graham J

    2009-01-01

    Full Text Available Abstract Background CDKN2A/p16INK4a is frequently altered in human cancers and it is the most important melanoma susceptibility gene identified to date. p16INK4a inhibits pRb phosphorylation and induces cell cycle arrest, which is considered its main tumour suppressor function. Nevertheless, additional activities may contribute to the tumour suppressor role of p16INK4a and could help explain its specific association with melanoma predisposition. To identify such functions we conducted a yeast-two-hybrid screen for novel p16INK4a binding partners. Results We now report that p16INK4a interacts with the chromatin remodelling factor BRG1. We investigated the cooperative roles of p16INK4a and BRG1 using a panel of cell lines and a melanoma cell model with inducible p16INK4a expression and BRG1 silencing. We found evidence that BRG1 is not required for p16INK4a-induced cell cycle inhibition and propose that the p16INK4a-BRG1 complex regulates BRG1 chromatin remodelling activity. Importantly, we found frequent loss of BRG1 expression in primary and metastatic melanomas, implicating this novel p16INK4a binding partner as an important tumour suppressor in melanoma. Conclusion This data adds to the increasing evidence implicating the SWI/SNF chromatin remodelling complex in tumour development and the association of p16INK4a with chromatin remodelling highlights potentially new functions that may be important in melanoma predisposition and chemoresistance.

  9. Phytophthora effector targets a novel component of small RNA pathway in plants to promote infection.

    Science.gov (United States)

    Qiao, Yongli; Shi, Jinxia; Zhai, Yi; Hou, Yingnan; Ma, Wenbo

    2015-05-05

    A broad range of parasites rely on the functions of effector proteins to subvert host immune response and facilitate disease development. The notorious Phytophthora pathogens evolved effectors with RNA silencing suppression activity to promote infection in plant hosts. Here we report that the Phytophthora Suppressor of RNA Silencing 1 (PSR1) can bind to an evolutionarily conserved nuclear protein containing the aspartate-glutamate-alanine-histidine-box RNA helicase domain in plants. This protein, designated PSR1-Interacting Protein 1 (PINP1), regulates the accumulation of both microRNAs and endogenous small interfering RNAs in Arabidopsis. A null mutation of PINP1 causes embryonic lethality, and silencing of PINP1 leads to developmental defects and hypersusceptibility to Phytophthora infection. These phenotypes are reminiscent of transgenic plants expressing PSR1, supporting PINP1 as a direct virulence target of PSR1. We further demonstrate that the localization of the Dicer-like 1 protein complex is impaired in the nucleus of PINP1-silenced or PSR1-expressing cells, indicating that PINP1 may facilitate small RNA processing by affecting the assembly of dicing complexes. A similar function of PINP1 homologous genes in development and immunity was also observed in Nicotiana benthamiana. These findings highlight PINP1 as a previously unidentified component of RNA silencing that regulates distinct classes of small RNAs in plants. Importantly, Phytophthora has evolved effectors to target PINP1 in order to promote infection.

  10. Transcriptional Inhibition of the Human Papilloma Virus Reactivates Tumor Suppressor p53 in Cervical Carcinoma Cells

    Science.gov (United States)

    Kochetkov, D. V.; Ilyinskaya, G. V.; Komarov, P. G.; Strom, E.; Agapova, L. S.; Ivanov, A. V.; Budanov, A. V.; Frolova, E. I.; Chumakov, P. M.

    2009-01-01

    Inactivation of tumor suppressor p53 accompanies the majority of human malignancies. Restoration of p53 function causes death of tumor cells and is potentially suitable for gene therapy of cancer. In cervical carcinoma, human papilloma virus (HPV) E6 facilitates proteasomal degradation of p53. Hence, a possible approach to p53 reactivation is the use of small molecules suppressing the function of viral proteins. HeLa cervical carcinoma cells (HPV-18) with a reporter construct containing the b-galactosidase gene under the control of a p53-responsive promoter were used as a test system to screen a library of small molecules for restoration of the transcriptional activity of p53. The effect of the two most active compounds was studied with cell lines differing in the state of p53-dependent signaling pathways. The compounds each specifically activated p53 in cells expressing HPV-18 and, to a lesser extent, HPV-16 and exerted no effect on control p53-negative cells or cells with the intact p53-dependent pathways. Activation of p53 in cervical carcinoma cells was accompanied by induction of p53-dependent CDKN1 (p21), inhibition of cell proliferation, and induction of apoptosis. In addition, the two compounds dramatically decreased transcription of the HPV genome, which was assumed to cause p53 reactivation. The compounds were low-toxic for normal cells and can be considered as prototypes of new anticancer drugs. PMID:17685229

  11. Saponin Inhibits Hepatitis C Virus Propagation by Up-regulating Suppressor of Cytokine Signaling 2

    Science.gov (United States)

    Kang, Sang-Min; Min, Saehong; Son, Kidong; Lee, Han Sol; Park, Eun Mee; Ngo, Huong T. T.; Tran, Huong T. L.; Lim, Yun-Sook; Hwang, Soon B.

    2012-01-01

    Saponins are a group of naturally occurring plant glycosides which possess a wide range of pharmacological properties, including anti-tumorigenic and antiviral activities. To investigate whether saponin has anti-hepatitis C virus (HCV) activity, we examined the effect of saponin on HCV replication. HCV replication was efficiently inhibited at a concentration of 10 µg/ml of saponin in cell culture grown HCV (HCVcc)-infected cells. Inhibitory effect of saponin on HCV replication was verified by quantitative real-time PCR, reporter assay, and immunoblot analysis. In addition, saponin potentiated IFN-α-induced anti-HCV activity. Moreover, saponin exerted antiviral activity even in IFN-α resistant mutant HCVcc-infected cells. To investigate how cellular genes were regulated by saponin, we performed microarray analysis using HCVcc-infected cells. We demonstrated that suppressor of cytokine signaling 2 (SOCS2) protein level was distinctively increased by saponin, which in turn resulted in inhibition of HCV replication. We further showed that silencing of SOCS2 resurrected HCV replication and overexpression of SOCS2 suppressed HCV replication. These data imply that saponin inhibits HCV replication via SOCS2 signaling pathway. These findings suggest that saponin may be a potent therapeutic agent for HCV patients. PMID:22745742

  12. Understanding the tumor suppressor PTEN in chronic alcoholism and hepatocellular carcinoma.

    Science.gov (United States)

    Shearn, Colin T; Petersen, Dennis R

    2015-01-01

    The tumor suppressor phosphatase and tensin homolog deleted on chromosome 10 (PTEN) is a phosphatidylinositol (PtdIns) phosphatase that regulates Akt activation via PtdIns 3 kinase. Changes in PTEN expression and/or activity have been identified in a variety of chronic hepatocellular disorders including obesity, NAFLD, NASH, and alcoholism. In cancer biology, PTEN is frequently mutated or deleted in a wide variety of tumors. Mutations, decreased promoter activity, and decreased expression in PTEN are frequently identified in patients with hepatocellular carcinoma. While the majority of research on PTEN concerns obesity and NASH, PTEN clearly has a role in hepatic insulin sensitivity and in the development of steatosis during chronic alcoholism. Yet, in chronic alcoholics and HCC, very little is known concerning PTEN mutation/deletion or low PTEN expression. This review is focused on an overview of the current knowledge on molecular mechanisms of dysregulation of PTEN expression/activity in the liver and their relationship to development of ethanol-induced hepatocellular damage and cancer.

  13. Isoform-specific interactions of the von Hippel-Lindau tumor suppressor protein

    OpenAIRE

    Minervini, Giovanni; Mazzotta, Gabriella M.; Masiero, Alessandro; Sartori, Elena; Corr?, Samantha; Potenza, Emilio; Costa, Rodolfo; Tosatto, Silvio C. E.

    2015-01-01

    Deregulation of the von Hippel-Lindau tumor suppressor protein (pVHL) is considered one of the main causes for malignant renal clear-cell carcinoma (ccRCC) insurgence. In human, pVHL exists in two isoforms, pVHL19 and pVHL30 respectively, displaying comparable tumor suppressor abilities. Mutations of the p53 tumor suppressor gene have been also correlated with ccRCC insurgence and ineffectiveness of treatment. A recent proteomic analysis linked full length pVHL30 with p53 pathway regulation t...

  14. Functional Analysis of In-frame Indel ARID1A Mutations Reveals New Regulatory Mechanisms of Its Tumor Suppressor Functions

    Directory of Open Access Journals (Sweden)

    Bin Guan

    2012-10-01

    Full Text Available AT-rich interactive domain 1A (ARID1A has emerged as a new tumor suppressor in which frequent somatic mutations have been identified in several types of human cancers. Although most ARID1A somatic mutations are frame-shift or nonsense mutations that contribute to mRNA decay and loss of protein expression, 5% of ARID1A mutations are in-frame insertions or deletions (indels that involve only a small stretch of peptides. Naturally occurring in-frame indel mutations provide unique and useful models to explore the biology and regulatory role of ARID1A. In this study, we analyzed indel mutations identified in gynecological cancers to determine how these mutations affect the tumor suppressor function of ARID1A. Our results demonstrate that all in-frame mutants analyzed lost their ability to inhibit cellular proliferation or activate transcription of CDKN1A, which encodes p21, a downstream effector of ARID1A. We also showed that ARID1A is a nucleocytoplasmic protein whose stability depends on its subcellular localization. Nuclear ARID1A is less stable than cytoplasmic ARID1A because ARID1A is rapidly degraded by the ubiquitin-proteasome system in the nucleus. In-frame deletions affecting the consensus nuclear export signal reduce steady-state protein levels of ARID1A. This defect in nuclear exportation leads to nuclear retention and subsequent degradation. Our findings delineate a mechanism underlying the regulation of ARID1A subcellular distribution and protein stability and suggest that targeting the nuclear ubiquitin-proteasome system can increase the amount of the ARID1A protein in the nucleus and restore its tumor suppressor functions.

  15. A multicopy suppressor screening approach as a means to identify antibiotic resistance determinant candidates in Yersinia pestis

    Directory of Open Access Journals (Sweden)

    Moy Richard L

    2008-07-01

    Full Text Available Abstract Background Yersinia pestis is the causative agent of plague and a potential agent of bioterrorism and biowarfare. The plague biothreat and the emergence of multidrug-resistant plague underscore the need to increase our understanding of the intrinsic potential of Y. pestis for developing antimicrobial resistance and to anticipate the mechanisms of resistance that may emerge in Y. pestis. Identification of Y. pestis genes that, when overexpressed, are capable of reducing antibiotic susceptibility is a useful strategy to expose genes that this pathogen may rely upon to evolve antibiotic resistance via a vertical modality. In this study, we explored the use of a multicopy suppressor, Escherichia coli host-based screening approach as a means to expose antibiotic resistance determinant candidates in Y. pestis. Results We constructed a multicopy plasmid-based, Y. pestis genome-wide expression library of nearly 16,000 clones in E. coli and screened the library for suppressors of the antimicrobial activity of ofloxacin, a fluoroquinolone antibiotic. The screen permitted the identification of a transcriptional regulator-encoding gene (robAYp that increased the MIC99 of ofloxacin by 23-fold when overexpressed from a multicopy plasmid in Y. pestis. Additionally, we found that robAYp overexpression in Y. pestis conferred low-level resistance to many other antibiotics and increased organic solvent tolerance. Overexpression of robAYp also upregulated the expression of several efflux pumps in Y. pestis. Conclusion Our study provides proof of principle for the use of multicopy suppressor screening based on the tractable and easy-to-manipulate E. coli host as a means to identify antibiotic resistance determinant candidates of Y. pestis.

  16. Novel interactions of the von Hippel-Lindau (pVHL) tumor suppressor with the CDKN1 family of cell cycle inhibitors

    OpenAIRE

    Giovanni Minervini; Raffaele Lopreiato; Raissa Bortolotto; Antonella Falconieri; Geppo Sartori; Silvio C. E. Tosatto

    2017-01-01

    Germline inactivation of the von Hippel-Lindau (VHL) tumor suppressor predisposes patients to develop different highly vascularized cancers. pVHL targets the hypoxia-inducible transcription factor (HIF-1?) for degradation, modulating the activation of various genes involved in hypoxia response. Hypoxia plays a relevant role in regulating cell cycle progression, inducing growth arrest in cells exposed to prolonged oxygen deprivation. However, the exact molecular details driving this transition...

  17. PML tumor suppressor protein is required for HCV production

    Energy Technology Data Exchange (ETDEWEB)

    Kuroki, Misao [Department of Tumor Virology, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical Sciences, 2-5-1, Shikata-cho, Okayama 700-8558 (Japan); Research Fellow of the Japan Society for the Promotion of Science (Japan); Center for AIDS Research, Kumamoto University, Kumamoto 860-0811 (Japan); Ariumi, Yasuo, E-mail: ariumi@kumamoto-u.ac.jp [Department of Tumor Virology, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical Sciences, 2-5-1, Shikata-cho, Okayama 700-8558 (Japan); Center for AIDS Research, Kumamoto University, Kumamoto 860-0811 (Japan); Hijikata, Makoto [Department of Viral Oncology, Institute for Virus Research, Kyoto University, Kyoto 606-8507 (Japan); Ikeda, Masanori; Dansako, Hiromichi [Department of Tumor Virology, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical Sciences, 2-5-1, Shikata-cho, Okayama 700-8558 (Japan); Wakita, Takaji [Department of Virology II, National Institute of Infectious Diseases, Tokyo 162-8640 (Japan); Shimotohno, Kunitada [Research Center for Hepatitis and Immunology, National Center for Global Health and Medicine, Ichikawa, Chiba 272-8516 (Japan); Kato, Nobuyuki [Department of Tumor Virology, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical Sciences, 2-5-1, Shikata-cho, Okayama 700-8558 (Japan)

    2013-01-11

    Highlights: Black-Right-Pointing-Pointer PML tumor suppressor protein is required for HCV production. Black-Right-Pointing-Pointer PML is dispensable for HCV RNA replication. Black-Right-Pointing-Pointer HCV could not alter formation of PML-NBs. Black-Right-Pointing-Pointer INI1 and DDX5, PML-related proteins, are involved in HCV life cycle. -- Abstract: PML tumor suppressor protein, which forms discrete nuclear structures termed PML-nuclear bodies, has been associated with several cellular functions, including cell proliferation, apoptosis and antiviral defense. Recently, it was reported that the HCV core protein colocalizes with PML in PML-NBs and abrogates the PML function through interaction with PML. However, role(s) of PML in HCV life cycle is unknown. To test whether or not PML affects HCV life cycle, we examined the level of secreted HCV core and the infectivity of HCV in the culture supernatants as well as the level of HCV RNA in HuH-7-derived RSc cells, in which HCV-JFH1 can infect and efficiently replicate, stably expressing short hairpin RNA targeted to PML. In this context, the level of secreted HCV core and the infectivity in the supernatants from PML knockdown cells was remarkably reduced, whereas the level of HCV RNA in the PML knockdown cells was not significantly affected in spite of very effective knockdown of PML. In fact, we showed that PML is unrelated to HCV RNA replication using the subgenomic HCV-JFH1 replicon RNA, JRN/3-5B. Furthermore, the infectivity of HCV-like particle in the culture supernatants was significantly reduced in PML knockdown JRN/3-5B cells expressing core to NS2 coding region of HCV-JFH1 genome using the trans-packaging system. Finally, we also demonstrated that INI1 and DDX5, the PML-related proteins, are involved in HCV production. Taken together, these findings suggest that PML is required for HCV production.

  18. ZNF649, a novel Kruppel type zinc-finger protein, functions as a transcriptional suppressor

    International Nuclear Information System (INIS)

    Yang Hong; Yuan Wuzhou; Wang Ying; Zhu Chuanbing; Liu Bisheng; Wang Yuequn; Yang, Dan; Li Yongqing; Wang Canding; Wu Xiushan; Liu Mingyao

    2005-01-01

    Cardiac differentiation involves a cascade of coordinated gene expression that regulates cell proliferation and matrix protein formation in a defined temporo-spatial manner. Many of the KRAB-ZFPs are involved in cardiac development or cardiovascular diseases. Here we report the identification and characterization of a novel human zinc-finger gene named ZNF649. The cDNA of ZNF649 is 3176 bp, encoding a protein of 505 amino acids in the nuclei. Northern blot analysis indicates that ZNF649 is expressed in most of the examined human adult and embryonic tissues. ZNF649 is a transcription suppressor when fused to GAL-4 DNA-binding domain and cotransfected with VP-16. Overexpression of ZNF649 in COS-7 cells inhibits the transcriptional activities of SRE and AP-1. Deletion analysis with a series of truncated fusion proteins indicates that the KRAB motif is a basal repression domain when the truncated fusion proteins were assayed for the transcriptional activities of SRE and AP-1. These results suggest that ZNF649 protein may act as a transcriptional repressor in mitogen-activated protein kinase signaling pathway to mediate cellular functions

  19. Gemfibrozil, a Lipid-lowering Drug, Induces Suppressor of Cytokine Signaling 3 in Glial Cells

    Science.gov (United States)

    Ghosh, Arunava; Pahan, Kalipada

    2012-01-01

    Glial inflammation is an important feature of several neurodegenerative disorders. Suppressor of cytokine signaling (SOCS) proteins play a crucial role in inhibiting cytokine signaling and inflammatory gene expression in various cell types, including glial cells. However, mechanisms by which SOCS genes could be up-regulated are poorly understood. This study underlines the importance of gemfibrozil, a Food and Drug Administration-approved lipid-lowering drug, in up-regulating the expression of SOCS3 in glial cells. Gemfibrozil increased the expression of Socs3 mRNA and protein in mouse astroglia and microglia in both a time- and dose-dependent manner. Interestingly, gemfibrozil induced the activation of type IA phosphatidylinositol (PI) 3-kinase and AKT. Accordingly, inhibition of PI 3-kinase and AKT by chemical inhibitors abrogated gemfibrozil-mediated up-regulation of SOCS3. Furthermore, we demonstrated that gemfibrozil induced the activation of Krüppel-like factor 4 (KLF4) via the PI 3-kinase-AKT pathway and that siRNA knockdown of KLF4 abrogated gemfibrozil-mediated up-regulation of SOCS3. Gemfibrozil also induced the recruitment of KLF4 to the distal, but not proximal, KLF4-binding site of the Socs3 promoter. This study delineates a novel property of gemfibrozil in up-regulating SOCS3 in glial cells via PI 3-kinase-AKT-mediated activation of KLF4 and suggests that gemfibrozil may find therapeutic application in neuroinflammatory and neurodegenerative disorders. PMID:22685291

  20. Axl acts as a tumor suppressor by regulating LIGHT expression in T lymphoma

    Science.gov (United States)

    Young, Kon-Ji; Park, A-Reum; Choi, Ha-Rim; Lee, Hwa-Youn; Kim, Su-Man; Chung, Byung Yeoup; Park, Chul-Hong; Choi, Hyo Jin; Ko, Young-Hyeh; Bai, Hyoung-Woo; Kang, Hyung-Sik

    2017-01-01

    Axl is an oncogenic receptor tyrosine kinase that plays a role in many cancers. LIGHT (Lymphotoxin-related inducible ligand that competes for glycoprotein D binding to herpesvirus entry mediator on T cells) is a ligand that induces robust anti-tumor immunity by enhancing the recruitment and activation of effector immune cells at tumor sites. We observed that mouse EL4 and human Jurkat T lymphoma cells that stably overexpressed Axl also showed high expression of LIGHT. When Jurkat-Axl cells were treated with Gas6, a ligand for Axl, LIGHT expression was upregulated through activation of the PI3K/AKT signaling pathway and transcriptional induction by Sp1. The lytic activity of cytotoxic T lymphocytes and natural killer cells was enhanced by EL4-Axl cells. In addition, tumor volume and growth were markedly reduced due to enhanced apoptotic cell death in EL4-Axl tumor-bearing mice as compared to control mice. We also observed upregulated expression of CCL5 and its receptor, CCR5, and enhanced intratumoral infiltration of cytotoxic T lymphocytes and natural killer cells in EL4-Axl-bearing mice as compared to mock controls. These data strongly suggested that Axl exerts novel tumor suppressor effects by inducing upregulation of LIGHT in the tumor microenvironment of T lymphoma. PMID:28423548

  1. Tumor Suppressor p53 Stimulates the Expression of Epstein-Barr Virus Latent Membrane Protein 1.

    Science.gov (United States)

    Wang, Qianli; Lingel, Amy; Geiser, Vicki; Kwapnoski, Zachary; Zhang, Luwen

    2017-10-15

    Epstein-Barr virus (EBV) is associated with multiple human malignancies. EBV latent membrane protein 1 (LMP1) is required for the efficient transformation of primary B lymphocytes in vitro and possibly in vivo The tumor suppressor p53 plays a seminal role in cancer development. In some EBV-associated cancers, p53 tends to be wild type and overly expressed; however, the effects of p53 on LMP1 expression is not clear. We find LMP1 expression to be associated with p53 expression in EBV-transformed cells under physiological and DNA damaging conditions. DNA damage stimulates LMP1 expression, and p53 is required for the stimulation. Ectopic p53 stimulates endogenous LMP1 expression. Moreover, endogenous LMP1 blocks DNA damage-mediated apoptosis. Regarding the mechanism of p53-mediated LMP1 expression, we find that interferon regulatory factor 5 (IRF5), a direct target of p53, is associated with both p53 and LMP1. IRF5 binds to and activates a LMP1 promoter reporter construct. Ectopic IRF5 increases the expression of LMP1, while knockdown of IRF5 leads to reduction of LMP1. Furthermore, LMP1 blocks IRF5-mediated apoptosis in EBV-infected cells. All of the data suggest that cellular p53 stimulates viral LMP1 expression, and IRF5 may be one of the factors for p53-mediated LMP1 stimulation. LMP1 may subsequently block DNA damage- and IRF5-mediated apoptosis for the benefits of EBV. The mutual regulation between p53 and LMP1 may play an important role in EBV infection and latency and its related cancers. IMPORTANCE The tumor suppressor p53 is a critical cellular protein in response to various stresses and dictates cells for various responses, including apoptosis. This work suggests that an Epstein-Bar virus (EBV) principal viral oncogene is activated by cellular p53. The viral oncogene blocks p53-mediated adverse effects during viral infection and transformation. Therefore, the induction of the viral oncogene by p53 provides a means for the virus to cope with infection and

  2. Is the gene encoding Chibby implicated as a tumour suppressor in colorectal cancer ?

    International Nuclear Information System (INIS)

    Gad, Sophie; Teboul, David; Lièvre, Astrid; Goasguen, Nicolas; Berger, Anne; Beaune, Philippe; Laurent-Puig, Pierre

    2004-01-01

    A novel member of the Wnt signalling pathway, Chibby, was recently identified. This protein inhibits Wnt/β-catenin mediated transcriptional activation by competing with Lef-1 (the transcription factor and target of β-catenin) to bind to β-catenin. This suggests that Chibby could be a tumour suppressor protein. The C22orf2 gene coding Chibby is located on chromosome 22, a region recurrently lost in colorectal cancer. Activation of the Wnt pathway is a major feature of colorectal cancer and occurs through inactivation of APC or activation of β-catenin. All of this led us to analyse the possible implication of Chibby in colorectal carcinogenesis. First, 36 tumour and matched normal colonic mucosa DNA were genotyped with five microsatellite markers located on chromosome 22 to search for loss of heterozygosity. Then, mutation screening of the C22orf2 coding sequence and splice sites was performed in the 36 tumour DNA. Finally, expression of Chibby was analysed by quantitative RT-PCR on 10 patients, 4 with loss of heterozygosity (LOH) on chromosome 22. Loss of heterozygosity involving the C22orf2 region was detected in 11 out of 36 patients (30%). Sequencing analysis revealed a known variant, rs3747174, in exon 5: T321C leading to a silent amino acid polymorphism A107A. Allelic frequencies were 0.69 and 0.31 for T and C variants respectively. No other mutation was detected. Among the 10 patients studied, expression analysis revealed that Chibby is overexpressed in 2 tumours and underexpressed in 1. No correlations were found with 22q LOH status. As no somatic mutation was detected in C22orf2 in 36 colorectal tumour DNA, our results do not support the implication of Chibby as a tumour suppressor in colorectal carcinogenesis. This was supported by the absence of underexpression of Chibby among the tumour samples with 22q LOH. The implication of other Wnt pathway members remains to be identified to explain the part of colorectal tumours without mutation in APC and β-catenin

  3. A virus-sensitive suppressor cell is involved in the regulation of human allospecific T cell-mediated cytotoxicity

    International Nuclear Information System (INIS)

    Muluk, S.C.; Bernstein, D.C.; Shearer, G.M.

    1989-01-01

    The in vitro generation of allospecific CTL by human PBMC was enhanced 4- to 16-fold by sequential plastic and nylon wool adherence, which depleted the PBMC of macrophages and B cells. The enhanced CTL response was suppressed by adding back irradiated, unfractionated PBMC or adherent cells to the depleted cells. This finding suggests that the enhanced CTL response was not simply a consequence of enrichment of T cells, but was instead due to active suppression by radioresistant cells contained in the adherent fraction. Of note is the finding that, unlike the CTL response, the proliferative response to allostimulation was not affected by the removal of adherent cells. The suppressor function could be abrogated by preincubation of irradiated PBMC with influenza A virus before the coculture with depleted cells. Furthermore, costimulation of unfractionated PBMC with influenza A virus and allogeneic stimulators augmented allospecific CTL activity. Thus, in the adherent fraction of human PBMC, there appears to be a native suppressor population that can be functionally inactivated by virus. This result may account for the clinical observation of increased allograft rejection after certain viral infections

  4. Immune suppression with supraoptimal doses of antigen in contact sensitivity. I. Demonstration of suppressor cells and their sensitivity to cyclophosphamide.

    Science.gov (United States)

    Sy, M S; Miller, S D; Claman, H N

    1977-07-01

    Immunologic suppression was induced in a mouse model of contact sensitization to DNFB by using supraoptimal doses of antigen. In these studies, in vivo measurement of ear swelling as an indication of immunologic responsiveness correlated well with measurement of in vitro antigen-induced cell proliferation. This unresponsiveness was specific, since supraoptimal doses of DNFB did not interfere with the development of contact sensitivity to another contactant, oxazolone. The decrease in responsiveness is a form of active suppression, as lymphoid cells from supraoptimally sensitized donors transferred suppression to normal recipients. Furthermore, pretreatment with cyclophosphamide (Cy) reversed the suppression seen in supraoptimally sensitized animals but had no effect on the optimal sensitization regimen. These results indicate that supraoptimal doses of contactants can activate suppressor cells and that precursors of these cells are sensitive to Cy. Such suppressors regenerate within 7 to 14 days after Cy treatment. The ability of Cy pretreatment to affect supraoptimal sensitization without affecting optimal sensitization confirms other reports indicating that the observed results of Cy treatment depend critically upon the dose of antigen used.

  5. RDE-1 slicer activity is required only for passenger-strand cleavage during RNAi in Caenorhabditis elegans.

    Science.gov (United States)

    Steiner, Florian A; Okihara, Kristy L; Hoogstrate, Suzanne W; Sijen, Titia; Ketting, René F

    2009-02-01

    RNA interference (RNAi) is a process in which double-stranded RNA is cleaved into small interfering RNAs (siRNAs) that induce the destruction of homologous single-stranded mRNAs. Argonaute proteins are essential components of this silencing process; they bind siRNAs directly and can cleave RNA targets using a conserved RNase H motif. In Caenorhabditis elegans, the Argonaute protein RDE-1 has a central role in RNAi. In animals lacking RDE-1, the introduction of double-stranded RNA does not trigger any detectable level of RNAi. Here we show that RNase H activity of RDE-1 is required only for efficient removal of the passenger strand of the siRNA duplex and not for triggering the silencing response at the target-mRNA level. These results uncouple the role of the RDE-1 RNase H activity in small RNA maturation from its role in target-mRNA silencing in vivo.

  6. Suppressor Effects of Positive and Negative Religious Coping on Academic Burnout Among Korean Middle School Students.

    Science.gov (United States)

    Noh, Hyunkyung; Chang, Eunbi; Jang, Yoojin; Lee, Ji Hae; Lee, Sang Min

    2016-02-01

    Statistical suppressor effects in prediction models can provide evidence of the interdependent relationship of independent variables. In this study, the suppressor effects of positive and negative religious coping on academic burnout were examined using longitudinal data. First, 388 middle school students reported their type of religion and use of positive and negative religious coping strategies. Four months later, they also reported their level of academic burnout. From structural equation modeling, significant suppressor effects were found among religious students. That is, the coefficients became larger when both positive and negative religious coping predicted academic burnout simultaneously, compared to when each religious coping predicted academic burnout alone. However, suppressor effects were not found among non-religious students.

  7. Changes in helper and suppressor T lymphocytes following radiotherapy for breast cancer

    International Nuclear Information System (INIS)

    Newman, G.H.; Rees, G.J.G.; Jones, R.S.J.; Grove, E.A.; Preece, A.W.

    1987-01-01

    Changes in total lymphocyte, T lymphocyte, T helper and T suppressor lymphocyte numbers were studied in 22 patients with breast cancer before and after radiotherapy. T lymphocyte subsets were measured using monoclonal antibodies and fluorescence microscopy. After treatment the total lymphocyte count fell significantly and was still reduced 9 months later, but the proportion of cells labelled as T lymphocytes was unchanged during this period. The helper-suppressor ratio, which was within the normal range before radiotherapy, was significantly reduced at 3 months and 9 months after. Following treatment both T helper and T suppressor cell numbers were significantly reduced. T helper cell numbers remained reduced throughout the study period but T suppressor cell numbers showed a recovery to normal values 9 months after radiotherapy. (author)

  8. PGC-1α functions as a co-suppressor of XBP1s to regulate glucose metabolism

    Directory of Open Access Journals (Sweden)

    Jaemin Lee

    2018-01-01

    Full Text Available Objective: Peroxisome proliferator-activated receptor γ (PPARγ coactivator-1α (PGC-1α promotes hepatic gluconeogenesis by activating HNF4α and FoxO1. PGC-1α expression in the liver is highly elevated in obese and diabetic conditions, leading to increased hepatic glucose production. We previously showed that the spliced form of X-box binding protein 1 (XBP1s suppresses FoxO1 activity and hepatic gluconeogenesis. The shared role of PGC-1α and XBP1s in regulating FoxO1 activity and gluconeogenesis led us to investigate the probable interaction between PGC-1α and XBP1s and its role in glucose metabolism. Methods: We investigated the biochemical interaction between PGC-1α and XBP1s and examined the role of their interaction in glucose homeostasis using animal models. Results: We show that PGC-1α interacts with XBP1s, which plays an anti-gluconeogenic role in the liver by suppressing FoxO1 activity. The physical interaction between PGC-1α and XBP1s leads to suppression of XBP1s activity rather than its activation. Upregulating PGC-1α expression in the liver of lean mice lessens XBP1s protein levels, and reducing PGC-1α levels in obese and diabetic mouse liver restores XBP1s protein induction. Conclusions: Our findings reveal a novel function of PGC-1α as a suppressor of XBP1s function, suggesting that hepatic PGC-1α promotes gluconeogenesis through multiple pathways as a co-activator for HNF4α and FoxO1 and also as a suppressor for anti-gluconeogenic transcription factor XBP1s. Keywords: PGC-1α, XBP1s, Glucose homeostasis, ER stress, UPR, Insulin resistance

  9. Potential of lactic acid bacteria as suppressors of wine allergies

    Directory of Open Access Journals (Sweden)

    Yıldırım Hatice Kalkan

    2017-01-01

    Full Text Available Allergens causes some symptoms as all asthma, allergic conjunctivitis, and allergic rhinitis. These symptoms are seen twice as many in women than in men. The major wine allergens reported in wines are endochitinase 4A and lipid-transfer protein (LTP. This review deal with possibilities of using lactic acid bacteria as suppressors of wine allergies. Phenolic compounds present in wines have not only antioxidant properties causing radical scavenging but also some special properties reported in many in vitro studies as regulating functions in inflammatory cells as mast cells. So what is the role of lactic acid bacteria in these cases? Lactic acid bacteria are used during malolactic fermentation step of wine production with purpose of malic acid reduction. During this bioconversion complex phenolic compounds could be hydrolysed by bacterial enzymes to their aglycone forms. Obtained aglycons could pass through the intestinal epithelium of human and allowed reduction of IgE antibody production by affecting Th1/ Th2 ratio. Considering different contents and quantities of phenols in different grape varieties and consequently in different wines more studies are required in order to determine which lactic acid bacteria and strains could be effective in suppressing wine allergens.

  10. Myeloid-Derived Suppressor Cells and Therapeutic Strategies in Cancer

    Directory of Open Access Journals (Sweden)

    Hiroshi Katoh

    2015-01-01

    Full Text Available Development of solid cancer depends on escape from host immunosurveillance. Various types of immune cells contribute to tumor-induced immune suppression, including tumor associated macrophages, regulatory T cells, type 2 NKT cells, and myeloid-derived suppressor cells (MDSCs. Growing body of evidences shows that MDSCs play pivotal roles among these immunosuppressive cells in multiple steps of cancer progression. MDSCs are immature myeloid cells that arise from myeloid progenitor cells and comprise a heterogeneous immune cell population. MDSCs are characterized by the ability to suppress both adaptive and innate immunities mainly through direct inhibition of the cytotoxic functions of T cells and NK cells. In clinical settings, the number of circulating MDSCs is associated with clinical stages and response to treatment in several cancers. Moreover, MDSCs are reported to contribute to chemoresistant phenotype. Collectively, targeting MDSCs could potentially provide a rationale for novel treatment strategies in cancer. This review summarizes recent understandings of MDSCs in cancer and discusses promissing clinical approaches in cancer patients.

  11. Chemical suppressors of mlo-mediated powdery mildew resistance

    Science.gov (United States)

    Wu, Hongpo; Kwaaitaal, Mark; Strugala, Roxana; Schaffrath, Ulrich; Bednarek, Paweł

    2017-01-01

    Loss-of-function of barley mildew locus o (Mlo) confers durable broad-spectrum penetration resistance to the barley powdery mildew pathogen, Blumeria graminis f. sp. hordei (Bgh). Given the importance of mlo mutants in agriculture, surprisingly few molecular components have been identified to be required for this type of resistance in barley. With the aim to identify novel cellular factors contributing to mlo-based resistance, we devised a pharmacological inhibitor screen. Of the 41 rationally chosen compounds tested, five caused a partial suppression of mlo resistance in barley, indicated by increased levels of Bgh host cell entry. These chemicals comprise brefeldin A (BFA), 2′,3′-dideoxyadenosine (DDA), 2-deoxy-d-glucose, spermidine, and 1-aminobenzotriazole. Further inhibitor analysis corroborated a key role for both anterograde and retrograde endomembrane trafficking in mlo resistance. In addition, all four ribonucleosides, some ribonucleoside derivatives, two of the five nucleobases (guanine and uracil), some guanine derivatives as well as various polyamines partially suppress mlo resistance in barley via yet unknown mechanisms. Most of the chemicals identified to be effective in partially relieving mlo resistance in barley also to some extent compromised powdery mildew resistance in an Arabidopsis mlo2 mlo6 double mutant. In summary, our study identified novel suppressors of mlo resistance that may serve as valuable probes to unravel further the molecular processes underlying this unusual type of disease resistance. PMID:29127104

  12. RASSF6; the Putative Tumor Suppressor of the RASSF Family

    Directory of Open Access Journals (Sweden)

    Hiroaki Iwasa

    2015-12-01

    Full Text Available Humans have 10 genes that belong to the Ras association (RA domain family (RASSF. Among them, RASSF7 to RASSF10 have the RA domain in the N-terminal region and are called the N-RASSF proteins. In contradistinction to them, RASSF1 to RASSF6 are referred to as the C-RASSF proteins. The C-RASSF proteins have the RA domain in the middle region and the Salvador/RASSF/Hippo domain in the C-terminal region. RASSF6 additionally harbors the PSD-95/Discs large/ZO-1 (PDZ-binding motif. Expression of RASSF6 is epigenetically suppressed in human cancers and is generally regarded as a tumor suppressor. RASSF6 induces caspase-dependent and -independent apoptosis. RASSF6 interacts with mammalian Ste20-like kinases (homologs of Drosophila Hippo and cross-talks with the Hippo pathway. RASSF6 binds MDM2 and regulates p53 expression. The interactions with Ras and Modulator of apoptosis 1 (MOAP1 are also suggested by heterologous protein-protein interaction experiments. RASSF6 regulates apoptosis and cell cycle through these protein-protein interactions, and is implicated in the NF-κB and JNK signaling pathways. We summarize our current knowledge about RASSF6 and discuss what common and different properties RASSF6 and the other C-RASSF proteins have.

  13. Polerovirus protein P0 prevents the assembly of small RNA-containing RISC complexes and leads to degradation of ARGONAUTE1.

    Science.gov (United States)

    Csorba, Tibor; Lózsa, Rita; Hutvágner, György; Burgyán, József

    2010-05-01

    RNA silencing plays an important role in plants in defence against viruses. To overcome this defence, plant viruses encode suppressors of RNA silencing. The most common mode of silencing suppression is sequestration of double-stranded RNAs involved in the antiviral silencing pathways. Viral suppressors can also overcome silencing responses through protein-protein interaction. The poleroviral P0 silencing suppressor protein targets ARGONAUTE (AGO) proteins for degradation. AGO proteins are the core component of the RNA-induced silencing complex (RISC). We found that P0 does not interfere with the slicer activity of pre-programmed siRNA/miRNA containing AGO1, but prevents de novo formation of siRNA/miRNA containing AGO1. We show that the AGO1 protein is part of a high-molecular-weight complex, suggesting the existence of a multi-protein RISC in plants. We propose that P0 prevents RISC assembly by interacting with one of its protein components, thus inhibiting formation of siRNA/miRNA-RISC, and ultimately leading to AGO1 degradation. Our findings also suggest that siRNAs enhance the stability of co-expressed AGO1 in both the presence and absence of P0.

  14. ERF is a Potential ERK-Modulated Tumor Suppressor in Prostate Cancer

    Science.gov (United States)

    2017-10-01

    AWARD NUMBER: W81XWH-15-1-0277 TITLE: ERF is a Potential ERK-Modulated Tumor Suppressor in Prostate Cancer PRINCIPAL INVESTIGATOR: Dr...Rohit Bose CONTRACTING ORGANIZATION: Sloan Kettering Institute for Cancer Research New York NY 10065 REPORT DATE: October 2017 TYPE OF REPORT...4. TITLE AND SUBTITLE ERF is a Potential ERK-Modulated Tumor Suppressor in Prostate Cancer 5a. CONTRACT NUMBER 5b. GRANT NUMBER W81XWH-15-1-0277 5c

  15. ERF is a Potential ERK Modulated Tumor Suppressor in Prostate Cancer

    Science.gov (United States)

    2016-10-01

    6/27/2016 - 6/27/2019 1.20 calendar Prostate Cancer Foundation (formerly CaP CURE) $ 75,000 Epigenetic ...AWARD NUMBER: W81XWH-15-1-0277 TITLE: ERF is a Potential ERK-Modulated Tumor Suppressor in Prostate Cancer PRINCIPAL INVESTIGATOR: Dr. Rohit...4. TITLE AND SUBTITLE ERF is a Potential ERK-Modulated Tumor Suppressor in Prostate Cancer 5a. CONTRACT NUMBER 5b. GRANT NUMBER W81XWH-15-1-0277

  16. A Convenient In Vivo Model Using Small Interfering RNA Silencing to Rapidly Assess Skeletal Gene Function.

    Directory of Open Access Journals (Sweden)

    Wen Zhang

    Full Text Available It is difficult to study bone in vitro because it contains various cell types that engage in cross-talk. Bone biologically links various organs, and it has thus become increasingly evident that skeletal physiology must be studied in an integrative manner in an intact animal. We developed a model using local intraosseous small interfering RNA (siRNA injection to rapidly assess the effects of a target gene on the local skeletal environment. In this model, 160-g male Sprague-Dawley rats were treated for 1-2 weeks. The left tibia received intraosseous injection of a parathyroid hormone 1 receptor (Pth1r or insulin-like growth factor 1 receptor (Igf-1r siRNA transfection complex loaded in poloxamer 407 hydrogel, and the right tibia received the same volume of control siRNA. All the tibias received an intraosseous injection of recombinant human parathyroid hormone (1-34 (rhPTH (1-34 or insulin-like growth factor-1 (IGF-1. Calcein green and alizarin red were injected 6 and 2 days before euthanasia, respectively. IGF-1R and PTH1R expression levels were detected via RT-PCR assays and immunohistochemistry. Bone mineral density (BMD, microstructure, mineral apposition rates (MARs, and strength were determined by dual-energy X-ray absorptiometry, micro-CT, histology and biomechanical tests. The RT-PCR and immunohistochemistry results revealed that IGF-1R and PTH1R expression levels were dramatically diminished in the siRNA-treated left tibias compared to the right tibias (both p<0.05. Using poloxamer 407 hydrogel as a controlled-release system prolonged the silencing effect of a single dose of siRNA; the mRNA expression levels of IGF-1R were lower at two weeks than at one week (p<0.01. The BMD, bone microstructure parameters, MAR and bone strength were significantly decreased in the left tibias compared to the right tibias (all p<0.05. This simple and convenient local intraosseous siRNA injection model achieved gene silencing with very small quantities of siRNA over a short treatment period (≤7 days.

  17. Exonuclease-mediated degradation of nascent RNA silences genes linked to severe malaria

    DEFF Research Database (Denmark)

    Zhang, Qingfeng; Siegel, T Nicolai; Martins, Rafael M

    2014-01-01

    -coding RNA. The presence of stable upsA var transcripts overcomes monoallelic expression, resulting in the simultaneous expression of both upsA and upsC type PfEMP1 proteins on the surface of individual infected red blood cells. In addition, we observe an inverse relationship between transcript levels of Pf...

  18. Robust RNA silencing-mediated resistance to Plum pox virus under variable abiotic and biotic conditions.

    Science.gov (United States)

    Di Nicola, Elisa; Tavazza, Mario; Lucioli, Alessandra; Salandri, Laura; Ilardi, Vincenza

    2014-10-01

    Some abiotic and biotic conditions are known to have a negative impact on post-transcriptional gene silencing (PTGS), thus representing a potential concern for the production of stable engineered virus resistance traits. However, depending on the strategy followed to achieve PTGS of the transgene, different responses to external conditions can be expected. In the present study, we utilized the Nicotiana benthamiana–Plum pox virus (PPV) pathosystem to evaluate in detail the stability of intron-hairpin(ihp)-mediated virus resistance under conditions known to adversely affect PTGS. The ihp plants grown at low or high temperatures were fully resistant to multiple PPV challenges, different PPV inoculum concentrations and even to a PPV isolate differing from the ihp construct by more than 28% at the nucleotide level. In addition, infections of ihp plants with viruses belonging to Cucumovirus, Potyvirus or Tombusvirus, all known to affect PTGS at different steps, were not able to defeat PPV resistance. Low temperatures did not affect the accumulation of transgenic small interfering RNAs (siRNAs), whereas a clear increase in the amount of siRNAs was observed during infections sustained by Cucumber mosaic virus and Potato virus Y. Our results show that the above stress factors do not represent an important concern for the production,through ihp-PTGS technology, of transgenic plants having robust virus resistance traits.

  19. Structure of the Cmr2 Subunit of the CRISPR-Cas RNA Silencing Complex

    Energy Technology Data Exchange (ETDEWEB)

    Cocozaki, Alexis I.; Ramia, Nancy F.; Shao, Yaming; Hale, Caryn R.; Terns, Rebecca M.; Terns, Michael P.; Li, Hong (FSU); (Georgia)

    2012-08-10

    Cmr2 is the largest and an essential subunit of a CRISPR RNA-Cas protein complex (the Cmr complex) that cleaves foreign RNA to protect prokaryotes from invading genetic elements. Cmr2 is thought to be the catalytic subunit of the effector complex because of its N-terminal HD nuclease domain. Here, however, we report that the HD domain of Cmr2 is not required for cleavage by the complex in vitro. The 2.3 {angstrom} crystal structure of Pyrococcus furiosus Cmr2 (lacking the HD domain) reveals two adenylyl cyclase-like and two {alpha}-helical domains. The adenylyl cyclase-like domains are arranged as in homodimeric adenylyl cyclases and bind ADP and divalent metals. However, mutagenesis studies show that the metal- and ADP-coordinating residues of Cmr2 are also not critical for cleavage by the complex. Our findings suggest that another component provides the catalytic function and that the essential role by Cmr2 does not require the identified ADP- or metal-binding or HD domains in vitro.

  20. Transitive RNA silencing signals induce cytosine methylation of a transgenic but not an endogenous target

    Czech Academy of Sciences Publication Activity Database

    Vermeersch, L.; De Winne, N.; Nolf, J.; Bleys, A.; Kovařík, Aleš; Depicker, A.

    2013-01-01

    Roč. 74, č. 5 (2013), s. 867-879 ISSN 0960-7412 R&D Projects: GA ČR GBP501/12/G090 Institutional research plan: CEZ:AV0Z50040702 Institutional support: RVO:68081707 Keywords : DIRECTED DNA METHYLATION * POTATO-VIRUS-X * DOUBLE-STRANDED-RNA Subject RIV: BO - Biophysics Impact factor: 6.815, year: 2013

  1. Restoration of tumor suppressor miR-34 inhibits human p53-mutant gastric cancer tumorspheres

    International Nuclear Information System (INIS)

    Ji, Qing; Hao, Xinbao; Meng, Yang; Zhang, Min; DeSano, Jeffrey; Fan, Daiming; Xu, Liang

    2008-01-01

    MicroRNAs (miRNAs), some of which function as oncogenes or tumor suppressor genes, are involved in carcinogenesis via regulating cell proliferation and/or cell death. MicroRNA miR-34 was recently found to be a direct target of p53, functioning downstream of the p53 pathway as a tumor suppressor. miR-34 targets Notch, HMGA2, and Bcl-2, genes involved in the self-renewal and survival of cancer stem cells. The role of miR-34 in gastric cancer has not been reported previously. In this study, we examined the effects of miR-34 restoration on p53-mutant human gastric cancer cells and potential target gene expression. Human gastric cancer cells were transfected with miR-34 mimics or infected with the lentiviral miR-34-MIF expression system, and validated by miR-34 reporter assay using Bcl-2 3'UTR reporter. Potential target gene expression was assessed by Western blot for proteins, and by quantitative real-time RT-PCR for mRNAs. The effects of miR-34 restoration were assessed by cell growth assay, cell cycle analysis, caspase-3 activation, and cytotoxicity assay, as well as by tumorsphere formation and growth. Human gastric cancer Kato III cells with miR-34 restoration reduced the expression of target genes Bcl-2, Notch, and HMGA2. Bcl-2 3'UTR reporter assay showed that the transfected miR-34s were functional and confirmed that Bcl-2 is a direct target of miR-34. Restoration of miR-34 chemosensitized Kato III cells with a high level of Bcl-2, but not MKN-45 cells with a low level of Bcl-2. miR-34 impaired cell growth, accumulated the cells in G1 phase, increased caspase-3 activation, and, more significantly, inhibited tumorsphere formation and growth. Our results demonstrate that in p53-deficient human gastric cancer cells, restoration of functional miR-34 inhibits cell growth and induces chemosensitization and apoptosis, indicating that miR-34 may restore p53 function. Restoration of miR-34 inhibits tumorsphere formation and growth, which is reported to be

  2. Suppressor of fused (Sufu) promotes epithelial-mesenchymal transition (EMT) in cervical squamous cell carcinoma

    Science.gov (United States)

    Zhang, Ziyu; Zou, Yang; Liang, Meirong; Chen, Yuanting; Luo, Yong; Yang, Bicheng; Liu, Faying; Qin, Yunna; He, Deming; Wang, Feng; Huang, Ouping

    2017-01-01

    Suppressor of fused is essential for the maximal activation of Sonic Hedgehog signaling in development and tumorigenesis. However, the role of Sufu in cervical carcinoma remains unknown. Here, we report new findings of Sufu in regulating the epithelial-to-mesenchymal transition through the FoxM1 transcriptional modulation by 14-3-3ζ protein in cervical carcinoma. Sufu is overexpressed in cervical squamous cell carcinoma and its level in clinical tumor tissues is positively correlated with 14-3-3ζ. Functionanlly, siSufu remarkably prevents the cancer cell migration and invasion. We further demonstrate that the transcriptional activity of Sufu is increased by FoxM1, of which stability is promoted by 14-3-3ζ. Knockdown FoxM1 decreases the invasion of SiHa cells and reconstitution of Sufu rescues the invasion of these cells.Finally, overexpression of Sufu is significantly associated with differentiation grade, FIGO stage, Depth of stromal invasion and vascular cancer embolus. Our findings highlight a novel role for Sufu in cervical carcinogenesis. PMID:29371981

  3. Suppressor of cytokine signaling 1 expression during LPS-induced inflammation and bone loss in rats

    Directory of Open Access Journals (Sweden)

    João Antonio Chaves de SOUZA

    2017-10-01

    Full Text Available Abstract This study aimed to characterize the dynamics of suppressor of cytokine signaling (SOCS1 expression in a rat model of lipopolysaccharide-induced periodontitis. Wistar rats in the experimental groups were injected three times/week with LPS from Escherichia coli on the palatal aspect of the first molars, and control animals were injected with vehicle (phosphate-buffered saline. Animals were sacrificed 7, 15, and 30 days after the first injection to analyze inflammation (stereometric analysis, bone loss (macroscopic analysis, gene expression (qRT-PCR, and protein expression/activation (Western blotting. The severity of inflammation and bone loss associated with LPS-induced periodontitis increased from day 7 to day 15, and it was sustained through day 30. Significant (p < 0.05 increases in SOCS1, RANKL, OPG, and IFN-γ gene expression were observed in the experimental group versus the control group at day 15. SOCS1 protein expression and STAT1 and NF-κB activation were increased throughout the 30-day experimental period. Gingival tissues affected by experimental periodontitis express SOCS1, indicating that this protein may potentially downregulate signaling events involved in inflammatory reactions and bone loss and thus may play a relevant role in the development and progression of periodontal disease.

  4. Regulatory T cells as suppressors of anti-tumor immunity: Role of metabolism.

    Science.gov (United States)

    De Rosa, Veronica; Di Rella, Francesca; Di Giacomo, Antonio; Matarese, Giuseppe

    2017-06-01

    Novel concepts in immunometabolism support the hypothesis that glucose consumption is also used to modulate anti-tumor immune responses, favoring growth and expansion of specific cellular subsets defined in the past as suppressor T cells and currently reborn as regulatory T (Treg) cells. During the 1920s, Otto Warburg and colleagues observed that tumors consumed high amounts of glucose compared to normal tissues, even in the presence of oxygen and completely functioning mitochondria. However, the role of the Warburg Effect is still not completely understood, particularly in the context of an ongoing anti-tumor immune response. Current experimental evidence suggests that tumor-derived metabolic restrictions can drive T cell hyporesponsiveness and immune tolerance. For example, several glycolytic enzymes, deregulated in cancer, contribute to tumor progression independently from their canonical metabolic activity. Indeed, they can control apoptosis, gene expression and activation of specific intracellular pathways, thus suggesting a direct link between metabolic switches and pro-tumorigenic transcriptional programs. Focus of this review is to define the specific metabolic pathways controlling Treg cell immunobiology in the context of anti-tumor immunity and tumor progression. Copyright © 2017 Elsevier Ltd. All rights reserved.

  5. Analysis of the interaction between human RITA and Drosophila Suppressor of Hairless.

    Science.gov (United States)

    Brockmann, Birgit; Mastel, Helena; Oswald, Franz; Maier, Dieter

    2014-12-01

    Notch signalling mediates intercellular communication, which is effected by the transcription factor CSL, an acronym for vertebrate CBF1/RBP-J, Drosophila Suppressor of Hairless [Su(H)] and C. elegans Lag1. Nuclear import of CBF1/RBP-J depends on co-activators and co-repressors, whereas the export relies on RITA. RITA is a tubulin and CBF1/RBP-J binding protein acting as a negative regulator of Notch signalling in vertebrates. RITA protein is highly conserved in eumatazoa, but no Drosophila homologue was yet identified. In this work, the activity of human RITA in the fly was addressed. To this end, we generated transgenic flies that allow a tissue specific induction of human RITA, which was demonstrated by Western blotting and in fly tissues. Unexpectedly, overexpression of RITA during fly development had little phenotypic consequences, even when overexpressed simultaneously with either Su(H) or the Notch antagonist Hairless. We demonstrate the in vivo binding of human RITA to Su(H) and to tubulin by co-immune precipitation. Moreover, RITA and tubulin co-localized to some degree in several Drosophila tissues. Overall our data show that human RITA, albeit binding to Drosophila Su(H) and tubulin, cannot influence the Notch signalling pathway in the fly, suggesting that a nuclear export mechanism of Su(H), if existent in Drosophila, does not depend on RITA. © 2015 The Authors.

  6. Further evidence for poly-ADP-ribosylated histones as DNA suppressors

    International Nuclear Information System (INIS)

    Yu, F.L.; Geronimo, I.H.; Bender, W.; Meginniss, K.E.

    1986-01-01

    For many years histones have been considered to be the gene suppressors in eukaryotic cells. Recently, the authors have found strong evidence indicating that poly-ADP-ribosylated histones, rather than histones, are the potent inhibitors of DNA-dependent RNA synthesis. They now report additional evidence for this concept: 1) using histone inhibitor isolated directly from nuclei, the authors are able to confirm their earlier findings that the inhibitor substances are sensitive to pronase, snake venom phosphodiesterase digestion and 0.1N KOH hydrolysis, and are resistant to DNase I and RNase A digestion, 2) the O.D. 260/O.D.280 ratio of the histone inhibitor is between pure protein and nuclei acid, suggesting the inhibitor substance is a nucleoprotein hybrid. This result directly supports the fact that the isolated histone inhibitor is radioactive poly (ADP-ribose) labeled, 3) commercial histones show big differences in inhibitor activity. The authors believe this reflects the variation in poly-ADP-ribosylation among commercial histones, and 4) 0.1N KOH hydrolysis eliminates the poly (ADP-ribose) radioactivity from the acceptor proteins as well as histone inhibitor activity. Yet, on gel, the inhibitor shows identical histone bands and stain intensity before and after hydrolysis, indicating the histones per se are qualitatively and quantitatively unaffected by alkaline treatment. This result strongly suggests that histones themselves are not capable of inhibiting DNA-dependent RNA synthesis

  7. Coordinating ERK signaling via the molecular scaffold Kinase Suppressor of Ras [version 1; referees: 2 approved

    Directory of Open Access Journals (Sweden)

    Danielle Frodyma

    2017-08-01

    Full Text Available Many cancers, including those of the colon, lung, and pancreas, depend upon the signaling pathways induced by mutated and constitutively active Ras. The molecular scaffolds Kinase Suppressor of Ras 1 and 2 (KSR1 and KSR2 play potent roles in promoting Ras-mediated signaling through the Raf/MEK/ERK kinase cascade. Here we summarize the canonical role of KSR in cells, including its central role as a scaffold protein for the Raf/MEK/ERK kinase cascade, its regulation of various cellular pathways mediated through different binding partners, and the phenotypic consequences of KSR1 or KSR2 genetic inactivation. Mammalian KSR proteins have a demonstrated role in cellular and organismal energy balance with implications for cancer and obesity. Targeting KSR1 in cancer using small molecule inhibitors has potential for therapy with reduced toxicity to the patient. RNAi and small molecule screens using KSR1 as a reference standard have the potential to expose and target vulnerabilities in cancer. Interestingly, although KSR1 and KSR2 are similar in structure, KSR2 has a distinct physiological role in regulating energy balance. Although KSR proteins have been studied for two decades, additional analysis is required to elucidate both the regulation of these molecular scaffolds and their potent effect on the spatial and temporal control of ERK activation in health and disease.

  8. Reduction of Myeloid-derived Suppressor Cells and Lymphoma Growth by a Natural Triterpenoid

    Science.gov (United States)

    Radwan, Faisal F. Y.; Hossain, Azim; God, Jason M.; Leaphart, Nathan; Elvington, Michelle; Nagarkatti, Mitzi; Tomlinson, Stephen; Haque, Azizul

    2016-01-01

    Lymphoma is a potentially life threatening disease. The goal of this study was to investigate the therapeutic potential of a natural triterpenoid, Ganoderic acid A (GA-A) in controlling lymphoma growth both in vitro and in vivo. Here, we show that GA-A treatment induces caspase-dependent apoptotic cell death characterized by a dose-dependent increase in active caspases 9 and 3, up-regulation of pro-apoptotic BIM and BAX proteins, and a subsequent loss of mitochondrial membrane potential with release of cytochrome c. In addition to GA-A’s anti-growth activity, we show that lower doses of GA-A enhance HLA class II-mediated antigen presentation and CD4+ T cell recognition of lymphoma in vitro. The therapeutic relevance of GA-A treatment was also tested in vivo using the EL4 syngeneic mouse model of metastatic lymphoma. GA-A-treatment significantly prolonged survival of EL4 challenged mice and decreased tumor metastasis to the liver, an outcome accompanied by a marked down-regulation of STAT3 phosphorylation, reduction myeloid-derived suppressor cells (MDSCs), and enhancement of cytotoxic CD8+ T cells in the host. Thus, GA-A not only selectively induces apoptosis in lymphoma cells, but also enhances cell-mediated immune responses by attenuating MDSCs, and elevating Ag presentation and T cell recognition. The demonstrated therapeutic benefit indicates that GA-A is a candidate for future drug design for the treatment of lymphoma. PMID:25142864

  9. Enlightened protein: Fhit tumor suppressor protein structure and function and its role in the toxicity of protoporphyrin IX-mediated photodynamic reaction

    International Nuclear Information System (INIS)

    Zawacka-Pankau, Joanna

    2009-01-01

    The Fhit tumor suppressor protein possesses Ap 3 A (diadenosine triphosphate - ApppA) hydrolytic activity in vitro and its gene is found inactive in many pre-malignant states due to gene inactivation. For several years Fhit has been a widely investigated protein as its cellular function still remains largely unsolved. Fhit was shown to act as a molecular 'switch' of cell death via cascade operating on the influence of ATR-Chk1 pathway but also through the mitochondrial apoptotic pathway. Notably, Fhit was reported by our group to enhance the overall eradication effect of porphyrin-mediated photodynamic treatment (PDT). In this review the up-to-date findings on Fhit protein as a tumor suppressor and its role in PDT are presented.

  10. Analysis of Tospovirus NSs Proteins in Suppression of Systemic Silencing

    OpenAIRE

    Hedil, Marcio; Sterken, Mark G.; de Ronde, Dryas; Lohuis, Dick; Kormelink, Richard

    2015-01-01

    RNA silencing is a sequence-specific gene regulation mechanism that in plants also acts antiviral. In order to counteract antiviral RNA silencing, viruses have evolved RNA silencing suppressors (RSS). In the case of tospoviruses, the non-structural NSs protein has been identified as the RSS. Although the tomato spotted wilt virus (TSWV) tospovirus NSs protein has been shown to exhibit affinity to long and small dsRNA molecules, its ability to suppress the non-cell autonomous part of RNA silen...

  11. Phytochemical Compositions of Immature Wheat Bran, and Its Antioxidant Capacity, Cell Growth Inhibition, and Apoptosis Induction through Tumor Suppressor Gene

    Directory of Open Access Journals (Sweden)

    Mi Jeong Kim

    2016-09-01

    Full Text Available The purpose of this study was to investigate the phytochemical compositions and antioxidant capacity, cell growth inhibition, and apoptosis induction in extracts of immature wheat bran. Immature wheat bran (IWB was obtained from immature wheat harvested 10 days earlier than mature wheat. The phytochemical compositions of bran extract samples were analyzed by ultra-high performance liquid chromatography. The total ferulic acid (3.09 mg/g and p-coumaric acid (75 µg/g in IWB were significantly higher than in mature wheat bran (MWB, ferulic acid: 1.79 mg/g; p-coumaric acid: 55 µg/g. The oxygen radical absorbance capacity (ORAC: 327 µM Trolox equivalents (TE/g and cellular antioxidant activity (CAA: 4.59 µM Quercetin equivalents (QE/g of the IWB were higher than those of the MWB (ORAC: 281 µM TE/g; CAA: 0.63 µM QE/g. When assessing cell proliferation, the IWB extracts resulted in the lowest EC50 values against HT-29 (18.9 mg/mL, Caco-2 (7.74 mg/mL, and HeLa cells (8.17 mg/mL among bran extract samples. Additionally, the IWB extracts increased the gene expression of p53 and PTEN (tumor suppressor genes in HT-29 cells, indicating inhibited cell growth and induced apoptosis through tumor suppressor genes.

  12. Metastasis suppressor proteins in cutaneous squamous cell carcinoma.

    Science.gov (United States)

    Bozdogan, Onder; Vargel, Ibrahim; Cavusoglu, Tarik; Karabulut, Ayse A; Karahan, Gurbet; Sayar, Nilufer; Atasoy, Pınar; Yulug, Isik G

    2016-07-01

    Cutaneous squamous cell carcinomas (cSCCs) are common human carcinomas. Despite having metastasizing capacities, they usually show less aggressive progression compared to squamous cell carcinoma (SCC) of other organs. Metastasis suppressor proteins (MSPs) are a group of proteins that control and slow-down the metastatic process. In this study, we established the importance of seven well-defined MSPs including NDRG1, NM23-H1, RhoGDI2, E-cadherin, CD82/KAI1, MKK4, and AKAP12 in cSCCs. Protein expression levels of the selected MSPs were detected in 32 cSCCs, 6 in situ SCCs, and two skin cell lines (HaCaT, A-431) by immunohistochemistry. The results were evaluated semi-quantitatively using the HSCORE system. In addition, mRNA expression levels were detected by qRT-PCR in the cell lines. The HSCOREs of NM23-H1 were similar in cSCCs and normal skin tissues, while RGHOGDI2, E-cadherin and AKAP12 were significantly downregulated in cSCCs compared to normal skin. The levels of MKK4, NDRG1 and CD82 were partially conserved in cSCCs. In stage I SCCs, nuclear staining of NM23-H1 (NM23-H1nuc) was significantly lower than in stage II/III SCCs. Only nuclear staining of MKK4 (MKK4nuc) showed significantly higher scores in in situ carcinomas compared to invasive SCCs. In conclusion, similar to other human tumors, we have demonstrated complex differential expression patterns for the MSPs in in-situ and invasive cSCCs. This complex MSP signature warrants further biological and experimental pathway research. Copyright © 2016 Elsevier GmbH. All rights reserved.

  13. Genistein up-regulates tumor suppressor microRNA-574-3p in prostate cancer.

    Directory of Open Access Journals (Sweden)

    Takeshi Chiyomaru

    Full Text Available Genistein has been shown to inhibit cancers both in vitro and in vivo, by altering the expression of several microRNAs (miRNAs. In this study, we focused on tumor suppressor miRNAs regulated by genistein and investigated their function in prostate cancer (PCa and target pathways. Using miRNA microarray analysis and real-time RT-PCR we observed that miR-574-3p was significantly up-regulated in PCa cells treated with genistein compared with vehicle control. The expression of miR-574-3p was significantly lower in PCa cell lines and clinical PCa tissues compared with normal prostate cells (RWPE-1 and adjacent normal tissues. Low expression level of miR-574-3p was correlated with advanced tumor stage and higher Gleason score in PCa specimens. Re-expression of miR-574-3p in PCa cells significantly inhibited cell proliferation, migration and invasion in vitro and in vivo. miR-574-3p restoration induced apoptosis through reducing Bcl-xL and activating caspase-9 and caspase-3. Using GeneCodis software analysis, several pathways affected by miR-574-3p were identified, such as 'Pathways in cancer', 'Jak-STAT signaling pathway', and 'Wnt signaling pathway'. Luciferase reporter assays demonstrated that miR-574-3p directly binds to the 3' UTR of several target genes (such as RAC1, EGFR and EP300 that are components of 'Pathways in cancer'. Quantitative real-time PCR and Western analysis showed that the mRNA and protein expression levels of the three target genes in PCa cells were markedly down-regulated with miR-574-3p. Loss-of-function studies demonstrated that the three target genes significantly affect cell proliferation, migration and invasion in PCa cell lines. Our results show that genistein up-regulates tumor suppressor miR-574-3p expression targeting several cell signaling pathways. These findings enhance understanding of how genistein regulates with miRNA in PCa.

  14. NFκB1 is a suppressor of neutrophil-driven hepatocellular carcinoma

    Science.gov (United States)

    Wilson, C. L.; Jurk, D.; Fullard, N.; Banks, P.; Page, A.; Luli, S.; Elsharkawy, A. M.; Gieling, R. G.; Chakraborty, J. Bagchi; Fox, C.; Richardson, C.; Callaghan, K.; Blair, G. E.; Fox, N.; Lagnado, A.; Passos, J. F.; Moore, A. J.; Smith, G. R.; Tiniakos, D. G.; Mann, J.; Oakley, F.; Mann, D. A.

    2015-04-01

    Hepatocellular carcinoma (HCC) develops on the background of chronic hepatitis. Leukocytes found within the HCC microenvironment are implicated as regulators of tumour growth. We show that diethylnitrosamine (DEN)-induced murine HCC is attenuated by antibody-mediated depletion of hepatic neutrophils, the latter stimulating hepatocellular ROS and telomere DNA damage. We additionally report a previously unappreciated tumour suppressor function for hepatocellular nfkb1 operating via p50:p50 dimers and the co-repressor HDAC1. These anti-inflammatory proteins combine to transcriptionally repress hepatic expression of a S100A8/9, CXCL1 and CXCL2 neutrophil chemokine network. Loss of nfkb1 promotes ageing-associated chronic liver disease (CLD), characterized by steatosis, neutrophillia, fibrosis, hepatocyte telomere damage and HCC. Nfkb1S340A/S340Amice carrying a mutation designed to selectively disrupt p50:p50:HDAC1 complexes are more susceptible to HCC; by contrast, mice lacking S100A9 express reduced neutrophil chemokines and are protected from HCC. Inhibiting neutrophil accumulation in CLD or targeting their tumour-promoting activities may offer therapeutic opportunities in HCC.

  15. Long Non-coding RNA, PANDA, Contributes to the Stabilization of p53 Tumor Suppressor Protein.

    Science.gov (United States)

    Kotake, Yojiro; Kitagawa, Kyoko; Ohhata, Tatsuya; Sakai, Satoshi; Uchida, Chiharu; Niida, Hiroyuki; Naemura, Madoka; Kitagawa, Masatoshi

    2016-04-01

    P21-associated noncoding RNA DNA damage-activated (PANDA) is induced in response to DNA damage and represses apoptosis by inhibiting the function of nuclear transcription factor Y subunit alpha (NF-YA) transcription factor. Herein, we report that PANDA affects regulation of p53 tumor-suppressor protein. U2OS cells were transfected with PANDA siRNAs. At 72 h post-transfection, cells were subjected to immunoblotting and quantitative reverse transcription-polymerase chain reaction. Depletion of PANDA was associated with decreased levels of p53 protein, but not p53 mRNA. The stability of p53 protein was markedly reduced by PANDA silencing. Degradation of p53 protein by silencing PANDA was prevented by treatment of MG132, a proteasome inhibitor. Moreover, depletion of PANDA prevented accumulation of p53 protein, as a result of DNA damage, induced by the genotoxic agent etoposide. These results suggest that PANDA stabilizes p53 protein in response to DNA damage, and provide new insight into the regulatory mechanisms of p53. Copyright© 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  16. Recurrent pregnancy failure is associated with a polymorphism in the p53 tumour suppressor gene.

    Science.gov (United States)

    Pietrowski, Detlef; Bettendorf, Hertha; Riener, Eva-Katrin; Keck, Christoph; Hefler, Lukas A; Huber, Johannes C; Tempfer, Clemens

    2005-04-01

    The p53 tumour suppressor gene is a well-known factor regulating apoptosis in a wide variety of cells and tissues. Alterations in the p53 gene are among the most common genetic changes in human cancers. In addition, recent data provide evidence that p53 plays a critical role in mediating pregnancy by regulating steroid hormone activation. In idiopathic recurrent miscarriages (IRM), causes and associations are much debated as the exact pathophysiological mechanisms are unknown. In this study, we assess whether an established polymorphism in the p53 gene is associated with the occurrence of IRM. Genotyping was performed by PCR-based amplification of the p53 Arg and Pro variants at codon 72 in 175 cases of IRM and 143 controls. We observed a statistically significant association between carriage of the Pro allele and the occurrence of IRM (P = 0.03, odds ratio 1.49, confidence interval 1.04-2.14). Distribution of genotypes was in Hardy-Weinberg equilibrium. Our results indicate an over-representation of the Pro allele of the p53 gene in women with IRM, giving support to the theory that p53 has a potential role during pregnancy.

  17. CCR5 is a suppressor for cortical plasticity and hippocampal learning and memory.

    Science.gov (United States)

    Zhou, Miou; Greenhill, Stuart; Huang, Shan; Silva, Tawnie K; Sano, Yoshitake; Wu, Shumin; Cai, Ying; Nagaoka, Yoshiko; Sehgal, Megha; Cai, Denise J; Lee, Yong-Seok; Fox, Kevin; Silva, Alcino J

    2016-12-20

    Although the role of CCR5 in immunity and in HIV infection has been studied widely, its role in neuronal plasticity, learning and memory is not understood. Here, we report that decreasing the function of CCR5 increases MAPK/CREB signaling, long-term potentiation (LTP), and hippocampus-dependent memory in mice, while neuronal CCR5 overexpression caused memory deficits. Decreasing CCR5 function in mouse barrel cortex also resulted in enhanced spike timing dependent plasticity and consequently, dramatically accelerated experience-dependent plasticity. These results suggest that CCR5 is a powerful suppressor for plasticity and memory, and CCR5 over-activation by viral proteins may contribute to HIV-associated cognitive deficits. Consistent with this hypothesis, the HIV V3 peptide caused LTP, signaling and memory deficits that were prevented by Ccr5 knockout or knockdown. Overall, our results demonstrate that CCR5 plays an important role in neuroplasticity, learning and memory, and indicate that CCR5 has a role in the cognitive deficits caused by HIV.

  18. Potential role of estrogen receptor beta as a tumor suppressor of epithelial ovarian cancer.

    Directory of Open Access Journals (Sweden)

    Carine Bossard

    Full Text Available Ovarian cancer is the gynecological cancer exhibiting the highest morbidity and improvement of treatments is still required. Previous studies have shown that Estrogen-receptor beta (ERβ levels decreased along with ovarian carcinogenesis. Here, we present evidence that reintroduction of ERβ in BG-1 epithelial ovarian cancer cells, which express ERα, leads in vitro to a decrease of basal and estradiol-promoted cell proliferation. ERβ reduced the frequency of cells in S phase and increased the one of cells in G2/M phase. At the molecular level, we found that ERβ downregulated total retinoblastoma (Rb, phosphorylated Rb and phospho-AKT cellular content as well as cyclins D1 and A2. In addition, ERβ had a direct effect on ERα, by strongly inhibiting its expression and activity, which could explain part of the anti-proliferative action of ERβ. By developing a novel preclinical model of ovarian cancer based on a luminescent orthotopic xenograft in athymic Nude mice, we further revealed that ERβ expression reduces tumor growth and the presence of tumor cells in sites of metastasis, hence resulting in improved survival of mice. Altogether, these findings unveil a potential tumor-suppressor role of ERβ in ovarian carcinogenesis, which could be of potential clinical relevance for the selection of the most appropriate treatment for patients.

  19. Potential Role of Estrogen Receptor Beta as a Tumor Suppressor of Epithelial Ovarian Cancer

    Science.gov (United States)

    Gaudin, Françoise; Machelon, Véronique; Brigitte, Madly; Jacquard, Carine; Pillon, Arnaud; Balaguer, Patrick; Balabanian, Karl; Lazennec, Gwendal

    2012-01-01

    Ovarian cancer is the gynecological cancer exhibiting the highest morbidity and improvement of treatments is still required. Previous studies have shown that Estrogen-receptor beta (ERβ) levels decreased along with ovarian carcinogenesis. Here, we present evidence that reintroduction of ERβ in BG-1 epithelial ovarian cancer cells, which express ERα, leads in vitro to a decrease of basal and estradiol-promoted cell proliferation. ERβ reduced the frequency of cells in S phase and increased the one of cells in G2/M phase. At the molecular level, we found that ERβ downregulated total retinoblastoma (Rb), phosphorylated Rb and phospho-AKT cellular content as well as cyclins D1 and A2. In addition, ERβ had a direct effect on ERα, by strongly inhibiting its expression and activity, which could explain part of the anti-proliferative action of ERβ. By developing a novel preclinical model of ovarian cancer based on a luminescent orthotopic xenograft in athymic Nude mice, we further revealed that ERβ expression reduces tumor growth and the presence of tumor cells in sites of metastasis, hence resulting in improved survival of mice. Altogether, these findings unveil a potential tumor-suppressor role of ERβ in ovarian carcinogenesis, which could be of potential clinical relevance for the selection of the most appropriate treatment for patients. PMID:22970307

  20. Specificity and autoregulation of Notch binding by tandem WW domains in suppressor of Deltex.

    Science.gov (United States)

    Jennings, Martin D; Blankley, Richard T; Baron, Martin; Golovanov, Alexander P; Avis, Johanna M

    2007-09-28

    WW domains target proline-tyrosine (PY) motifs and frequently function as tandem pairs. When studied in isolation, single WW domains are notably promiscuous and regulatory mechanisms are undoubtedly required to ensure selective interactions. Here, we show that the fourth WW domain (WW4) of Suppressor of Deltex, a modular Nedd4-like protein that down-regulates the Notch receptor, is the primary mediator of a direct interaction with a Notch-PY motif. A natural Trp to Phe substitution in WW4 reduces its affinity for general PY sequences and enhances selective interaction with the Notch-PY motif via compensatory specificity-determining interactions with PY-flanking residues. When WW4 is paired with WW3, domain-domain association, impeding proper folding, competes with Notch-PY binding to WW4. This novel mode of autoinhibition is relieved by binding of another ligand to WW3. Such cooperativity may facilitate the transient regulatory interactions observed in vivo between Su(dx) and Notch in the endocytic pathway. The highly conserved tandem arrangement of WW domains in Nedd4 proteins, and similar arrangements in more diverse proteins, suggests domain-domain communication may be integral to regulation of their associated cellular activities.

  1. Mechanisms maintaining enhancement of allografts. I. Demonstration of a specific suppressor cell

    International Nuclear Information System (INIS)

    Hall, B.M.

    1985-01-01

    DA rats treated with hyperimmune anti-PVG serum and grafted with (DA X PVG)F1 heart grafts in which graft survival was prolonged for greater than 75 d were used to examine the cellular mechanisms that maintain the state of specific unresponsiveness found in these animals. The capacity of lymphocytes from these animals to effect or inhibit graft rejection on adoptive transfer to irradiated heart-grafted hosts was tested. Spleen cell populations and the T cell subpopulation separated from spleen cells in vitro failed to restore rejection of PVG heart grafts in irradiated DA recipients but restored third party Lew graft rejection. Whole spleen cells had the capacity to suppress the ability of normal DA LNC to cause graft rejection, but T cells from spleen only delayed the restoration of rejection. LNC and recirculating T cells from rats with enhanced grafts adoptively restored PVG rejection, however. These studies show that the state of specific unresponsiveness that follows the induction of passive enhancement is dependent in part upon active suppression, which is induced or mediated by T lymphocytes. The recirculating pool of lymphocytes in these animals is not depleted of specific alloreactive cells with the capacity to initiate and effect rejection. Thus, these animals responsiveness is not like that found in transplantation tolerance induced in neonatal rats, but is, in part, due to a suppressor response that can inhibit normal alloreactive cells capacity to initiate and effect rejection

  2. Targeting Myeloid-Derived Suppressor Cells to Bypass Tumor-Induced Immunosuppression

    Directory of Open Access Journals (Sweden)

    Viktor Fleming

    2018-03-01

    Full Text Available The immune system has many sophisticated mechanisms to balance an extensive immune response. Distinct immunosuppressive cells could protect from excessive tissue damage and autoimmune disorders. Tumor cells take an advantage of those immunosuppressive mechanisms and establish a strongly immunosuppressive tumor microenvironment (TME, which inhibits antitumor immune responses, supporting the disease progression. Myeloid-derived suppressor cells (MDSC play a crucial role in this immunosuppressive TME. Those cells represent a heterogeneous population of immature myeloid cells with a strong immunosuppressive potential. They inhibit an antitumor reactivity of T cells and NK cells. Furthermore, they promote angiogenesis, establish pre-metastatic niches, and recruit other immunosuppressive cells such as regulatory T cells. Accumulating evidences demonstrated that the enrichment and activation of MDSC correlated with tumor progression, recurrence, and negative clinical outcome. In the last few years, various preclinical studies and clinical trials targeting MDSC showed promising results. In this review, we discuss different therapeutic approaches on MDSC targeting to overcome immunosuppressive TME and enhance the efficiency of current tumor immunotherapies.

  3. Requirement of the ATM/p53 tumor suppressor pathway for glucose homeostasis.

    Science.gov (United States)

    Armata, Heather L; Golebiowski, Diane; Jung, Dae Young; Ko, Hwi Jin; Kim, Jason K; Sluss, Hayla K

    2010-12-01

    Ataxia telangiectasia (A-T) patients can develop multiple clinical pathologies, including neuronal degeneration, an elevated risk of cancer, telangiectasias, and growth retardation. Patients with A-T can also exhibit an increased risk of insulin resistance and type 2 diabetes. The ATM protein kinase, the product of the gene mutated in A-T patients (Atm), has been implicated in metabolic disease, which is characterized by insulin resistance and increased cholesterol and lipid levels, blood pressure, and atherosclerosis. ATM phosphorylates the p53 tumor suppressor on a site (Ser15) that regulates transcription activity. To test whether the ATM pathway that regulates insulin resistance is mediated by p53 phosphorylation, we examined insulin sensitivity in mice with a germ line mutation that replaces the p53 phosphorylation site with alanine. The loss of p53 Ser18 (murine Ser15) led to increased metabolic stress, including severe defects in glucose homeostasis. The mice developed glucose intolerance and insulin resistance. The insulin resistance correlated with the loss of antioxidant gene expression and decreased insulin signaling. N-Acetyl cysteine (NAC) treatment restored insulin signaling in late-passage primary fibroblasts. The addition of an antioxidant in the diet rendered the p53 Ser18-deficient mice glucose tolerant. This analysis demonstrates that p53 phosphorylation on an ATM site is an important mechanism in the physiological regulation of glucose homeostasis.

  4. Characterization of cell-surface receptors for monoclonal-nonspecific suppressor factor (MNSF)

    International Nuclear Information System (INIS)

    Nakamura, M.; Ogawa, H.; Tsunematsu, T.

    1990-01-01

    Monoclonal-nonspecific suppressor factor (MNSF) is a lymphokine derived from murine T cell hybridoma. The target tissues are both LPS-stimulated B cells and Con A-stimulated T cells. Since the action of MNSF may be mediated by its binding to specific cell surface receptors, we characterized the mode of this binding. The purified MNSF was labeled with 125 I, using the Bolton-Hunter reagent. The labeled MNSF bound specifically to a single class of receptor (300 receptors per cell) on mitogen-stimulated murine B cells or T cells with an affinity of 16 pM at 24 degrees C, in the presence of sodium azide. Competitive experiments showed that MNSF bound to the specific receptor and that the binding was not shared with IL2, IFN-gamma, and TNF. Various cell types were surveyed for the capacity to specifically bind 125 I-MNSF. 125 I-MNSF bound to MOPC-31C (a murine plasmacytoma line) and to EL4 (a murine T lymphoma line). The presence of specific binding correlates with the capacity of the cells to respond to MNSF. These data support the view that like other polypeptide hormones, the action of MNSF is mediated by specific cell surface membrane receptor protein. Identification of these receptors will provide insight into the apparently diverse activities of MNSF

  5. Pam2 lipopeptides systemically increase myeloid-derived suppressor cells through TLR2 signaling

    Energy Technology Data Exchange (ETDEWEB)

    Maruyama, Akira; Shime, Hiroaki, E-mail: shime@med.hokudai.ac.jp; Takeda, Yohei; Azuma, Masahiro; Matsumoto, Misako; Seya, Tsukasa, E-mail: seya-tu@pop.med.hokudai.ac.jp

    2015-02-13

    Myeloid-derived suppressor cells (MDSCs) are immature myeloid cells that exhibit potent immunosuppressive activity. They are increased in tumor-bearing hosts and contribute to tumor development. Toll-like receptors (TLRs) on MDSCs may modulate the tumor-supporting properties of MDSCs through pattern-recognition. Pam2 lipopeptides represented by Pam2CSK4 serve as a TLR2 agonist to exert anti-tumor function by dendritic cell (DC)-priming that leads to NK cell activation and cytotoxic T cell proliferation. On the other hand, TLR2 enhances tumor cell progression/invasion by activating tumor-infiltrating macrophages. How MDSCs respond to TLR2 agonists has not yet been determined. In this study, we found intravenous administration of Pam2CSK4 systemically up-regulated the frequency of MDSCs in EG7 tumor-bearing mice. The frequency of tumor-infiltrating MDSCs was accordingly increased in response to Pam2CSK4. MDSCs were not increased by Pam2CSK4 stimuli in TLR2 knockout (KO) mice. Adoptive transfer experiments using CFSE-labeled MDSCs revealed that the TLR2-positive MDSCs survived long in tumor-bearing mice in response to Pam2CSK4 treatment. Since the increased MDSC population sustained immune-suppressive properties, our study suggests that Pam2CSK4-triggered TLR2 activation enhances the MDSC potential and suppress antitumor immune response in tumor microenvironment. - Highlights: • Pam2CSK4 administration induces systemic accumulation of CD11b{sup +}Gr1{sup +} MDSCs. • TLR2 is essential for Pam2CSK4-induced accumulation of CD11b{sup +}Gr1{sup +} MDSCs. • Pam2CSK4 supports survival of CD11b{sup +}Gr1{sup +} MDSCs in vivo.

  6. LACTB, a novel epigenetic silenced tumor suppressor, inhibits colorectal cancer progression by attenuating MDM2-mediated p53 ubiquitination and degradation.

    Science.gov (United States)

    Zeng, Kaixuan; Chen, Xiaoxiang; Hu, Xiuxiu; Liu, Xiangxiang; Xu, Tao; Sun, Huiling; Pan, Yuqin; He, Bangshun; Wang, Shukui

    2018-06-13

    Colorectal cancer (CRC) is one of the most common aggressive malignancies. Like other solid tumors, inactivation of tumor suppressor genes and activation of oncogenes occur during CRC development and progression. Recently, a novel tumor suppressor, LACTB, was proposed to inhibit tumor progression, but the functional and clinical significance of this tumor suppressor in CRC remains unexplored. Herein, we found LACTB was significantly downregulated in CRC due to promoter methylation and histone deacetylation, which was associated with metastasis and advanced clinical stage. CRC patients with low LACTB expression had poorer overall survival and LACTB also determined to be an independent prognostic factor for poorer outcome. Ectopic expression of LACTB suppressed CRC cells proliferation, migration, invasion, and epithelial-mesenchymal transition (EMT) in vitro and inhibited CRC growth and metastasis in vivo, while knockout of LACTB by CRISPR/Cas9 gene editing technique resulted in an opposite phenotype. Interestingly, LACTB could exert antitumorigenic effect only in HCT116 and HCT8 cells harboring wild-type TP53, but not in HT29 and SW480 cells harboring mutant TP53 or HCT116 p53 -/- cells. Mechanistic studies demonstrated that LACTB could directly bind to the C terminus of p53 to inhibit p53 degradation by preventing MDM2 from interacting with p53. Moreover, ablation of p53 attenuated the antitumorigenic effects of LACTB overexpression in CRC. Collectively, our findings successfully demonstrate for the first time that LACTB is a novel epigenetic silenced tumor suppressor through modulating the stability of p53, supporting the pursuit of LACTB as a potential therapeutic target for CRC.

  7. Antihelminthic drug niclosamide inhibits CIP2A and reactivates tumor suppressor protein phosphatase 2A in non-small cell lung cancer cells.

    Science.gov (United States)

    Kim, Myeong-Ok; Choe, Min Ho; Yoon, Yi Na; Ahn, Jiyeon; Yoo, Minjin; Jung, Kwan-Young; An, Sungkwan; Hwang, Sang-Gu; Oh, Jeong Su; Kim, Jae-Sung

    2017-11-15

    Protein phosphatase 2A (PP2A) is a critical tumor suppressor complex responsible for the inactivation of various oncogenes. Recently, PP2A reactivation has emerged asan anticancer strategy. Cancerous inhibitor of protein phosphatase 2A (CIP2A), an endogenous inhibitor of PP2A, is upregulated in many cancer cells, including non-small cell lung cancer (NSCLC) cells. We demonstrated that the antihelminthic drug niclosamide inhibited the expression of CIP2A and reactivated the tumor suppressor PP2A in NSCLC cells. We performed a drug-repurposing screen and identified niclosamide asa CIP2A suppressor in NSCLC cells. Niclosamide inhibited cell proliferation, colony formation, and tumor sphere formation, and induced mitochondrial dysfunction through increased mitochondrial ROS production in NSCLC cells; however, these effects were rescued by CIP2A overexpression, which indicated that the antitumor activity of niclosamide was dependent on CIP2A. We found that niclosamide increased PP2A activity through CIP2A inhibition, which reduced the phosphorylation of several oncogenic proteins. Moreover, we found that a niclosamide analog inhibited CIP2A expression and increased PP2A activity in several types of NSCLC cells. Finally, we showed that other well-known PP2A activators, including forskolin and FTY720, did not inhibit CIP2A and that their activities were not dependent on CIP2A. Collectively, our data suggested that niclosamide effectively suppressed CIP2A expression and subsequently activated PP2A in NSCLC cells. This provided strong evidence for the potential use of niclosamide asa PP2A-activating drug in the clinical treatment of NSCLC. Copyright © 2017 Elsevier Inc. All rights reserved.

  8. Suppressor cells in transplantation tolerance. III. The role of antigen in the maintenance of transplantation tolerance

    International Nuclear Information System (INIS)

    Tutschka, P.J.; Hess, A.D.; Beschorner, W.E.; Santos, G.W.

    1982-01-01

    Suppressor cells, which in an alloantigen-specific manner inhibit proliferation of donor cells to host antigens in a mixed lymphocyte culture and adoptively transfer the suppression of graft-versus-host disease (GVHD), undergo a gradual clonal reduction in long-term, allogeneic, histoincompatible rat radiation chimeras until they can no longer be measured in an in vitro suppressor cell assay. When lymphohematopoietic cells from these chimeras are transferred into lethally irradiated secondary recipients of original donor strain, the suppressor cells, now in a target antigen-free environment, undergo a further clonal reduction. After parking for 120 days, the chimeric cells are specifically tolerant to original host antigens, but cannot adoptively transfer suppression of GVHD. When chimeric cells, parked for 120 days in secondary recipients of original donor strain, are stimulated with original host-type antigen repeatedly during or once at the end of the parking period, the suppressor cell clone is expanded, suppressor cells can be identified in vitro, and suppression of GVHD can adoptively be transferred to tertiary recipients

  9. Power consumption in positive ion beam converter with electrostatic electron suppressor

    International Nuclear Information System (INIS)

    Hashimoto, Kiyoshi; Sugawara, Tohru

    1985-01-01

    The power recovery characteristics of an in-line direct beam converter provided with electrostatic electron suppressor were studied numerically by tracing the orbits of fast primary ions and secondary charged particles generated along their beam path by collision with background gas molecules. It is shown that, in reference to the electrostatic field potential at the point of impact, the energy distribution of secondary ions impinging on the suppressor has two peaks-one corresponding to a zone of high positive potential surrounding the collector and the other to one of slightly negative potential around the electron suppressor. Secondary electron emission from the suppressor is ascribed mainly to the latter peak, associated with impingement of slower secondary ions. Far much power consumed in secondary particle acceleration is spent for emitting electrons from the suppressor than for secondary ions generated by beam-gas collision. The upper limit of background pressure is discussed on the basis of criteria prescribed for restricting the power consumed in this secondary particle acceleration, as for practical convenience of electrode cooling. Numerical examples are given of calculations based on particle trajectory analysis of both primary ions and secondary particles, for the case of a 100 keV-proton sheet beam 10 cm thick of 35 mA/cm 2 current density. (author)

  10. Photoreactivation of conversion and de novo suppressor mutation in Escherichia coli

    Energy Technology Data Exchange (ETDEWEB)

    Bockrath, R C; Plamer, J E [Indiana Univ., Indianapolis (USA). Dept. of Microbiology

    1977-04-01

    Studies of mutagenesis and photoreactivation in various E.coli strains have shown that conversion mutation of a mutant containing an amber suppressor to one containing an ochre suppressor is sensitive to photoreactivation. Direct photoreactivation by photoreactivating light (PRL) after uv mutagenesis reduced mutation frequencies by a factor of about 2 for each minute of exposure during the first 5 to 8 min of exposure for cells with normal repair capacity. Conversion and potential de novo suppressor mutations were about equally sensitive. For conversion, the sensitivities to PRL were identical in the repair-normal and excisions-repair-deficient strains. For de novo suppressor mutation, the rate of mutation frequency reduction by PRL in the repair-deficient strain was about one-half that in the other strains. The results suggest that ultraviolet radiation produces both de novo suppressor mutation and conversion at the sup(E,B) locus by photoreversible pyrimidine dimers in the DNA. The causative dimers could be Thy()Cyt dimers in the transcribed strand or the non-transcribed strand, respectively.

  11. Effect of hydroxyurea on the promoter occupancy profiles of tumor suppressor p53 and p73

    Directory of Open Access Journals (Sweden)

    Lu Xin

    2009-06-01

    Full Text Available Abstract Background The p53 tumor suppressor and its related protein, p73, share a homologous DNA binding domain, and mouse genetics studies have suggested that they have overlapping as well as distinct biological functions. Both p53 and p73 are activated by genotoxic stress to regulate an array of cellular responses. Previous studies have suggested that p53 and p73 independently activate the cellular apoptotic program in response to cytotoxic drugs. The goal of this study was to compare the promoter-binding activity of p53 and p73 at steady state and after genotoxic stress induced by hydroxyurea. Results We employed chromatin immunoprecipitation, the NimbleGen promoter arrays and a model-based algorithm for promoter arrays to identify promoter sequences enriched in anti-p53 or anti-p73 immunoprecipitates, either before or after treatment with hydroxyurea, which increased the expression of both p53 and p73 in the human colon cancer cell line HCT116-3(6. We calculated a model-based algorithm for promoter array score for each promoter and found a significant correlation between the promoter occupancy profiles of p53 and p73. We also found that after hydroxyurea treatment, the p53-bound promoters were still bound by p73, but p73 became associated with additional promoters that that did not bind p53. In particular, we showed that hydroxyurea induces the binding of p73 but not p53 to the promoter of MLH3, which encodes a mismatch repair protein, and causes an up-regulation of the MLH3 mRNA. Conclusion These results suggest that hydroxyurea exerts differential effects on the promoter-binding functions of p53 and p73 and illustrate the power of model-based algorithm for promoter array in the analyses of promoter occupancy profiles of highly homologous transcription factors.

  12. Structural Studies of the SET Domain from RIZ1 Tumor Suppressor

    Energy Technology Data Exchange (ETDEWEB)

    Briknarova, Klara; Zhou, Xinliang; Satterthwait, Arnold C.; Hoyt, David W.; Ely, Kathryn R.; Huang, Shi

    2008-02-15

    Histone lysine methyltransferases (HKMTs) are involved in regulation of chromatin structure, and, as such, are important for longterm gene activation and repression that is associated with cell memory and establishment of cell-type specific transcriptional programs. Most HKMTs contain a SET domain, which is responsible for their catalytic activity. RIZ1 is a transcription regulator and tumor suppressor that catalyzes methylation of lysine 9 of histone H3 and contains a rather distinct SET domain. Similar SET domains, sometimes refererred to as PR (PRDI-BF1 and RIZ1 homology) domains, are also found in other proteins including Blimp-1/PRDI-BF1, MDS1-EVI1 and Meisetz. We determined the solution structure of the PR domain from RIZ1 and characterized its interaction with S-adenosyl homocysteine (SAH) and a peptide from histone H3. Despite low sequence identity with canonical SET domains, the PR domain displays a typical SET fold including a pseudo-knot at the C-terminus. The N-flanking sequence of RIZ1 PR domain adopts a novel conformation and interacts closely with the SET fold. The C-flanking sequence contains an α-helix that exhibits higher mobility than the SET fold and points away from the protein face that harbors active site in other SET domains. Residues that interact with the methylation cofactor in SET domains are not conserved in RIZ1 or other PR domains, and the SET fold of RIZ1 does not bind SAH. However, the PR domain of RIZ1 interacts specifically with a synthetic peptide comprising residues 1-20 of histone H3.

  13. Mechanism of inhibition of the tumor suppressor Patched by Sonic Hedgehog.

    Science.gov (United States)

    Tukachinsky, Hanna; Petrov, Kostadin; Watanabe, Miyako; Salic, Adrian

    2016-10-04

    The Hedgehog cell-cell signaling pathway is crucial for animal development, and its misregulation is implicated in numerous birth defects and cancers. In unstimulated cells, pathway activity is inhibited by the tumor suppressor membrane protein, Patched. Hedgehog signaling is triggered by the secreted Hedgehog ligand, which binds and inhibits Patched, thus setting in motion the downstream events in signal transduction. Despite its critical importance, the mechanism by which Hedgehog antagonizes Patched has remained unknown. Here, we show that vertebrate Patched1 inhibition is caused by direct, palmitate-dependent interaction with the Sonic Hedgehog ligand. We find that a short palmitoylated N-terminal fragment of Sonic Hedgehog binds Patched1 and, strikingly, is sufficient to inhibit it and to activate signaling. The rest of Sonic Hedgehog confers high-affinity Patched1 binding and internalization through a distinct binding site, but, surprisingly, it is not absolutely required for signaling. The palmitate-dependent interaction with Patched1 is specifically impaired in a Sonic Hedgehog mutant causing human holoprosencephaly, the most frequent congenital brain malformation, explaining its drastically reduced potency. The palmitate-dependent interaction is also abolished in constitutively inhibited Patched1 point mutants causing the Gorlin cancer syndrome, suggesting that they might adopt a conformation distinct from the wild type. Our data demonstrate that Sonic Hedgehog signals via the palmitate-dependent arm of a two-pronged contact with Patched1. Furthermore, our results suggest that, during Hedgehog signaling, ligand binding inhibits Patched by trapping it in an inactive conformation, a mechanism that explains the dramatically reduced activity of oncogenic Patched1 mutants.

  14. Repression of estrogen receptor β function by putative tumor suppressor DBC1

    International Nuclear Information System (INIS)

    Koyama, Satoshi; Wada-Hiraike, Osamu; Nakagawa, Shunsuke; Tanikawa, Michihiro; Hiraike, Haruko; Miyamoto, Yuichiro; Sone, Kenbun; Oda, Katsutoshi; Fukuhara, Hiroshi; Nakagawa, Keiichi; Kato, Shigeaki; Yano, Tetsu; Taketani, Yuji

    2010-01-01

    It has been well established that estrogen is involved in the pathophysiology of breast cancer. Estrogen receptor (ER) α appears to promote the proliferation of cancer tissues, while ERβ can protect against the mitogenic effect of estrogen in breast tissue. The expression status of ERα and ERβ may greatly influence on the development, treatment, and prognosis of breast cancer. Previous studies have indicated that the deleted in breast cancer 1 (DBC1/KIAA1967) gene product has roles in regulating functions of nuclear receptors. The gene encoding DBC1 is a candidate for tumor suppressor identified by genetic search for breast cancer. Caspase-dependent processing of DBC1 promotes apoptosis, and depletion of the endogenous DBC1 negatively regulates p53-dependent apoptosis through its specific inhibition of SIRT1. In addition, DBC1 modulates ERα expression and promotes breast cancer cell survival by binding to ERα. Here we report an ERβ-specific repressive function of DBC1. Immunoprecipitation and immunofluorescence studies show that ERβ and DBC1 interact in a ligand-independent manner similar to ERα. In vitro pull-down assays revealed a direct interaction between DBC1 amino-terminus and activation function-1/2 domain of ERβ. Although DBC1 shows no influence on the ligand-dependent transcriptional activation function of ERα, the expression of DBC1 negatively regulates the ligand-dependent transcriptional activation function of ERβin vivo, and RNA interference-mediated depletion of DBC1 stimulates the transactivation function of ERβ. These results implicate the principal role of DBC1 in regulating ERβ-dependent gene expressions.

  15. Myeloid-derived suppressor cells as a potential therapy for experimental autoimmune myasthenia gravis.

    Science.gov (United States)

    Li, Yan; Tu, Zhidan; Qian, Shiguang; Fung, John J; Markowitz, Sanford D; Kusner, Linda L; Kaminski, Henry J; Lu, Lina; Lin, Feng

    2014-09-01

    We recently demonstrated that hepatic stellate cells induce the differentiation of myeloid-derived suppressor cells (MDSCs) from myeloid progenitors. In this study, we found that adoptive transfer of these MDSCs effectively reversed disease progression in experimental autoimmune myasthenia gravis (EAMG), a T cell-dependent and B cell-mediated model for myasthenia gravis. In addition to ameliorated disease severity, MDSC-treated EAMG mice showed suppressed acetylcholine receptor (AChR)-specific T cell responses, decreased levels of serum anti-AChR IgGs, and reduced complement activation at the neuromuscular junctions. Incubating MDSCs with B cells activated by anti-IgM or anti-CD40 Abs inhibited the proliferation of these in vitro-activated B cells. Administering MDSCs into mice immunized with a T cell-independent Ag inhibited the Ag-specific Ab production in vivo. MDSCs directly inhibit B cells through multiple mechanisms, including PGE2, inducible NO synthase, and arginase. Interestingly, MDSC treatment in EAMG mice does not appear to significantly inhibit their immune response to a nonrelevant Ag, OVA. These results demonstrated that hepatic stellate cell-induced MDSCs concurrently suppress both T and B cell autoimmunity, leading to effective treatment of established EAMG, and that the MDSCs inhibit AChR-specific immune responses at least partially in an Ag-specific manner. These data suggest that MDSCs could be further developed as a novel approach to treating myasthenia gravis and, even more broadly, other diseases in which T and B cells are involved in pathogenesis. Copyright © 2014 by The American Association of Immunologists, Inc.

  16. The potential for tumor suppressor gene therapy in head and neck cancer.

    Science.gov (United States)

    Birkeland, Andrew C; Ludwig, Megan L; Spector, Matthew E; Brenner, J Chad

    2016-01-01

    Head and neck squamous cell carcinoma remains a highly morbid and fatal disease. Importantly, genomic sequencing of head and neck cancers has identified frequent mutations in tumor suppressor genes. While targeted therapeutics increasingly are being investigated in head and neck cancer, the majority of these agents are against overactive/overexpressed oncogenes. Therapy to restore lost tumor suppressor gene function remains a key and under-addressed niche in trials for head and neck cancer. Recent advances in gene editing have captured the interest of both the scientific community and the public. As our technology for gene editing and gene expression modulation improves, addressing lost tumor suppressor gene function in head and neck cancers is becoming a reality. This review will summarize new techniques, challenges to implementation, future directions, and ethical ramifications of gene therapy in head and neck cancer.

  17. Suppressor of cytokine signaling 4 (SOCS4 protects against severe cytokine storm and enhances viral clearance during influenza infection.

    Directory of Open Access Journals (Sweden)

    Lukasz Kedzierski

    2014-05-01

    Full Text Available Suppressor of cytokine signaling (SOCS proteins are key regulators of innate and adaptive immunity. There is no described biological role for SOCS4, despite broad expression in the hematopoietic system. We demonstrate that mice lacking functional SOCS4 protein rapidly succumb to infection with a pathogenic H1N1 influenza virus (PR8 and are hypersusceptible to infection with the less virulent H3N2 (X31 strain. In SOCS4-deficient animals, this led to substantially greater weight loss, dysregulated pro-inflammatory cytokine and chemokine production in the lungs and delayed viral clearance. This was associated with impaired trafficking of influenza-specific CD8 T cells to the site of infection and linked to defects in T cell receptor activation. These results demonstrate that SOCS4 is a critical regulator of anti-viral immunity.

  18. Microcell-mediated chromosome transfer identifies EPB41L3 as a functional suppressor of epithelial ovarian cancers

    DEFF Research Database (Denmark)

    Dafou, Dimitra; Grun, Barbara; Sinclair, John

    2010-01-01

    lines. Using immunohistochemistry, 66% of 794 invasive ovarian tumors showed no EPB41L3 expression compared with only 24% of benign ovarian tumors and 0% of normal ovarian epithelial tissues. EPB41L3 was extensively methylated in ovarian cancer cell lines and primary ovarian tumors compared with normal...... (erythrocyte membrane protein band 4.1-like 3, alternative names DAL-1 and 4.1B) was a candidate ovarian cancer-suppressor gene. Immunoblot analysis showed that EPB41L3 was activated in TOV21G(+18) hybrids, expressed in normal ovarian epithelial cell lines, but was absent in 15 (78%) of 19 ovarian cancer cell...... tissues (P = .00004), suggesting this may be the mechanism of gene inactivation in ovarian cancers. Constitutive reexpression of EPB41L3 in a three-dimensional multicellular spheroid model of ovarian cancer caused significant growth suppression and induced apoptosis. Transmission and scanning electron...

  19. Silibinin inhibits accumulation of myeloid-derived suppressor cells and tumor growth of murine breast cancer

    International Nuclear Information System (INIS)

    Forghani, Parvin; Khorramizadeh, Mohammad R; Waller, Edmund K

    2014-01-01

    Myeloid-derived suppressor cells (MDSC)s increase in blood and accumulate in the tumor microenvironment of tumor-bearing animals, contributing to immune suppression in cancer. Silibinin, a natural flavonoid from the seeds of milk thistle, has been developed as an anti-inflammatory agent and supportive care agent to reduce the toxicity of cancer chemotherapy. The goals of this study were to evaluate the effect of silibinin on MDSCs in tumor-bearing mice and antitumor activity of silibinin in a mouse model of breast cancer. 4T1 luciferase-transfected mammary carcinoma cells were injected into in the mammary fat pad female BALB/c mice, and female CB17-Prkdc Scid/J mice. Silibinin treatment started on day 4 or day 14 after tumor inoculation continued every other day. Tumor growth was monitored by bioluminescent imaging (BLI) measuring total photon flux. Flow cytometry measured total leukocytes, CD11b + Gr-1 + MDSC, and T cells in the blood and tumors of tumor-bearing mice. The effects of silibinin on 4T1 cell viability in vitro were measured by BLI. Treatment with silibinin increased overall survival in mice harboring tumors derived from the 4T1-luciferase breast cancer cell line, and reduced tumor volumes and numbers of CD11b + Gr-1 + MDSCs in the blood and tumor, and increased the content of T cells in the tumor microenvironment. Silibinin failed to inhibit tumor growth in immunocompromised severe combined immunodeficiency mice, supporting the hypothesis that anticancer effect of silibinin is immune-mediated. The antitumor activity of silibinin requires an intact host immune system and is associated with decreased accumulation of blood and tumor-associated MDSCs

  20. Nuclear γ-tubulin associates with nucleoli and interacts with tumor suppressor protein C53.

    Science.gov (United States)

    Hořejší, Barbora; Vinopal, Stanislav; Sládková, Vladimíra; Dráberová, Eduarda; Sulimenko, Vadym; Sulimenko, Tetyana; Vosecká, Věra; Philimonenko, Anatoly; Hozák, Pavel; Katsetos, Christos D; Dráber, Pavel

    2012-01-01

    γ-Tubulin is assumed to be a typical cytosolic protein necessary for nucleation of microtubules from microtubule organizing centers. Using immunolocalization and cell fractionation techniques in combination with siRNAi and expression of FLAG-tagged constructs, we have obtained evidence that γ-tubulin is also present in nucleoli of mammalian interphase cells of diverse cellular origins. Immunoelectron microscopy has revealed γ-tubulin localization outside fibrillar centers where transcription of ribosomal DNA takes place. γ-Tubulin was associated with nucleolar remnants after nuclear envelope breakdown and could be translocated to nucleoli during mitosis. Pretreatment of cells with leptomycin B did not affect the distribution of nuclear γ-tubulin, making it unlikely that rapid active transport via nuclear pores participates in the transport of γ-tubulin into the nucleus. This finding was confirmed by heterokaryon assay and time-lapse imaging of photoconvertible protein Dendra2 tagged to γ-tubulin. Immunoprecipitation from nuclear extracts combined with mass spectrometry revealed an association of γ-tubulin with tumor suppressor protein C53 located at multiple subcellular compartments including nucleoli. The notion of an interaction between γ-tubulin and C53 was corroborated by pull-down and co-immunoprecipitation experiments. Overexpression of γ-tubulin antagonized the inhibitory effect of C53 on DNA damage G(2) /M checkpoint activation. The combined results indicate that aside from its known role in microtubule nucleation, γ-tubulin may also have nuclear-specific function(s). Copyright © 2011 Wiley Periodicals, Inc.

  1. Tumor suppressor roles of CENP-E and Nsl1 in Drosophila epithelial tissues.

    Science.gov (United States)

    Clemente-Ruiz, Marta; Muzzopappa, Mariana; Milán, Marco

    2014-01-01

    Depletion of spindle assembly checkpoint (SAC) genes in Drosophila epithelial tissues leads to JNK-dependent programmed cell death and additional blockade of the apoptotic program drives tumorigenesis. A recent report proposes that chromosomal instability (CIN) is not the driving force in the tumorigenic response of the SAC-deficient tissue, and that checkpoint proteins exert a SAC-independent tumor suppressor role. This notion is based on observations that the depletion of CENP-E levels or prevention of Bub3 from binding to the kinetochore in Drosophila tissues unable to activate the apoptotic program induces CIN but does not cause hyperproliferation. Here we re-examined this proposal. In contrast to the previous report, we observed that depletion of CENP-E or Nsl1-the latter mediating kinetochore targeting of Bub3-in epithelial tissues unable to activate the apoptotic program induces significant levels of aneuploidy and drives tumor-like growth. The induction of the JNK transcriptional targets Wingless, a mitogenic molecule, and MMP1, a matrix metaloproteinase 1 involved in basement membrane degradation was also observed in these tumors. An identical response of the tissue was previously detected upon depletion of several SAC genes or genes involved in spindle assembly, chromatin condensation, and cytokinesis, all of which have been described to cause CIN. All together, these results reinforce the role of CIN in driving tumorigenesis in Drosophila epithelial tissues and question the proposed SAC-independent roles of checkpoint proteins in suppressing tumorigenesis. Differences in aneuploidy rates might explain the discrepancy between the previous report and our results.

  2. Gamma-aminobutyric acid, a potential tumor suppressor for small airway-derived lung adenocarcinoma.

    Science.gov (United States)

    Schuller, Hildegard M; Al-Wadei, Hussein A N; Majidi, Mourad

    2008-10-01

    Pulmonary adenocarcinoma (PAC) is the leading type of lung cancer in smokers and non-smokers that arises in most cases from small airway epithelial cells. PAC has a high mortality due to its aggressive behavior and resistance to cancer therapeutics. We have shown previously that the proliferation of human PAC cells NCI-H322 and immortalized human small airway epithelial cells HPL1D is stimulated by cyclic adenosine monophosphate (cAMP)/protein kinase A-dependent phosphorylation of cyclic adenosine monophosphate response element-binding (CREB) protein and transactivation of the epidermal growth factor receptor and that this pathway is activated by beta-1-adrenoreceptors (beta(1)-ARs) and the non-genomic estrogen receptor beta. Our current in vitro studies with HPL1D and NCI-H322 cells showed that signaling via the gamma-amino butyric acid receptor (GABA(B)R) strongly inhibited base level and isoproterenol-induced cAMP, p-CREB, cyclic adenosine monophosphate response element-luciferase activity and p-extracellular regulated kinase-1 (ERK1)/2 and effectively blocked DNA synthesis and cell migration. The inhibitory effects of gamma-amino butyric acid (GABA) were disinhibited by the GABA(B)R antagonist CGP-35348 or GABA(B)R knockdown. Immunohistochemical investigation of hamster lungs showed significant underexpression of GABA in animals with small airway-derived PACs induced by the nicotine-derived carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK). These findings suggest that GABA may have tumor suppressor function in small airway epithelia and the PACs derived from them and that downregulation of GABA by NNK may contribute to the development of this cancer in smokers. Our findings suggest that marker-guided treatment with GABA or a GABA(B)R agonist of individuals with downregulated pulmonary GABA may provide a novel targeted approach for the prevention of PAC in smokers.

  3. 15-Hydroxyprostaglandin dehydrogenase is an in vivo suppressor of colon tumorigenesis.

    Science.gov (United States)

    Myung, Seung-Jae; Rerko, Ronald M; Yan, Min; Platzer, Petra; Guda, Kishore; Dotson, Angela; Lawrence, Earl; Dannenberg, Andrew J; Lovgren, Alysia Kern; Luo, Guangbin; Pretlow, Theresa P; Newman, Robert A; Willis, Joseph; Dawson, Dawn; Markowitz, Sanford D

    2006-08-08

    15-Hydroxyprostaglandin dehydrogenase (15-PGDH) is a prostaglandin-degrading enzyme that is highly expressed in normal colon mucosa but is ubiquitously lost in human colon cancers. Herein, we demonstrate that 15-PGDH is active in vivo as a highly potent suppressor of colon neoplasia development and acts in the colon as a required physiologic antagonist of the prostaglandin-synthesizing activity of the cyclooxygenase 2 (COX-2) oncogene. We first show that 15-PGDH gene knockout induces a marked 7.6-fold increase in colon tumors arising in the Min (multiple intestinal neoplasia) mouse model. Furthermore, 15-PGDH gene knockout abrogates the normal resistance of C57BL/6J mice to colon tumor induction by the carcinogen azoxymethane (AOM), conferring susceptibility to AOM-induced adenomas and carcinomas in situ. Susceptibility to AOM-induced tumorigenesis is mediated by a marked induction of dysplasia, proliferation, and cyclin D1 expression throughout microscopic aberrant crypt foci arising in 15-PGDH null colons and is concomitant with a doubling of prostaglandin E(2) in 15-PGDH null colonic mucosa. A parallel role for 15-PGDH loss in promoting the earliest steps of colon neoplasia in humans is supported by our finding of a universal loss of 15-PGDH expression in microscopic colon adenomas recovered from patients with familial adenomatous polyposis, including adenomas as small as a single crypt. These models thus delineate the in vivo significance of 15-PGDH-mediated negative regulation of the COX-2 pathway and moreover reveal the particular importance of 15-PGDH in opposing the neoplastic progression of colonic aberrant crypt foci.

  4. Reduction of myeloid-derived suppressor cells and lymphoma growth by a natural triterpenoid.

    Science.gov (United States)

    Radwan, Faisal F Y; Hossain, Azim; God, Jason M; Leaphart, Nathan; Elvington, Michelle; Nagarkatti, Mitzi; Tomlinson, Stephen; Haque, Azizul

    2015-01-01

    Lymphoma is a potentially life threatening disease. The goal of this study was to investigate the therapeutic potential of a natural triterpenoid, Ganoderic acid A (GA-A) in controlling lymphoma growth both in vitro and in vivo. Here, we show that GA-A treatment induces caspase-dependent apoptotic cell death characterized by a dose-dependent increase in active caspases 9 and 3, up-regulation of pro-apoptotic BIM and BAX proteins, and a subsequent loss of mitochondrial membrane potential with release of cytochrome c. In addition to GA-A's anti-growth activity, we show that lower doses of GA-A enhance HLA class II-mediated antigen (Ag) presentation and CD4+ T cell recognition of lymphoma cells in vitro. The therapeutic relevance of GA-A treatment was also tested in vivo using the EL4 syngeneic mouse model of metastatic lymphoma. GA-A-treatment significantly prolonged survival of EL4 challenged mice and decreased tumor metastasis to the liver, an outcome accompanied by a marked down-regulation of STAT3 phosphorylation, reduction myeloid-derived suppressor cells (MDSCs), and enhancement of cytotoxic CD8+ T cells in the host. Thus, GA-A not only selectively induces apoptosis in lymphoma cells, but also enhances cell-mediated immune responses by attenuating MDSCs, and elevating Ag presentation and T cell recognition. The demonstrated therapeutic benefit indicates that GA-A is a candidate for future drug design for the treatment of lymphoma. © 2014 Wiley Periodicals, Inc.

  5. Clinical impact of the immunome in lymphoid malignancies: the role of Myeloid-Derived Suppressor Cells

    Directory of Open Access Journals (Sweden)

    Calogero eVetro

    2015-05-01

    Full Text Available The better definition of the mutual sustainment between neoplastic cells and immune system has been translated from the bench to the bedside acquiring value as prognostic factor. Additionally, it represents a promising tool for improving therapeutic strategies. In this context, myeloid-derived suppressor cells have gained a central role in tumor developing with consequent therapeutic implications. In this review, we will focus on the biological and clinical impact of the study of myeloid-derived suppressor cells in the settings of lymphoid malignancies.

  6. The role of tumor suppressor p15Ink4b in the regulation of hematopoietic progenitor cell fate

    International Nuclear Information System (INIS)

    Humeniuk, R; Rosu-Myles, M; Fares, J; Koller, R; Bies, J; Wolff, L

    2013-01-01

    Epigenetic silencing of the tumor suppressor gene p15Ink4b (CDKN2B) is a frequent event in blood disorders like acute myeloid leukemia and myelodysplastic syndromes. The molecular function of p15Ink4b in hematopoietic differentiation still remains to be elucidated. Our previous study demonstrated that loss of p15Ink4b in mice results in skewing of the differentiation pattern of the common myeloid progenitor towards the myeloid lineage. Here, we investigated a function of p15Ink4b tumor suppressor gene in driving erythroid lineage commitment in hematopoietic progenitors. It was found that p15Ink4b is expressed more highly in committed megakaryocyte–erythroid progenitors than granulocyte–macrophage progenitors. More importantly, mice lacking p15Ink4b have lower numbers of primitive red cell progenitors and a severely impaired response to 5-fluorouracil- and phenylhydrazine-induced hematopoietic stress. Introduction of p15Ink4b into multipotential progenitors produced changes at the molecular level, including activation of mitogen-activated protein kinase/extracellular signal-regulated kinase (MEK/ERK) signaling, increase GATA-1, erythropoietin receptor (EpoR) and decrease Pu1, GATA-2 expression. These changes rendered cells more permissive to erythroid commitment and less permissive to myeloid commitment, as demonstrated by an increase in early burst-forming unit-erythroid formation with concomitant decrease in myeloid colonies. Our results indicate that p15Ink4b functions in hematopoiesis, by maintaining proper lineage commitment of progenitors and assisting in rapid red blood cells replenishment following stress

  7. Loss of Mitochondrial Tumor Suppressor Genes Expression Is Associated with Unfavorable Clinical Outcome in Head and Neck Squamous Cell Carcinoma: Data from Retrospective Study.

    Directory of Open Access Journals (Sweden)

    Ishrat Mahjabeen

    Full Text Available Mitochondrial genes play important roles in cellular energy metabolism, free radical generation, and apoptosis. Dysregulation of these genes have long been suspected to contribute to the generation of reactive oxygen species (ROS, increased proliferation and progression of cancer. A family of orthologues of yeast silent information regulator 3 (SIRT3, 4 (SIRT4 and mitochondrial tumor suppressor 1 (MTUS1 are important mitochondrial tumor suppressor genes which play an important role in the progression of multiple cancers. However, their role in the development of oxidative stress, enhanced proliferation and progression of head and neck squamous cell carcinoma (HNSCC has not yet been studied. In this study we aimed to test the association between reduced mitochondrial tumor suppressor genes' activities and enhancement in tissue oxidative stress and cell proliferation in HNSCC cases. The expression of mitochondrial tumor suppressor genes (SIRT3, SIRT4 and MTUS1, mitochondrial DNA repair gene (OGG1-2a and a proliferation marker (Ki-67 was studied in a study cohort of 120 HNSCC patients and controls with reverse transcriptase polymerase chain reaction (RT-PCR and real-time PCR (qPCR in order to determine the potential prognostic significance of these genes. A statistically significant downregulation of SIRT3 (p<0.001, SIRT4 (p<0.0001, MTUS1 (p<0.002 and OGG1 (p<0.0001 was observed in HNSCC compared to control samples. Ki-67 was also overexpressed (p<0.0001 in HNSCC versus control samples. Additionally, to explore gene-gene relationship, we observed a positive spearmen correlation between SIRT3 versus SIRT4 (r = 0.523***, p<0.0001, SIRT3 versus MTUS1 (r = 0.273***, p<0.001, SIRT3 versus OGG1-2a (r = 0.213*, p<0.03, SIRT4 versus OGG1 (r = 0.338***, p<0.0001 and MTUS1 versus OGG1-2a (r = 0.215*, p<0.03 in HNSCC cases. A negative spearman correlation was observed between OGG1 versus Ki-67 (r = -0.224**, p<0.01 and OGG1-2a versus Ki-67 (r = -0.224**, p<0

  8. DMPD: Suppressor of cytokine signaling (SOCS) 2, a protein with multiple functions. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 17070092 Suppressor of cytokine signaling (SOCS) 2, a protein with multiple function...Epub 2006 Oct 27. (.png) (.svg) (.html) (.csml) Show Suppressor of cytokine signaling (SOCS) 2, a protein with multiple function...SOCS) 2, a protein with multiple functions. Authors Rico-Bautista E, Flores-Morales A, Fernandez-Perez L. Pu

  9. Cancer cell specific cytotoxic gene expression mediated by ARF tumor suppressor promoter constructs

    International Nuclear Information System (INIS)

    Kurayoshi, Kenta; Ozono, Eiko; Iwanaga, Ritsuko; Bradford, Andrew P.; Komori, Hideyuki; Ohtani, Kiyoshi

    2014-01-01

    Highlights: • ARF promoter showed higher responsiveness to deregulated E2F activity than the E2F1 promoter. • ARF promoter showed higher cancer cell-specificity than E2F1 promoter to drive gene expression. • HSV-TK driven by ARF promoter showed higher cancer cell-specific cytotoxicity than that driven by E2F1 promoter. - Abstract: In current cancer treatment protocols, such as radiation and chemotherapy, side effects on normal cells are major obstacles to radical therapy. To avoid these side effects, a cancer cell-specific approach is needed. One way to specifically target cancer cells is to utilize a cancer specific promoter to express a cytotoxic gene (suicide gene therapy) or a viral gene required for viral replication (oncolytic virotherapy). For this purpose, the selected promoter should have minimal activity in normal cells to avoid side effects, and high activity in a wide variety of cancers to obtain optimal therapeutic efficacy. In contrast to the AFP, CEA and PSA promoters, which have high activity only in a limited spectrum of tumors, the E2F1 promoter exhibits high activity in wide variety of cancers. This is based on the mechanism of carcinogenesis. Defects in the RB pathway and activation of the transcription factor E2F, the main target of the RB pathway, are observed in almost all cancers. Consequently, the E2F1 promoter, which is mainly regulated by E2F, has high activity in wide variety of cancers. However, E2F is also activated by growth stimulation in normal growing cells, suggesting that the E2F1 promoter may also be highly active in normal growing cells. In contrast, we found that the tumor suppressor ARF promoter is activated by deregulated E2F activity, induced by forced inactivation of pRB, but does not respond to physiological E2F activity induced by growth stimulation. We also found that the deregulated E2F activity, which activates the ARF promoter, is detected only in cancer cell lines. These observations suggest that ARF promoter

  10. Cancer cell specific cytotoxic gene expression mediated by ARF tumor suppressor promoter constructs

    Energy Technology Data Exchange (ETDEWEB)

    Kurayoshi, Kenta [Department of Bioscience, School of Science and Technology, Kwansei Gakuin University, 2-1 Gakuen, Sanda, Hyogo 669-1337 (Japan); Ozono, Eiko [Centre for Molecular Oncology, Barts Cancer Institute, Queen Mary, University of London, John Vane Science Centre, Charterhouse Square, London EC1M 6BQ (United Kingdom); Iwanaga, Ritsuko; Bradford, Andrew P. [Department of Obstetrics and Gynecology, University of Colorado School of Medicine, Anschutz Medical Campus, 12800 East 19th Avenue, Aurora, CO 80045 (United States); Komori, Hideyuki [Center for Stem Cell Biology, Life Sciences Institute, University of Michigan, Ann Arbor, MI 48109 (United States); Ohtani, Kiyoshi, E-mail: btm88939@kwansei.ac.jp [Department of Bioscience, School of Science and Technology, Kwansei Gakuin University, 2-1 Gakuen, Sanda, Hyogo 669-1337 (Japan)

    2014-07-18

    Highlights: • ARF promoter showed higher responsiveness to deregulated E2F activity than the E2F1 promoter. • ARF promoter showed higher cancer cell-specificity than E2F1 promoter to drive gene expression. • HSV-TK driven by ARF promoter showed higher cancer cell-specific cytotoxicity than that driven by E2F1 promoter. - Abstract: In current cancer treatment protocols, such as radiation and chemotherapy, side effects on normal cells are major obstacles to radical therapy. To avoid these side effects, a cancer cell-specific approach is needed. One way to specifically target cancer cells is to utilize a cancer specific promoter to express a cytotoxic gene (suicide gene therapy) or a viral gene required for viral replication (oncolytic virotherapy). For this purpose, the selected promoter should have minimal activity in normal cells to avoid side effects, and high activity in a wide variety of cancers to obtain optimal therapeutic efficacy. In contrast to the AFP, CEA and PSA promoters, which have high activity only in a limited spectrum of tumors, the E2F1 promoter exhibits high activity in wide variety of cancers. This is based on the mechanism of carcinogenesis. Defects in the RB pathway and activation of the transcription factor E2F, the main target of the RB pathway, are observed in almost all cancers. Consequently, the E2F1 promoter, which is mainly regulated by E2F, has high activity in wide variety of cancers. However, E2F is also activated by growth stimulation in normal growing cells, suggesting that the E2F1 promoter may also be highly active in normal growing cells. In contrast, we found that the tumor suppressor ARF promoter is activated by deregulated E2F activity, induced by forced inactivation of pRB, but does not respond to physiological E2F activity induced by growth stimulation. We also found that the deregulated E2F activity, which activates the ARF promoter, is detected only in cancer cell lines. These observations suggest that ARF promoter

  11. Mimetics of Suppressor of cytokine signalling 3: novel potential therapeutics in triple breast cancer.

    Science.gov (United States)

    La Manna, Sara; Lee, Eunmi; Ouzounova, Maria; Di Natale, Concetta; Novellino, Ettore; Merlino, Antonello; Korkaya, Hasan; Marasco, Daniela

    2018-05-11

    Suppressor of cytokine signaling (SOCS) family of proteins plays critical role in the regulation of immune responses controlling JAK/STAT mediated inflammatory cytokines. Among the members, SOCS1 and SOCS3 contain a kinase inhibitory region (KIR) and SOCS3 binds to JAK/STAT/gp130 complex by inhibiting the downstream signaling and suppressing inflammatory cytokines. Loss or reduced levels of SOCS3 have been linked to cancer-associated inflammation and suppressive immunity leading to enhanced tumour growth and metastasis. In line with these reports, we previously demonstrated that proteolytic degradation of SOCS3 in triple negative breast cancer (TNBC) subtype drives the expression of inflammatory cytokines. Therefore, we postulated that SOCS3 mimetics might suppress the inflammatory cytokine production in TNBC subtype and inhibit tumor growth and metastasis. Here we designed and characterized five linear peptides derived from the N-terminal region of SOCS3 encompassing regions that interface with the JAK2/gp130 complex by using the Circular Dichroism and Surface Plasmon Resonance spectroscopies. The KIRESS peptide resulted the sequence containing the most part of the hot-spots required for binding to JAK2 and was further investigated in vivo in mouse xenografts of MDA-MB-231-luci tumours as models of human TNBC subtype. Expectedly, this peptide showed a significant inhibition of primary tumour growth and pulmonary metastasis. Our studies suggest that SOCS3 peptidomimetics may possess a therapeutic potential in aggressive cancers, such as TNBC subtype, with activated inflammatory cytokines. This article is protected by copyright. All rights reserved. © 2018 UICC.

  12. miR-126 Functions as a Tumor Suppressor in Osteosarcoma by Targeting Sox2

    Directory of Open Access Journals (Sweden)

    Chenglin Yang

    2013-12-01

    Full Text Available Osteosarcoma (OS is the most common malignant bone tumor in children and young adults, the early symptoms and signs of which are non-specific. The discovery of microRNAs (miRNAs provides a new avenue for the early diagnosis and treatment of OS. miR-126 has been reported to be highly expressed in vascularized tissues, and is recently widely studied in cancers. Herein, we explored the expression and significance of miR-126 in OS. Using TaqMan RT-PCR analysis, we analyzed the expression of miR-126 in 32 paired OS tumor tissues and 4 OS cell lines and found that miR-126 was consistently under-expressed in OS tissues and cell lines compared with normal bone tissues and normal osteoblast cells (NHOst, respectively. As miR-126 is significantly decreased in OS tissues and cell lines, we sought to compensate for its loss through exogenous transfection into MG-63 cells with a miR-126 mimic. Ectopic expression of miR-126 inhibited cell proliferation, migration and invasion, and induced apoptosis of MG-63 cells. Moreover, bioinformatic prediction suggested that the sex-determining region Y-box 2 (Sox2 is a target gene of miR-126. Using mRNA and protein expression analysis, luciferase assays and rescue assays, we demonstrate that restored expression of Sox2 dampened miR-126-mediated suppression of tumor progression, which suggests the important role of miR-126/Sox2 interaction in tumor progression. Taken together, our data indicate that miR-126 functions as a tumor suppressor in OS, which exerts its activity by suppressing the expression of Sox2.

  13. BAX and tumor suppressor TRP53 are important in regulating mutagenesis in spermatogenic cells in mice.

    Science.gov (United States)

    Xu, Guogang; Vogel, Kristine S; McMahan, C Alex; Herbert, Damon C; Walter, Christi A

    2010-12-01

    During the first wave of spermatogenesis, and in response to ionizing radiation, elevated mutant frequencies are reduced to a low level by unidentified mechanisms. Apoptosis is occurring in the same time frame that the mutant frequency declines. We examined the role of apoptosis in regulating mutant frequency during spermatogenesis. Apoptosis and mutant frequencies were determined in spermatogenic cells obtained from Bax-null or Trp53-null mice. The results showed that spermatogenic lineage apoptosis was markedly decreased in Bax-null mice and was accompanied by a significantly increased spontaneous mutant frequency in seminiferous tubule cells compared to that of wild-type mice. Apoptosis profiles in the seminiferous tubules for Trp53-null were similar to control mice. Spontaneous mutant frequencies in pachytene spermatocytes and in round spermatids from Trp53-null mice were not significantly different from those of wild-type mice. However, epididymal spermatozoa from Trp53-null mice displayed a greater spontaneous mutant frequency compared to that from wild-type mice. A greater proportion of spontaneous transversions and a greater proportion of insertions/deletions 15 days after ionizing radiation were observed in Trp53-null mice compared to wild-type mice. Base excision repair activity in mixed germ cell nuclear extracts prepared from Trp53-null mice was significantly lower than that for wild-type controls. These data indicate that BAX-mediated apoptosis plays a significant role in regulating spontaneous mutagenesis in seminiferous tubule cells obtained from neonatal mice, whereas tumor suppressor TRP53 plays a significant role in regulating spontaneous mutagenesis between postmeiotic round spermatid and epididymal spermatozoon stages of spermiogenesis.

  14. Identification of essential sequences for cellular localization in BRMS1 metastasis suppressor.

    Directory of Open Access Journals (Sweden)

    José Rivera

    Full Text Available BACKGROUND: Breast cancer metastasis suppressor 1 (BRMS1 reduces the number and the size of secondary tumours in a mouse model without affecting the growth of the primary foci upon its re-expression. Knockdown of BRMS1 expression associates with metastasis. The molecular details on BRMS1 mechanism of action include its ability to function as a transcriptional co-repressor and consistently BRMS1 has been described as a predominantly nuclear protein. Since cellular distribution could represent a potential mechanism of regulation, we wanted to characterize BRMS1 sequence motifs that might regulate its cellular distribution. According to its amino acids sequence, BRMS1 contain two putative nuclear localization signals, however none of them has been proved to work so far. METHODOLOGY/PRINCIPAL FINDINGS: By using well known in vivo assays to detect both nuclear import and export signal, we have characterized, in the present study, one functional nuclear localisation signal as necessary and sufficient to promote nuclear transport. Additionally, the outcome of a directed yeast two-hybrid assay identify importin alpha6 as a specific partner of BRMS1 thus speculating that BRMS1 nuclear import could be specifically mediated by the reported nuclear transporter. Besides, the combination of a computational searching approach along the utilization of a nuclear export assay, identified a functional motif within the BRMS1 sequence responsible for its nuclear export, that resulted not affected by the highly specific CRM1 inhibitor Leptomycin-B. Interspecies heterokaryon assay demonstrate the capability of BRMS1 to shuttle between the nuclear and cytosolic compartments CONCLUSIONS/SIGNIFICANCE: Our results show for the first time that BRMS1 contains both nuclear import and export signals enabling its nucleo-cytoplasmic shuttling. These findings contributes new data for the understanding of the BRMS1 functions and allow us to speculate that this phenomenon could

  15. Absence of mutations in the coding sequence of the potential tumor suppressor 3pK in metastatic melanoma

    Directory of Open Access Journals (Sweden)

    Houben Roland

    2005-12-01

    Full Text Available Abstract Background Activation of Ras or Raf contributes to tumorigenesis of melanoma. However, constitutive Raf activation is also a characteristic of the majority of benign melanocytic nevi and high intensity signaling of either Ras or Raf was found to induce growth inhibition and senescence rather than transformation. Since the chromosome 3p kinase (3pK is a target of the Ras/Raf/Mek/Erk signaling pathway which antagonizes the function of the oncogene and anti-differentiation factor Bmi-1, 3pK may function as a tumor suppressor in tumors with constitutive Ras/Raf activation. Consequently, we tested whether inactivating 3pK mutations are present in melanoma. Methods 30 metastatic melanoma samples, which were positive for activating mutations of either BRaf or NRas, were analyzed for possible mutations in the 3pk gene. The 10 coding exons and their flanking intron sequences were amplified by PCR and direct sequencing of the PCR products was performed. Results This analysis revealed that besides the presence of some single nucleotide polymorphisms in the 3pk gene, we could not detect any possible loss of function mutation in any of these 30 metastatic melanoma samples selected for the presence of activating mutations within the Ras/Raf/Mek/Erk signaling pathway. Conclusion Hence, in melanoma with constitutively active Ras/Raf inactivating mutations within the 3pk gene do not contribute to the oncogenic phenotype of this highly malignant tumor.

  16. A study on tumor suppressor genes mutations associated with different pathological colorectal lesions

    International Nuclear Information System (INIS)

    Matar, S.N.A.

    2011-01-01

    Colorectal cancer (CRC) is the second leading cause of cancer-related death in the Western world. In Egypt; there is an increasing incidence of the disease, especially among patients ≤40 years age. While CRC have been reported in low incidence rate in developing countries, it is the third most common tumor in male and the fifth common tumor in females in Egypt. Early diagnosis and surgical interference guarantee long survival of most CRC patients. Early diagnosis is impeded by the disease onset at young age and imprecise symptoms at the initial stages of the disease. As in most solid tumors, the malignant transformation of colonic epithelial cells is to arise through a multistep process during which they acquire genetic changes involving the activation of proto-oncogenes and the loss of tumor suppressor genes. Recently, a candidate tumor suppressor gene, KLF6, which is mapped to chromosome 10p, was found to be frequently mutated in a number of cancers. There are some evidences suggesting that the disruption of the functional activity of KLF6 gene products may be one of the early events in tumor genesis of the colon. The main objective of the present study was to detect mutational changes of KLF6 tumor suppressor gene and to study the loss of heterozygosity (LOH) markers at chromosome 10p15 (KLF6 locus) in colorectal lesions and colorectal cancer in Egyptian patients. The patients included in this study were 83 presented with different indications for colonoscopic examination. Selecting patients with colorectal pre-cancerous lesions or colorectal cancer was done according to the results of tissue biopsy from lesion and adjacent normal. The patients were classified into three main groups; (G I) Cancerous group, (G II) polyps group including patients with adenomatous polyps (AP), familial adenomatous polyps (FAP) and hyperplastic polyps (HP) and (G III) Inflammatory Bowel Diseases (IBD) including patients with ulcerative colitis (UC) and Crohn's disease (CD

  17. Influence of anticancer drugs on interactions of tumor suppressor protein p53 with DNA

    Czech Academy of Sciences Publication Activity Database

    Pivoňková, Hana; Němcová, Kateřina; Brázdová, Marie; Kašpárková, Jana; Brabec, Viktor; Fojta, Miroslav

    2005-01-01

    Roč. 272, Suppl. 1 (2005), s. 562 ISSN 1474-3833. [FEBS Congress /30./ and IUBMB Conference /9./. 02.07.2005-07.07.2005, Budapest] R&D Projects: GA MZd(CZ) NC7574 Institutional research plan: CEZ:AV0Z50040507 Keywords : tumour suppressor protein p53 * anticancer drugs * interaction with DNA Subject RIV: BO - Biophysics

  18. TFPI-2 is a putative tumor suppressor gene frequently inactivated by promoter hypermethylation in nasopharyngeal carcinoma

    International Nuclear Information System (INIS)

    Wang, Shumin; Ma, Ning; Murata, Mariko; Huang, Guangwu; Zhang, Zhe; Xiao, Xue; Zhou, Xiaoying; Huang, Tingting; Du, Chunping; Yu, Nana; Mo, Yingxi; Lin, Longde; Zhang, Jinyan

    2010-01-01

    Epigenetic silencing of tumor suppressor genes play important roles in NPC tumorgenesis. Tissue factor pathway inhibitor-2 (TFPI-2), is a protease inhibitor. Recently, TFPI-2 was suggested to be a tumor suppressor gene involved in tumorigenesis and metastasis in some cancers. In this study, we investigated whether TFPI-2 was inactivated epigenetically in nasopharyngeal carcinoma (NPC). Transcriptional expression levels of TFPI-2 was evaluated by RT-PCR. Methylation status were investigated by methylation specific PCR and bisulfate genomic sequencing. The role of TFPI-2 as a tumor suppressor gene in NPC was addressed by re-introducing TFPI-2 expression into the NPC cell line CNE2. TFPI-2 mRNA transcription was inactivated in NPC cell lines. TFPI-2 was aberrantly methylated in 66.7% (4/6) NPC cell lines and 88.6% (62/70) of NPC primary tumors, but not in normal nasopharyngeal epithelia. TFPI-2 expression could be restored in NPC cells after demethylation treatment. Ectopic expression of TFPI-2 in NPC cells induced apoptosis and inhibited cell proliferation, colony formation and cell migration. Epigenetic inactivation of TFPI-2 by promoter hypermethylation is a frequent and tumor specific event in NPC. TFPI-2 might be considering as a putative tumor suppressor gene in NPC

  19. Mechanism of inhibition of growth hormone receptor signaling by suppressor of cytokine signaling proteins

    DEFF Research Database (Denmark)

    Hansen, J A; Lindberg, K; Hilton, D J

    1999-01-01

    In this study we have investigated the role of suppressor of cytokine signaling (SOCS) proteins in GH receptor-mediated signaling. GH-induced transcription was inhibited by SOCS-1 and SOCS-3, while SOCS-2 and cytokine inducible SH2-containing protein (CIS) had no effect By using chimeric SOCS pro...

  20. Suppressor of cytokine signaling 3 knockdown in the mediobasal hypothalamus: counterintuitive effects on energy balance

    NARCIS (Netherlands)

    de Backer, M. W. A.; Brans, M. A. D.; van Rozen, A. J.; van der Zwaal, E. M.; Luijendijk, M. C. M.; Garner, K. G.; de Krom, M.; van Beekum, O.; La Fleur, S. E.; Adan, R. A. H.

    2010-01-01

    An increase in brain suppressor of cytokine signaling 3 (SOCS3) has been implicated in the development of both leptin and insulin resistance. Socs3 mRNA is localized throughout the brain, and it remains unclear which brain areas are involved in the effect of SOCS3 levels on energy balance. We

  1. Suppressor Effects in Coping Research with African American Adolescents from Low-Income Communities

    Science.gov (United States)

    Gaylord-Harden, Noni K.; Cunningham, Jamila A.; Holmbeck, Grayson N.; Grant, Kathryn E.

    2010-01-01

    Objective: The purpose of the current study was to demonstrate the replicable nature of statistical suppressor effects in coping research through 2 examples with African American adolescents from low-income communities. Method: Participants in the 1st example included 497 African American adolescents (mean age = 12.61 years, SD = 0.99; 57% female)…

  2. Identification of new adventitious rooting mutants amongst suppressors of the Arabidopsis thaliana superroot2 mutation.

    Science.gov (United States)

    Pacurar, Daniel Ioan; Pacurar, Monica Lacramioara; Bussell, John Desmond; Schwambach, Joseli; Pop, Tiberia Ioana; Kowalczyk, Mariusz; Gutierrez, Laurent; Cavel, Emilie; Chaabouni, Salma; Ljung, Karin; Fett-Neto, Arthur Germano; Pamfil, Doru; Bellini, Catherine

    2014-04-01

    The plant hormone auxin plays a central role in adventitious rooting and is routinely used with many economically important, vegetatively propagated plant species to promote adventitious root initiation and development on cuttings. Nevertheless the molecular mechanisms through which it acts are only starting to emerge. The Arabidopsis superroot2-1 (sur2-1) mutant overproduces auxin and, as a consequence, develops excessive adventitious roots in the hypocotyl. In order to increase the knowledge of adventitious rooting and of auxin signalling pathways and crosstalk, this study performed a screen for suppressors of superroot2-1 phenotype. These suppressors provide a new resource for discovery of genetic players involved in auxin signalling pathways or at the crosstalk of auxin and other hormones or environmental signals. This study reports the identification and characterization of 26 sur2-1 suppressor mutants, several of which were identified as mutations in candidate genes involved in either auxin biosynthesis or signalling. In addition to confirming the role of auxin as a central regulator of adventitious rooting, superroot2 suppressors indicated possible crosstalk with ethylene signalling in this process.

  3. Alterations in tumour suppressor gene p53 in human gliomas from ...

    Indian Academy of Sciences (India)

    Unknown

    Alterations in the tumour suppressor p53 gene are among the most common defects seen in a variety of human cancers. ..... rangement of the EGF receptor gene in primary human brain tumors ... the INK4A gene in superficial bladder tumors.

  4. The Polerovirus silencing suppressor P0 targets ARGONAUTE proteins for degradation.

    Science.gov (United States)

    Baumberger, Nicolas; Tsai, Ching-Hsui; Lie, Miranda; Havecker, Ericka; Baulcombe, David C

    2007-09-18

    Plant and animal viruses encode suppressor proteins of an adaptive immunity mechanism in which viral double-stranded RNA is processed into 21-25 nt short interfering (si)RNAs. The siRNAs guide ARGONAUTE (AGO) proteins so that they target viral RNA. Most viral suppressors bind long dsRNA or siRNAs and thereby prevent production of siRNA or binding of siRNA to AGO. The one exception is the 2b suppressor of Cucumoviruses that binds to and inhibits AGO1. Here we describe a novel suppressor mechanism in which a Polerovirus-encoded F box protein (P0) targets the PAZ motif and its adjacent upstream sequence in AGO1 and mediates its degradation. F box proteins are components of E3 ubiquitin ligase complexes that add polyubiquitin tracts on selected lysine residues and thereby mark a protein for proteasome-mediated degradation. With P0, however, the targeted degradation of AGO is insensitive to inhibition of the proteasome, indicating that the proteasome is not involved. We also show that P0 does not block a mobile signal of silencing, indicating that the signal molecule does not have AGO protein components. The ability of P0 to block silencing without affecting signal movement may contribute to the phloem restriction of viruses in the Polerovirus group.

  5. Suppressor of cytokine signalling (SOCS)-3 protects beta cells against IL-1beta-mediated toxicity through inhibition of multiple nuclear factor-kappaB-regulated proapoptotic pathways

    DEFF Research Database (Denmark)

    Karlsen, Allan Ertman; Heding, P E; Frobøse, H

    2004-01-01

    The proinflammatory cytokine IL-1beta induces apoptosis in pancreatic beta cells via pathways dependent on nuclear factor-kappaB (NF-kappaB), mitogen-activated protein kinase, and protein kinase C. We recently showed suppressor of cytokine signalling (SOCS)-3 to be a natural negative feedback reg...... regulator of IL-1beta- and IFN-gamma-mediated signalling in rat islets and beta cell lines, preventing their deleterious effects. However, the mechanisms underlying SOCS-3 inhibition of IL-1beta signalling and prevention against apoptosis remain unknown....

  6. MicroRNA-34a is a potent tumor suppressor molecule in vivo in neuroblastoma

    LENUS (Irish Health Repository)

    Tivnan, Amanda

    2011-01-25

    ABSTRACT Background Neuroblastoma is a paediatric cancer which originates from precursor cells of the sympathetic nervous system and accounts for 15% of childhood cancer mortalities. With regards to the role of miRNAs in neuroblastoma, miR-34a, mapping to a chromosome 1p36 region that is commonly deleted, has been found to act as a tumor suppressor through targeting of numerous genes associated with cell proliferation and apoptosis. Methods A synthetic miR-34a (or negative control) precursor molecule was transfected into NB1691luc and SK-N-ASluc neuroblastoma cells. Quantitative PCR was used to verify increased miR-34a levels in NB1691luc and SK-N-ASluc cell lines prior to in vitro and in vivo analysis. In vitro analysis of the effects of miR-34a over expression on cell growth, cell cycle and phosphoprotein activation in signal transduction pathways was performed. Neuroblastoma cells over expressing miR-34a were injected retroperitoneally into immunocompromised CB17-SCID mice and tumor burden was assessed over a 21 day period by measuring bioluminescence (photons\\/sec\\/cm2). Results Over expression of miR-34a in both NB1691luc and SK-N-ASluc neuroblastoma cell lines led to a significant decrease in cell number relative to premiR-negative control treated cells over a 72 hour period. Flow cytometry results indicated that miR-34a induced cell cycle arrest and subsequent apoptosis activation. Phosphoprotein analysis highlighted key elements involved in signal transduction, whose activation was dysregulated as a result of miR-34a introduction into cells. As a potential mechanism of miR-34a action on phosphoprotein levels, we demonstrate that miR-34a over-expression results in a significant reduction of MAP3K9 mRNA and protein levels. Although MAP3K9 is a predicted target of miR-34a, direct targeting could not be validated with luciferase reporter assays. Despite this fact, any functional effects of reduced MAP3K9 expression as a result of miR-34a would be expected to

  7. Acrolein-induced activation of mitogen-activated protein kinase signaling is mediated by alkylation of thioredoxin reductase and thioredoxin 1.

    Science.gov (United States)

    Randall, Matthew J; Spiess, Page C; Hristova, Milena; Hondal, Robert J; van der Vliet, Albert

    2013-01-01

    Cigarette smoking remains a major health concern worldwide, and many of the adverse effects of cigarette smoke (CS) can be attributed to its abundant electrophilic aldehydes, such as acrolein (2-propenal). Previous studies indicate that acrolein readily reacts with thioredoxin reductase 1 (TrxR1), a critical enzyme involved in regulation of thioredoxin (Trx)-mediated redox signaling, by alkylation at its selenocysteine (Sec) residue. Because alkylation of Sec within TrxR1 has significant implications for its enzymatic function, we explored the potential importance of TrxR1 alkylation in acrolein-induced activation or injury of bronchial epithelial cells. Exposure of human bronchial epithelial HBE1 cells to acrolein (1-30 μM) resulted in dose-dependent loss of TrxR thioredoxin reductase activity, which coincided with its alkylation, as determined by biotin hydrazide labeling, and was independent of initial GSH status. To test the involvement of TrxR1 in acrolein responses in HBE1 cells, we suppressed TrxR1 using siRNA silencing or augmented TrxR1 by cell supplementation with sodium selenite. Acrolein exposure of HBE1 cells induced dose-dependent activation of the MAP kinases, extracellular regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38, and activation of JNK was markedly enhanced after selenite-mediated induction of TrxR1, and was associated with increased alkylation of TrxR1. Conversely, siRNA silencing of TrxR1 significantly suppressed the ability of acrolein to activate JNK, and also appeared to attenuate acrolein-dependent activation of ERK and p38. Alteration of initial TrxR1 levels by siRNA or selenite supplementation also affected initial Trx1 redox status and acrolein-mediated alkylation of Trx1, but did not significantly affect acrolein-mediated activation of HO-1 or cytotoxicity. Collectively, our findings indicate that alkylation of TrxR1 and/or Trx1 may contribute directly to acrolein-mediated activation of MAP kinases such as JNK, and

  8. Acrolein-induced activation of mitogen-activated protein kinase signaling is mediated by alkylation of thioredoxin reductase and thioredoxin 1

    Directory of Open Access Journals (Sweden)

    Matthew J. Randall

    2013-01-01

    Full Text Available Cigarette smoking remains a major health concern worldwide, and many of the adverse effects of cigarette smoke (CS can be attributed to its abundant electrophilic aldehydes, such as acrolein (2-propenal. Previous studies indicate that acrolein readily reacts with thioredoxin reductase 1 (TrxR1, a critical enzyme involved in regulation of thioredoxin (Trx-mediated redox signaling, by alkylation at its selenocysteine (Sec residue. Because alkylation of Sec within TrxR1 has significant implications for its enzymatic function, we explored the potential importance of TrxR1 alkylation in acrolein-induced activation or injury of bronchial epithelial cells. Exposure of human bronchial epithelial HBE1 cells to acrolein (1–30 μM resulted in dose-dependent loss of TrxR thioredoxin reductase activity, which coincided with its alkylation, as determined by biotin hydrazide labeling, and was independent of initial GSH status. To test the involvement of TrxR1 in acrolein responses in HBE1 cells, we suppressed TrxR1 using siRNA silencing or augmented TrxR1 by cell supplementation with sodium selenite. Acrolein exposure of HBE1 cells induced dose-dependent activation of the MAP kinases, extracellular regulated kinase (ERK, c-Jun N-terminal kinase (JNK, and p38, and activation of JNK was markedly enhanced after selenite-mediated induction of TrxR1, and was associated with increased alkylation of TrxR1. Conversely, siRNA silencing of TrxR1 significantly suppressed the ability of acrolein to activate JNK, and also appeared to attenuate acrolein-dependent activation of ERK and p38. Alteration of initial TrxR1 levels by siRNA or selenite supplementation also affected initial Trx1 redox status and acrolein-mediated alkylation of Trx1, but did not significantly affect acrolein-mediated activation of HO-1 or cytotoxicity. Collectively, our findings indicate that alkylation of TrxR1 and/or Trx1 may contribute directly to acrolein-mediated activation of MAP kinases

  9. Acrolein-induced activation of mitogen-activated protein kinase signaling is mediated by alkylation of thioredoxin reductase and thioredoxin 1☆☆☆

    Science.gov (United States)

    Randall, Matthew J.; Spiess, Page C.; Hristova, Milena; Hondal, Robert J.; van der Vliet, Albert

    2013-01-01

    Cigarette smoking remains a major health concern worldwide, and many of the adverse effects of cigarette smoke (CS) can be attributed to its abundant electrophilic aldehydes, such as acrolein (2-propenal). Previous studies indicate that acrolein readily reacts with thioredoxin reductase 1 (TrxR1), a critical enzyme involved in regulation of thioredoxin (Trx)-mediated redox signaling, by alkylation at its selenocysteine (Sec) residue. Because alkylation of Sec within TrxR1 has significant implications for its enzymatic function, we explored the potential importance of TrxR1 alkylation in acrolein-induced activation or injury of bronchial epithelial cells. Exposure of human bronchial epithelial HBE1 cells to acrolein (1–30 μM) resulted in dose-dependent loss of TrxR thioredoxin reductase activity, which coincided with its alkylation, as determined by biotin hydrazide labeling, and was independent of initial GSH status. To test the involvement of TrxR1 in acrolein responses in HBE1 cells, we suppressed TrxR1 using siRNA silencing or augmented TrxR1 by cell supplementation with sodium selenite. Acrolein exposure of HBE1 cells induced dose-dependent activation of the MAP kinases, extracellular regulated1 kinase (ERK), c-Jun N-terminal kinase (JNK), and p38, and activation of JNK was markedly enhanced after selenite-mediated induction of TrxR1, and was associated with increased alkylation of TrxR1. Conversely, siRNA silencing of TrxR1 significantly suppressed the ability of acrolein to activate JNK, and also appeared to attenuate acrolein-dependent activation of ERK and p38. Alteration of initial TrxR1 levels by siRNA or selenite supplementation also affected initial Trx1 redox status and acrolein-mediated alkylation of Trx1, but did not significantly affect acrolein-mediated activation of HO-1 or cytotoxicity. Collectively, our findings indicate that alkylation of TrxR1 and/or Trx1 may contribute directly to acrolein-mediated activation of MAP kinases such as JNK

  10. Tumor suppressor p16 INK4a: Downregulation of galectin-3, an endogenous competitor of the pro-anoikis effector galectin-1, in a pancreatic carcinoma model.

    Science.gov (United States)

    Sanchez-Ruderisch, Hugo; Fischer, Christian; Detjen, Katharina M; Welzel, Martina; Wimmel, Anja; Manning, Joachim C; André, Sabine; Gabius, Hans-Joachim

    2010-09-01

    The tumor suppressor p16(INK4a) has functions beyond cell-cycle control via cyclin-dependent kinases. A coordinated remodeling of N- and O-glycosylation, and an increase in the presentation of the endogenous lectin galectin-1 sensing these changes on the surface of p16(INK4a)-expressing pancreatic carcinoma cells (Capan-1), lead to potent pro-anoikis signals. We show that the p16(INK4a)-dependent impact on growth-regulatory lectins is not limited to galectin-1, but also concerns galectin-3. By monitoring its expression in relation to p16(INK4a) status, as well as running anoikis assays with galectin-3 and cell transfectants with up- or downregulated lectin expression, a negative correlation between anoikis and the presence of this lectin was established. Nuclear run-off and northern blotting experiments revealed an effect of the presence of p16(INK4a) on steady-state levels of galectin-3-specific mRNA that differed from decreasing the transcriptional rate. On the cell surface, galectin-3 interferes with galectin-1, which initiates signaling toward its pro-anoikis activity via caspase-8 activation. The detected opposite effects of p16(INK4a) at the levels of growth-regulatory galectins-1 and -3 shift the status markedly towards the galectin-1-dependent pro-anoikis activity. A previously undescribed orchestrated fine-tuning of this effector system by a tumor suppressor is discovered.

  11. Cyclophosphamide-induced myeloid-derived suppressor cell population is immunosuppressive but not identical to myeloid-derived suppressor cells induced by growing TC-1 tumors

    Czech Academy of Sciences Publication Activity Database

    Mikyšková, Romana; Indrová, Marie; Polláková, Veronika; Bieblová, Jana; Šímová, Jana; Reiniš, Milan

    2012-01-01

    Roč. 35, č. 5 (2012), s. 374-384 ISSN 1524-9557 R&D Projects: GA ČR(CZ) GPP301/11/P220; GA ČR GA301/09/1024; GA ČR GA301/07/1410 EU Projects: European Commission(XE) 18933 - CLINIGENE Institutional research plan: CEZ:AV0Z50520514 Institutional support: RVO:68378050 Keywords : myeloid-derived suppressor cells * cyclophosphamide * all-trans-retinoic acid * IL-12 * HPV16 Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.463, year: 2012

  12. Effect of duct shape, Mach number, and lining construction on measured suppressor attenuation and comparison with theory

    Science.gov (United States)

    Olsen, W. A.; Krejsa, E. A.; Coats, J. W.

    1972-01-01

    Noise attenuation was measured for several types of cylindrical suppressors that use a duct lining composed of honeycomb cells covered with a perforated plate. The experimental technique used gave attenuation data that were repeatable and free of noise floors and other sources of error. The suppressor length, the effective acoustic diameter, suppressor shape and flow velocity were varied. The agreement among the attenuation data and two widely used analytical models was generally satisfactory. Changes were also made in the construction of the acoustic lining to measure their effect on attenuation. One of these produced a very broadband muffler.

  13. ETS transcription factors control transcription of EZH2 and epigenetic silencing of the tumor suppressor gene Nkx3.1 in prostate cancer.

    Directory of Open Access Journals (Sweden)

    Paolo Kunderfranco

    2010-05-01

    Full Text Available ETS transcription factors regulate important signaling pathways involved in cell differentiation and development in many tissues and have emerged as important players in prostate cancer. However, the biological impact of ETS factors in prostate tumorigenesis is still debated.We performed an analysis of the ETS gene family using microarray data and real-time PCR in normal and tumor tissues along with functional studies in normal and cancer cell lines to understand the impact in prostate tumorigenesis and identify key targets of these transcription factors. We found frequent dysregulation of ETS genes with oncogenic (i.e., ERG and ESE1 and tumor suppressor (i.e., ESE3 properties in prostate tumors compared to normal prostate. Tumor subgroups (i.e., ERG(high, ESE1(high, ESE3(low and NoETS tumors were identified on the basis of their ETS expression status and showed distinct transcriptional and biological features. ERG(high and ESE3(low tumors had the most robust gene signatures with both distinct and overlapping features. Integrating genomic data with functional studies in multiple cell lines, we demonstrated that ERG and ESE3 controlled in opposite direction transcription of the Polycomb Group protein EZH2, a key gene in development, differentiation, stem cell biology and tumorigenesis. We further demonstrated that the prostate-specific tumor suppressor gene Nkx3.1 was controlled by ERG and ESE3 both directly and through induction of EZH2.These findings provide new insights into the role of the ETS transcriptional network in prostate tumorigenesis and uncover previously unrecognized links between aberrant expression of ETS factors, deregulation of epigenetic effectors and silencing of tumor suppressor genes. The link between aberrant ETS activity and epigenetic gene silencing may be relevant for the clinical management of prostate cancer and design of new therapeutic strategies.

  14. C/EBPalpha: a tumour suppressor in multiple tissues?

    DEFF Research Database (Denmark)

    Schuster, Mikkel Bruhn; Porse, Bo Torben

    2006-01-01

    . In line with its anti-mitotic properties, C/EBPalpha has been shown to interact with, and alter the activities of, several cell cycle related proteins and a number of models as to the mechanistics of C/EBPalpha-mediated growth repression have been proposed. More recently, several reports have indicated...

  15. Janus-Faced Myeloid-Derived Suppressor Cell Exosomes for the Good and the Bad in Cancer and Autoimmune Disease

    Directory of Open Access Journals (Sweden)

    Margot Zöller

    2018-02-01

    Full Text Available Myeloid-derived suppressor cells (MDSCs are a heterogeneous population of immature myeloid cells originally described to hamper immune responses in chronic infections. Meanwhile, they are known to be a major obstacle in cancer immunotherapy. On the other hand, MDSC can interfere with allogeneic transplant rejection and may dampen autoreactive T cell activity. Whether MDSC-Exosomes (Exo can cope with the dangerous and potentially therapeutic activities of MDSC is not yet fully explored. After introducing MDSC and Exo, it will be discussed, whether a blockade of MDSC-Exo could foster the efficacy of immunotherapy in cancer and mitigate tumor progression supporting activities of MDSC. It also will be outlined, whether application of native or tailored MDSC-Exo might prohibit autoimmune disease progression. These considerations are based on the steadily increasing knowledge on Exo composition, their capacity to distribute throughout the organism combined with selectivity of targeting, and the ease to tailor Exo and includes open questions that answers will facilitate optimizing protocols for a MDSC-Exo blockade in cancer as well as for strengthening their therapeutic efficacy in autoimmune disease.

  16. Cystic Fibrosis Transmembrane Conductance Regulator Attaches Tumor Suppressor PTEN to the Membrane and Promotes Anti Pseudomonas aeruginosa Immunity.

    Science.gov (United States)

    Riquelme, Sebastián A; Hopkins, Benjamin D; Wolfe, Andrew L; DiMango, Emily; Kitur, Kipyegon; Parsons, Ramon; Prince, Alice

    2017-12-19

    The tumor suppressor PTEN controls cell proliferation by regulating phosphatidylinositol-3-kinase (PI3K) activity, but the participation of PTEN in host defense against bacterial infection is less well understood. Anti-inflammatory PI3K-Akt signaling is suppressed in patients with cystic fibrosis (CF), a disease characterized by hyper-inflammatory responses to airway infection. We found that Ptenl -/- mice, which lack the NH 2 -amino terminal splice variant of PTEN, were unable to eradicate Pseudomonas aeruginosa from the airways and could not generate sufficient anti-inflammatory PI3K activity, similar to what is observed in CF. PTEN and the CF transmembrane conductance regulator (CFTR) interacted directly and this interaction was necessary to position PTEN at the membrane. CF patients under corrector-potentiator therapy, which enhances CFTR transport to the membrane, have increased PTEN amounts. These findings suggest that improved CFTR trafficking could enhance P. aeruginosa clearance from the CF airway by activating PTEN-mediated anti-bacterial responses and might represent a therapeutic strategy. Published by Elsevier Inc.

  17. Gemfibrozil, a lipid-lowering drug, induces suppressor of cytokine signaling 3 in glial cells: implications for neurodegenerative disorders.

    Science.gov (United States)

    Ghosh, Arunava; Pahan, Kalipada

    2012-08-03

    Glial inflammation is an important feature of several neurodegenerative disorders. Suppressor of cytokine signaling (SOCS) proteins play a crucial role in inhibiting cytokine signaling and inflammatory gene expression in various cell types, including glial cells. However, mechanisms by which SOCS genes could be up-regulated are poorly understood. This study underlines the importance of gemfibrozil, a Food and Drug Administration-approved lipid-lowering drug, in up-regulating the expression of SOCS3 in glial cells. Gemfibrozil increased the expression of Socs3 mRNA and protein in mouse astroglia and microglia in both a time- and dose-dependent manner. Interestingly, gemfibrozil induced the activation of type IA phosphatidylinositol (PI) 3-kinase and AKT. Accordingly, inhibition of PI 3-kinase and AKT by chemical inhibitors abrogated gemfibrozil-mediated up-regulation of SOCS3. Furthermore, we demonstrated that gemfibrozil induced the activation of Krüppel-like factor 4 (KLF4) via the PI 3-kinase-AKT pathway and that siRNA knockdown of KLF4 abrogated gemfibrozil-mediated up-regulation of SOCS3. Gemfibrozil also induced the recruitment of KLF4 to the distal, but not proximal, KLF4-binding site of the Socs3 promoter. This study delineates a novel property of gemfibrozil in up-regulating SOCS3 in glial cells via PI 3-kinase-AKT-mediated activation of KLF4 and suggests that gemfibrozil may find therapeutic application in neuroinflammatory and neurodegenerative disorders.

  18. CD40 dependent exacerbation of immune mediated hepatitis by hepatic CD11b+ Gr-1+ myeloid derived suppressor cells in tumor bearing mice

    Science.gov (United States)

    Kapanadze, Tamar; Medina-Echeverz, José; Gamrekelashvili, Jaba; Weiss, Jonathan M.; Wiltrout, Robert H.; Kapoor, Veena; Hawk, Nga; Terabe, Masaki; Berzofsky, Jay A.; Manns, Michael P.; Wang, Ena; Marincola, Francesco M.; Korangy, Firouzeh; Greten, Tim F.

    2015-01-01

    Immunosuppressive CD11b+Gr-1+ myeloid-derived suppressor cells (MDSC) accumulate in the livers of tumor-bearing mice. We studied hepatic MDSC in two murine models of immune mediated hepatitis. Unexpectedly, treatment of tumor bearing mice with Concanavalin A or α-Galactosylceramide resulted in increased ALT and AST serum levels in comparison to tumor free mice. Adoptive transfer of hepatic MDSC into naïve mice exacerbated Concanavalin A induced liver damage. Hepatic CD11b+Gr-1+ cells revealed a polarized pro-inflammatory gene signature after Concanavalin A treatment. An interferon gamma- dependent up-regulation of CD40 on hepatic CD11b+Gr-1+ cells along with an up-regulation of CD80, CD86, and CD1d after Concanavalin A treatment was observed. Concanavalin A treatment resulted in a loss of suppressor function by tumor-induced CD11b+Gr-1+ MDSC as well as enhanced reactive oxygen species-mediated hepatotoxicity. CD40 knockdown in hepatic MDSC led to increased arginase activity upon Concanavalin A treatment and lower ALT/AST serum levels. Finally, blockade of arginase activity in Cd40−/− tumor-induced myeloid cells resulted in exacerbation of hepatitis and increased reactive oxygen species production in vivo. Our findings indicate that in a setting of acute hepatitis, tumor-induced hepatic MDSC act as pro-inflammatory immune effector cells capable of killing hepatocytes in a CD40-dependent manner. PMID:25616156

  19. A novel cervical cancer suppressor 3 (CCS-3) interacts with the BTB domain of PLZF and inhibits the cell growth by inducing apoptosis.

    Science.gov (United States)

    Rho, Seung Bae; Park, Young Gyo; Park, Kyoungsook; Lee, Seung-Hoon; Lee, Je-Ho

    2006-07-24

    Promyelocytic leukemia zinc finger protein (PLZF) is a sequence-specific, DNA binding, transcriptional repressor differentially expressed during embryogenesis and in adult tissues. PLZF is known to be a negative regulator of cell cycle progression. We used PLZF as bait in a yeast two-hybrid screen with a cDNA library from the human ovary tissue. A novel cervical cancer suppressor 3 (CCS-3) was identified as a PLZF interacting partner. Further characterization revealed the BTB domain as an interacting domain of PLZF. Interaction of CCS-3 with PLZF in mammalian cells was also confirmed by co-immunoprecipitation and in vitro binding assays. It was found that, although CCS-3 shares similar homology with eEF1A, the study determined CCS-3 to be an isoform. CCS-3 was observed to be downregulated in human cervical cell lines as well as in cervical tumors when compared to those from normal tissues. Overexpression of CCS-3 in human cervical cell lines inhibits cell growth by inducing apoptosis and suppressing human cyclin A2 promoter activity. These combined results suggest that the potential tumor suppressor activity of CCS-3 may be mediated by its interaction with PLZF.

  20. In vivo evidence for CD4+ and CD8+ suppressor T cells in vaccination-induced suppression of murine experimental autoimmune thyroiditis

    International Nuclear Information System (INIS)

    Flynn, J.C.; Kong, Y.C.

    1991-01-01

    In several experimental autoimmune diseases, including experimental autoimmune thyroiditis (EAT), vaccination with attenuated autoantigen-specific T cells has provided protection against subsequent induction of disease. However, the mechanism(s) of vaccination-induced suppression remains to be clarified. Since the authors have previously shown that suppression generated by pretreatment with mouse thyroglobulin (MTg) or thyroid-stimulating hormone in EAT is mediated by CD4+, not CD8+, suppressor T cells, they examined the role of T cell subsets in vaccination-induced suppression of EAT. Mice were vaccinated with irradiated, MTg-primed, and MTg-activated spleen cells and then challenged. Pretreatment with these cells suppressed EAT induced by immunization with MTg and adjuvant, but not by adoptive transfer of thyroiditogenic cells, suggesting a mechanism of afferent suppression. The activation of suppressor mechanisms did not require CD8+ cells, since mice depleted of CD8+ cells before vaccination showed reduced EAT comparable to control vaccinated mice. Furthermore, depletion of either the CD4+ or the CD8+ subset after vaccination did not significantly abrogate suppression. However, suppression was eliminated by the depletion of both CD4+ and CD8+ cells in vaccinated mice. These results provide evidence for the cooperative effects of CD4+ and CD8+ T cells in vaccination-induced suppression of EAT

  1. System and method for multi-stage bypass, low operating temperature suppressor for automatic weapons

    Science.gov (United States)

    Moss, William C.; Anderson, Andrew T.

    2015-06-09

    The present disclosure relates to a suppressor for use with a weapon. The suppressor may be formed to have a body portion having a bore extending concentric with a bore axis of the weapon barrel. An opening in the bore extends at least substantially circumferentially around the bore. A flow path communicates with the opening and defines a channel for redirecting gasses flowing in the bore out from the bore, through the opening, into a rearward direction in the flow path. The flow path raises a pressure at the opening to generate a Mach disk within the bore at a location approximately coincident with the opening. The Mach disk forms as a virtual baffle to divert at least a portion of the gasses into the opening and into the flow path.

  2. Tumor suppressor microRNAs are downregulated in myelodysplastic syndrome with spliceosome mutations

    DEFF Research Database (Denmark)

    Aslan, Derya; Garde, Christian; Nygaard, Mette Katrine

    2016-01-01

    Spliceosome mutations are frequently observed in patients with myelodysplastic syndromes (MDS). However, it is largely unknown how these mutations contribute to the disease. MicroRNAs (miRNAs) are small noncoding RNAs, which have been implicated in most human cancers due to their role in post...... the most downregulated miRNAs were several tumor-suppressor miRNAs, including several let-7 family members, miR-423, and miR-103a. Finally, we observed that the predicted targets of the most downregulated miRNAs were involved in apoptosis, hematopoiesis, and acute myeloid leukemia among other cancer......- and metabolic pathways. Our data indicate that spliceosome mutations may play an important role in MDS pathophysiology by affecting the expression of tumor suppressor miRNA genes involved in the development and progression of MDS....

  3. Noise suppression and crosstalk analysis of on-chip magnetic film-type noise suppressor

    Science.gov (United States)

    Ma, Jingyan; Muroga, Sho; Endo, Yasushi; Hashi, Shuichiro; Naoe, Masayuki; Yokoyama, Hiroo; Hayashi, Yoshiaki; Ishiyama, Kazushi

    2018-05-01

    This paper discusses near field, conduction and crosstalk noise suppression of magnetic films with uniaxial anisotropy on transmission lines for a film-type noise suppressor in the GHz frequency range. The electromagnetic noise suppressions of magnetic films with different permeability and resistivity were measured and simulated with simple microstrip lines. The experimental and simulated results of Co-Zr-Nb and CoPd-CaF2 films agreed with each other. The results indicate that the higher permeability leads to a better near field shielding, and in the frequency range of 2-7 GHz, a higher conduction noise suppression. It also suggests that the higher resistivity results in a better crosstalk suppression in the frequency range below 2 GHz. These results can support the design guidelines of the magnetic film-type noise suppressor used in the next generation IC chip.

  4. Proton cross-talk and losses in the dispersion suppressor regions at the FCC-hh

    CERN Document Server

    AUTHOR|(CDS)2100784; Appleby, Robert Barrie; Krainer, Alexander; Langner, Andy Sven; Abelleira, Jose

    2017-01-01

    Protons that collide at the interaction points of the FCC-hh may contribute to the background in the subsequent detector. Due to the high luminosity of the proton beams this may be of concern. Using DPMJET-III to model 50 TeV proton-proton collisions, tracking studies have been performed with PTC and MERLIN in order to gauge the elastic and inelastic proton cross-talk. High arc losses, particularly in the dispersion suppressor regions, have been revealed. These losses originate mainly from particles with a momentum deviation, either from interaction with a primary collimator in the betatron cleaning insertion, or from the proton-proton collisions. This issue can be mitigated by introducing additional collimators in the dispersion suppressor region. The specific design, lattice integration, and the effect of these collimators on cross-talk is assessed.

  5. Characteristics of DTH suppressor cells in mice infected with Candida albicans.

    Science.gov (United States)

    Valdez, J C; Mesón, O E; Sirena, A; de Alderete, N G

    1987-05-01

    Inoculation of 10(8) C. albicans intraperitoneally into Balb/c mice at given dosage was reported to induce suppression of antigen-specific delayed-type hypersensitivity. Adoptive transfer of spleen cells into normal syngeneic mice pre-treated with Cyclophosphamide confirmed the existence of suppressor cells in mice. Such cells were sensitive to treatment with anti-theta serum and complement, non-adherent to Sephadex G-10. A pretreatment of the mice with Cyclophosphamide eliminated DTH suppression. Treatment with antimacrophage agents via intraperitoneal abrogated suppression only if being effected before inoculation of alive 10(8) Candida albicans. It is concluded that the spleen suppressor cell is a T-lymphocyte whose precursor is Cyclophosphamide-sensitive, requiring the macrophage to be induced.

  6. Analysis of a Novel 17q25 Cell Cycle Gene Homolog: Is it a Breast Tumor Suppressor Gene?

    National Research Council Canada - National Science Library

    Kalikin, Linda

    2000-01-01

    ... of these molecular reagents into successful tools for the medical management of breast cancer. We hypothesize that a 350 kb region on 17q25 detected by our allelic imbalance studies harbors a novel breast tumor suppressor gene...

  7. Dissecting functions of the retinoblastoma tumor suppressor and the related pocket proteins by integrating genetic, cell biology, and electrophoretic techniques

    DEFF Research Database (Denmark)

    Hansen, Klaus; Lukas, J; Holm, K

    1999-01-01

    The members of the 'pocket protein' family, comprising the retinoblastoma tumor suppressor (pRB) and its relatives, p107 and p130, negatively regulate cell proliferation and modulate fundamental biological processes including embryonic development, differentiation, homeostatic tissue renewal...

  8. Generation of two modified mouse alleles of the Hic1 tumor suppressor gene

    Czech Academy of Sciences Publication Activity Database

    Pospíchalová, Vendula; Turečková, Jolana; Fafílek, Bohumil; Vojtěchová, Martina; Krausová, Michaela; Lukáš, Jan; Šloncová, Eva; Takacova, S.; Divoký, V.; Leprince, D.; Plachý, Jiří; Kořínek, Vladimír

    2011-01-01

    Roč. 49, č. 3 (2011), s. 142-151 ISSN 1526-954X R&D Projects: GA ČR(CZ) GA204/07/1567; GA ČR(CZ) GD204/09/H058 Institutional research plan: CEZ:AV0Z50520514 Keywords : Hypermethylated In Cancer 1 * Hic1 tumor suppressor * gene targeting Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.527, year: 2011

  9. Clinical Impact of the Immunome in Lymphoid Malignancies: The Role of Myeloid-Derived Suppressor Cells

    Science.gov (United States)

    Vetro, Calogero; Romano, Alessandra; Ancora, Flavia; Coppolino, Francesco; Brundo, Maria V.; Raccuia, Salvatore A.; Puglisi, Fabrizio; Tibullo, Daniele; La Cava, Piera; Giallongo, Cesarina; Parrinello, Nunziatina L.

    2015-01-01

    The better definition of the mutual sustainment between neoplastic cells and immune system has been translated from the bench to the bedside acquiring value as prognostic factor. Additionally, it represents a promising tool for improving therapeutic strategies. In this context, myeloid-derived suppressor cells (MDSCs) have gained a central role in tumor developing with consequent therapeutic implications. In this review, we will focus on the biological and clinical impact of the study of MDSCs in the settings of lymphoid malignancies. PMID:26052505

  10. Electrochemical sensing of tumor suppressor protein p53-deoxyribonucleic acid complex stability at an electrified interface

    Czech Academy of Sciences Publication Activity Database

    Paleček, Emil; Černocká, Hana; Ostatná, Veronika; Navrátilová, Lucie; Brázdová, Marie

    2014-01-01

    Roč. 828, MAY2014 (2014), s. 1-8 ISSN 0003-2670 R&D Projects: GA ČR(CZ) GAP301/11/2055; GA ČR(CZ) GA13-00956S; GA ČR(CZ) GA13-36108S Institutional support: RVO:68081707 Keywords : Deoxyribonucleic acid-protein binding * Tumor suppressor protein p53 * Electrochemical sensing Subject RIV: BO - Biophysics Impact factor: 4.513, year: 2014

  11. The Value of Suppressor Effects in Explicating the Construct Validity of Symptom Measures

    Science.gov (United States)

    Watson, David; Clark, Lee Anna; Chmielewski, Michael; Kotov, Roman

    2013-01-01

    Suppressor effects are operating when the addition of a predictor increases the predictive power of another variable. We argue that suppressor effects can play a valuable role in explicating the construct validity of symptom measures by bringing into clearer focus opposing elements that are inherent—but largely hidden—in the measure’s overall score. We illustrate this point using theoretically grounded, replicated suppressor effects that have emerged in analyses of the original Inventory of Depression and Anxiety Symptoms (IDAS; Watson et al., 2007) and its expanded second version (IDAS-II; Watson et al., 2012). In Study 1, we demonstrate that the IDAS-II Appetite Gain and Appetite Loss scales contain both (a) a shared distress component that creates a positive correlation between them and (b) a specific symptom component that produces a natural negative association between them (i.e., people who recently have experienced decreased interest in food/loss of appetite are less likely to report a concomitant increase in appetite/weight). In Study 2, we establish that mania scales also contain two distinct elements—namely, high energy/positive emotionality and general distress/dysfunction—that oppose each another in many instances. In both studies, we obtained evidence of suppression effects that were highly robust across different types of respondents (e.g., clinical outpatients, community adults, college students) and using both self-report and interview-based measures. These replicable suppressor effects establish that many homogeneous, unidimensional symptom scales actually contain distinguishable components with distinct—at times, even antagonistic—properties. PMID:23795886

  12. MicroRNA-34a is a potent tumor suppressor molecule in vivo in neuroblastoma.

    LENUS (Irish Health Repository)

    Tivnan, Amanda

    2011-01-01

    Neuroblastoma is a paediatric cancer which originates from precursor cells of the sympathetic nervous system and accounts for 15% of childhood cancer mortalities. With regards to the role of miRNAs in neuroblastoma, miR-34a, mapping to a chromosome 1p36 region that is commonly deleted, has been found to act as a tumor suppressor through targeting of numerous genes associated with cell proliferation and apoptosis.

  13. Identification of myeloid derived suppressor cells in the peripheral blood of tumor bearing dogs

    Directory of Open Access Journals (Sweden)

    Sherger Matthew

    2012-10-01

    Full Text Available Abstract Background Myeloid derived suppressor cells (MDSCs are a recently described population of immune cells that significantly contribute to the immunosuppression seen in cancer patients. MDSCs are one of the most important factors that limit the efficacy of cancer immunotherapy (e.g. cancer vaccines and MDSC levels are increased in cancer in multiple species. Identifying and targeting MDSCs is actively being investigated in the field of human oncology and is increasingly being investigated in veterinary oncology. The treatment of canine cancer not only benefits dogs, but is being used for translational studies evaluating and modifcying candidate therapies for use in humans. Thus, it is necessary to understand the immune alterations seen in canine cancer patients which, to date, have been relatively limited. This study investigates the use of commercially available canine antibodies to detect an immunosuppressive (CD11blow/CADO48low cell population that is increased in the peripheral blood of tumor-bearing dogs. Results Commercially available canine antibodies CD11b and CADO48A were used to evaluate white blood cells from the peripheral blood cells of forty healthy control dogs and forty untreated, tumor-bearing dogs. Tumor-bearing dogs had a statistically significant increase in CD11blow/CADO48Alow cells (7.9% as compared to the control dogs (3.6%. Additionally, sorted CD11blow/CADO48Alow generated in vitro suppressed the proliferation of canine lymphocytes. Conclusions The purpose of this study was aimed at identifying potential canine specific markers for identifying MDSCs in the peripheral blood circulation of dogs. This study demonstrates an increase in a unique CD11blow/CADO48Alow cell population in tumor-bearing dogs. This immunophenotype is consistent with described phenotypes of MDSCs in other species (i.e. mice and utilizes commercially available canine-specific antibodies. Importantly, CD11blow/CADO48Alow from a tumor environment

  14. Human microRNA oncogenes and tumor suppressors show significantly different biological patterns: from functions to targets.

    Directory of Open Access Journals (Sweden)

    Dong Wang

    Full Text Available MicroRNAs (miRNAs are small noncoding RNAs which play essential roles in many important biological processes. Therefore, their dysfunction is associated with a variety of human diseases, including cancer. Increasing evidence shows that miRNAs can act as oncogenes or tumor suppressors, and although there is great interest in research into these cancer-associated miRNAs, little is known about them. In this study, we performed a comprehensive analysis of putative human miRNA oncogenes and tumor suppressors. We found that miRNA oncogenes and tumor suppressors clearly show different patterns in function, evolutionary rate, expression, chromosome distribution, molecule size, free energy, transcription factors, and targets. For example, miRNA oncogenes are located mainly in the amplified regions in human cancers, whereas miRNA tumor suppressors are located mainly in the deleted regions. miRNA oncogenes tend to cleave target mRNAs more frequently than miRNA tumor suppressors. These results indicate that these two types of cancer-associated miRNAs play different roles in cancer formation and development. Moreover, the patterns identified here can discriminate novel miRNA oncogenes and tumor suppressors with a high degree of accuracy. This study represents the first large-scale bioinformatic analysis of human miRNA oncogenes and tumor suppressors. Our findings provide help for not only understanding of miRNAs in cancer but also for the specific identification of novel miRNAs as miRNA oncogenes and tumor suppressors. In addition, the data presented in this study will be valuable for the study of both miRNAs and cancer.

  15. Resistance to EGF receptor inhibitors in glioblastoma mediated by phosphorylation of the PTEN tumor suppressor at tyrosine 240.

    Science.gov (United States)

    Fenton, Tim R; Nathanson, David; Ponte de Albuquerque, Claudio; Kuga, Daisuke; Iwanami, Akio; Dang, Julie; Yang, Huijun; Tanaka, Kazuhiro; Oba-Shinjo, Sueli Mieko; Uno, Miyuki; Inda, Maria del Mar; Wykosky, Jill; Bachoo, Robert M; James, C David; DePinho, Ronald A; Vandenberg, Scott R; Zhou, Huilin; Marie, Suely K N; Mischel, Paul S; Cavenee, Webster K; Furnari, Frank B

    2012-08-28

    Glioblastoma multiforme (GBM) is the most aggressive of the astrocytic malignancies and the most common intracranial tumor in adults. Although the epidermal growth factor receptor (EGFR) is overexpressed and/or mutated in at least 50% of GBM cases and is required for tumor maintenance in animal models, EGFR inhibitors have thus far failed to deliver significant responses in GBM patients. One inherent resistance mechanism in GBM is the coactivation of multiple receptor tyrosine kinases, which generates redundancy in activation of phosphoinositide-3'-kinase (PI3K) signaling. Here we demonstrate that the phosphatase and tensin homolog deleted on chromosome 10 (PTEN) tumor suppressor is frequently phosphorylated at a conserved tyrosine residue, Y240, in GBM clinical samples. Phosphorylation of Y240 is associated with shortened overall survival and resistance to EGFR inhibitor therapy in GBM patients and plays an active role in mediating resistance to EGFR inhibition in vitro. Y240 phosphorylation can be mediated by both fibroblast growth factor receptors and SRC family kinases (SFKs) but does not affect the ability of PTEN to antagonize PI3K signaling. These findings show that, in addition to genetic loss and mutation of PTEN, its modulation by tyrosine phosphorylation has important implications for the development and treatment of GBM.

  16. Heterozygous Germline Mutations in the CBL Tumor-Suppressor Gene Cause a Noonan Syndrome-like Phenotype

    Science.gov (United States)

    Martinelli, Simone; De Luca, Alessandro; Stellacci, Emilia; Rossi, Cesare; Checquolo, Saula; Lepri, Francesca; Caputo, Viviana; Silvano, Marianna; Buscherini, Francesco; Consoli, Federica; Ferrara, Grazia; Digilio, Maria C.; Cavaliere, Maria L.; van Hagen, Johanna M.; Zampino, Giuseppe; van der Burgt, Ineke; Ferrero, Giovanni B.; Mazzanti, Laura; Screpanti, Isabella; Yntema, Helger G.; Nillesen, Willy M.; Savarirayan, Ravi; Zenker, Martin; Dallapiccola, Bruno; Gelb, Bruce D.; Tartaglia, Marco

    2010-01-01

    RAS signaling plays a key role in controlling appropriate cell responses to extracellular stimuli and participates in early and late developmental processes. Although enhanced flow through this pathway has been established as a major contributor to oncogenesis, recent discoveries have revealed that aberrant RAS activation causes a group of clinically related developmental disorders characterized by facial dysmorphism, a wide spectrum of cardiac disease, reduced growth, variable cognitive deficits, ectodermal and musculoskeletal anomalies, and increased risk for certain malignancies. Here, we report that heterozygous germline mutations in CBL, a tumor-suppressor gene that is mutated in myeloid malignancies and encodes a multivalent adaptor protein with E3 ubiquitin ligase activity, can underlie a phenotype with clinical features fitting or partially overlapping Noonan syndrome (NS), the most common condition of this disease family. Independent CBL mutations were identified in two sporadic cases and two families from among 365 unrelated subjects who had NS or suggestive features and were negative for mutations in previously identified disease genes. Phenotypic heterogeneity and variable expressivity were documented. Mutations were missense changes altering evolutionarily conserved residues located in the RING finger domain or the linker connecting this domain to the N-terminal tyrosine kinase binding domain, a known mutational hot spot in myeloid malignancies. Mutations were shown to affect CBL-mediated receptor ubiquitylation and dysregulate signal flow through RAS. These findings document that germline mutations in CBL alter development to cause a clinically variable condition that resembles NS and that possibly predisposes to malignancies. PMID:20619386

  17. DNA damage tolerance pathway involving DNA polymerase ι and the tumor suppressor p53 regulates DNA replication fork progression.

    Science.gov (United States)

    Hampp, Stephanie; Kiessling, Tina; Buechle, Kerstin; Mansilla, Sabrina F; Thomale, Jürgen; Rall, Melanie; Ahn, Jinwoo; Pospiech, Helmut; Gottifredi, Vanesa; Wiesmüller, Lisa

    2016-07-26

    DNA damage tolerance facilitates the progression of replication forks that have encountered obstacles on the template strands. It involves either translesion DNA synthesis initiated by proliferating cell nuclear antigen monoubiquitination or less well-characterized fork reversal and template switch mechanisms. Herein, we characterize a novel tolerance pathway requiring the tumor suppressor p53, the translesion polymerase ι (POLι), the ubiquitin ligase Rad5-related helicase-like transcription factor (HLTF), and the SWI/SNF catalytic subunit (SNF2) translocase zinc finger ran-binding domain containing 3 (ZRANB3). This novel p53 activity is lost in the exonuclease-deficient but transcriptionally active p53(H115N) mutant. Wild-type p53, but not p53(H115N), associates with POLι in vivo. Strikingly, the concerted action of p53 and POLι decelerates nascent DNA elongation and promotes HLTF/ZRANB3-dependent recombination during unperturbed DNA replication. Particularly after cross-linker-induced replication stress, p53 and POLι also act together to promote meiotic recombination enzyme 11 (MRE11)-dependent accumulation of (phospho-)replication protein A (RPA)-coated ssDNA. These results implicate a direct role of p53 in the processing of replication forks encountering obstacles on the template strand. Our findings define an unprecedented function of p53 and POLι in the DNA damage response to endogenous or exogenous replication stress.

  18. Neem leaf glycoprotein prevents post-surgical sarcoma recurrence in Swiss mice by differentially regulating cytotoxic T and myeloid-derived suppressor cells.

    Directory of Open Access Journals (Sweden)

    Madhurima Sarkar

    Full Text Available Post-surgical tumor recurrence is a common problem in cancer treatment. In the present study, the role of neem leaf glycoprotein (NLGP, a novel immunomodulator, in prevention of post-surgical recurrence of solid sarcoma was examined. Data suggest that NLGP prevents tumor recurrence after surgical removal of sarcoma in Swiss mice and increases their tumor-free survival time. In NLGP-treated tumor-free mice, increased cytotoxic CD8+ T cells and a decreased population of suppressor cells, especially myeloid-derived suppressor cells (MDSCs was observed. NLGP-treated CD8+ T cells showed greater cytotoxicity towards tumor-derived MDSCs and supernatants from the same CD8+ T cell culture caused upregulation of FasR and downregulation of cFLIP in MDSCs. To elucidate the role of CD8+ T cells, specifically in association with the downregulation in MDSCs, CD8+ T cells were depleted in vivo before NLGP immunization in surgically tumor removed mice and tumor recurrence was noted. These mice also exhibited increased MDSCs along with decreased levels of Caspase 3, Caspase 8 and increased cFLIP expression. In conclusion, it can be stated that NLGP, by activating CD8+ T cells, down regulates the proportion of MDSCs. Accordingly, suppressive effects of MDSCs on CD8+ T cells are minimized and optimum immune surveillance in tumor hosts is maintained to eliminate the residual tumor mass appearing during recurrence.

  19. Genetic and Epigenetic Tumor Suppressor Gene Silencing are Distinct Molecular Phenotypes Driven by Growth Promoting Mutations in Non small Cell Lung Cancer

    International Nuclear Information System (INIS)

    Marsit, C. J.; Kelsey, K. T.; Houseman, E. A.; Kelsey, K. T.; Houseman, E. A.; Nelson, H. H.

    2008-01-01

    Both genetic and epigenetic alterations characterize human non small cell lung cancer (NSCLC), but the biological processes that create or select these alterations remain incompletely investigated. Our hypothesis posits that a roughly reciprocal relationship between the propensity for promoter hyper methylation and a propensity for genetic deletion leads to distinct molecular phenotypes of lung cancer. To test this hypothesis, we examined promoter hyper methylation of 17 tumor suppressor genes, as a marker of epigenetic alteration propensity, and deletion events at the 3p21 region, as a marker of genetic alteration. To model the complex biology between these somatic alterations, we utilized an item response theory model. We demonstrated that tumors exhibiting LOH at greater than 30% of informative alleles in the 3p21 region have a significantly reduced propensity for hyper methylation. At the same time, tumors with activating KRAS mutations showed a significantly increased propensity for hyper methylation of the loci examined, a result similar to what has been observed in colon cancer. These data suggest that NSCLCs have distinct epigenetic or genetic alteration phenotypes acting upon tumor suppressor genes and that mutation of oncogenic growth promoting genes, such as KRAS, is associated with the epigenetic phenotype.

  20. Epidermal growth factor receptor signaling promotes metastatic prostate cancer through microRNA-96-mediated downregulation of the tumor suppressor ETV6.

    Science.gov (United States)

    Tsai, Yuan-Chin; Chen, Wei-Yu; Siu, Man Kit; Tsai, Hong-Yuan; Yin, Juan Juan; Huang, Jiaoti; Liu, Yen-Nien

    2017-01-01

    It has been suggested that ETV6 serves as a tumor suppressor; however, its molecular regulation and cellular functions remain unclear. We used prostate cancer as a model system and demonstrated a molecular mechanism in which ETV6 can be regulated by epidermal growth factor receptor (EGFR) signaling through microRNA-96 (miR-96)-mediated downregulation. In addition, EGFR acts as a transcriptional coactivator that binds to the promoter of primary miR-96 and transcriptionally regulates miR-96 levels. We analyzed two sets of clinical prostate cancer samples, confirmed association patterns that were consistent with the EGFR-miR-96-ETV6 signaling model and demonstrated that the reduced ETV6 levels were associated with malignant prostate cancer. Based on results derived from multiple approaches, we identified the biological functions of ETV6 as a tumor suppressor that inhibits proliferation and metastasis in prostate cancer. We present a molecular mechanism in which EGFR activation leads to the induction of miR-96 expression and suppression of ETV6, which contributes to prostate cancer progression. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  1. YNK1, the yeast homolog of human metastasis suppressor NM23, is required for repair of UV radiation- and etoposide-induced DNA damage

    Energy Technology Data Exchange (ETDEWEB)

    Yang Mengmeng; Jarrett, Stuart G.; Craven, Rolf [Department of Molecular and Biomedical Pharmacology, College of Medicine, University of Kentucky, Lexington, KY 40536-0298 (United States); Kaetzel, David M. [Department of Molecular and Biomedical Pharmacology, College of Medicine, University of Kentucky, Lexington, KY 40536-0298 (United States)], E-mail: dmkaetz@uky.edu

    2009-01-15

    In humans, NM23-H1 is a metastasis suppressor whose expression is reduced in metastatic melanoma and breast carcinoma cells, and which possesses the ability to inhibit metastatic growth without significant impact on the transformed phenotype. NM23-H1 exhibits three enzymatic activities in vitro, each with potential to maintain genomic stability, a 3'-5' exonuclease and two kinases, nucleoside diphosphate kinase (NDPK), and protein histidine kinase. Herein we have investigated the potential contributions of NM23 proteins to DNA repair in the yeast, Saccharomyces cerevisiae, which contains a single NM23 homolog, YNK1. Ablation of YNK1 delayed repair of UV- and etoposide-induced nuclear DNA damage by 3-6 h. However, YNK1 had no impact upon the kinetics of MMS-induced DNA repair. Furthermore, YNK1 was not required for repair of mitochondrial DNA damage. To determine whether the nuclear DNA repair deficit manifested as an increase in mutation frequency, the CAN1 forward assay was employed. An YNK1 deletion was associated with increased mutation rates following treatment with either UV (2.6x) or MMS (1.6x). Mutation spectral analysis further revealed significantly increased rates of base substitution and frameshift mutations following UV treatment in the ynk1{delta} strain. This study indicates a novel role for YNK1 in DNA repair in yeast, and suggests an anti-mutator function that may contribute to the metastasis suppressor function of NM23-H1 in humans.

  2. YNK1, the yeast homolog of human metastasis suppressor NM23, is required for repair of UV radiation- and etoposide-induced DNA damage

    International Nuclear Information System (INIS)

    Yang Mengmeng; Jarrett, Stuart G.; Craven, Rolf; Kaetzel, David M.

    2009-01-01

    In humans, NM23-H1 is a metastasis suppressor whose expression is reduced in metastatic melanoma and breast carcinoma cells, and which possesses the ability to inhibit metastatic growth without significant impact on the transformed phenotype. NM23-H1 exhibits three enzymatic activities in vitro, each with potential to maintain genomic stability, a 3'-5' exonuclease and two kinases, nucleoside diphosphate kinase (NDPK), and protein histidine kinase. Herein we have investigated the potential contributions of NM23 proteins to DNA repair in the yeast, Saccharomyces cerevisiae, which contains a single NM23 homolog, YNK1. Ablation of YNK1 delayed repair of UV- and etoposide-induced nuclear DNA damage by 3-6 h. However, YNK1 had no impact upon the kinetics of MMS-induced DNA repair. Furthermore, YNK1 was not required for repair of mitochondrial DNA damage. To determine whether the nuclear DNA repair deficit manifested as an increase in mutation frequency, the CAN1 forward assay was employed. An YNK1 deletion was associated with increased mutation rates following treatment with either UV (2.6x) or MMS (1.6x). Mutation spectral analysis further revealed significantly increased rates of base substitution and frameshift mutations following UV treatment in the ynk1Δ strain. This study indicates a novel role for YNK1 in DNA repair in yeast, and suggests an anti-mutator function that may contribute to the metastasis suppressor function of NM23-H1 in humans

  3. Genetic and Epigenetic Tumor Suppressor Gene Silencing Are Distinct Molecular Phenotypes Driven by Growth Promoting Mutations in Nonsmall Cell Lung Cancer

    Directory of Open Access Journals (Sweden)

    Carmen J. Marsit

    2008-01-01

    Full Text Available Both genetic and epigenetic alterations characterize human nonsmall cell lung cancer (NSCLC, but the biological processes that create or select these alterations remain incompletely investigated. Our hypothesis posits that a roughly reciprocal relationship between the propensity for promoter hypermethylation and a propensity for genetic deletion leads to distinct molecular phenotypes of lung cancer. To test this hypothesis, we examined promoter hypermethylation of 17 tumor suppressor genes, as a marker of epigenetic alteration propensity, and deletion events at the 3p21 region, as a marker of genetic alteration. To model the complex biology between these somatic alterations, we utilized an item response theory model. We demonstrated that tumors exhibiting LOH at greater than 30% of informative alleles in the 3p21 region have a significantly reduced propensity for hypermethylation. At the same time, tumors with activating KRAS mutations showed a significantly increased propensity for hypermethylation of the loci examined, a result similar to what has been observed in colon cancer. These data suggest that NSCLCs have distinct epigenetic or genetic alteration phenotypes acting upon tumor suppressor genes and that mutation of oncogenic growth promoting genes, such as KRAS, is associated with the epigenetic phenotype.

  4. Suppressor T-cell factor(s) display an altered pattern of Igh (immunoglobulin heavy chain locus) genetic restriction when developed in an Igh-congeneic host

    International Nuclear Information System (INIS)

    HayGlass, K.T.; Naides, S.J.; Benacerraf, B.; Sy, M.S.

    1985-01-01

    Suppressor T cell factor(s) (TsF 1 ) inhibit the in vivo priming of azobenzenearsonate-specific cytotoxic T-cell responses. The activity of TsF 1 is restricted by genes linked to Igh-1 allotypic markers. TsF 1 obtained from B6.Igh-1/sup n/ mice was unable to suppress the immune response in B6.Igh-1/sup b/ mice and vice versa. However, TsF 1 prepared from B6.Igh-1/sup n/ T cells parked in an Igh-congeneic B6.Igh-1/sup b/ environment displays an additional restriction specificity of the host. Thus, TsF 1 prepared from these Igh-chimeric mice suppressed immune responses in both B6.Igh-1/sup n/ (donor) and B6.Igh-1/sup b/ (recipient) mice but not in mice of the unrelated strain BALB/c.Igh-1/sup a/. The results indicate that the establishment of the suppressor T-cell repertoire is dependent not only upon the genetic background of the individual T cell but also upon the influence of Igh-linked determinants present when T-cell clones are selected during the response

  5. Suppressor of Cytokine Signaling (SOCS 5 utilises distinct domains for regulation of JAK1 and interaction with the adaptor protein Shc-1.

    Directory of Open Access Journals (Sweden)

    Edmond M Linossi

    Full Text Available Suppressor of Cytokine Signaling (SOCS5 is thought to act as a tumour suppressor through negative regulation of JAK/STAT and epidermal growth factor (EGF signaling. However, the mechanism/s by which SOCS5 acts on these two distinct pathways is unclear. We show for the first time that SOCS5 can interact directly with JAK via a unique, conserved region in its N-terminus, which we have termed the JAK interaction region (JIR. Co-expression of SOCS5 was able to specifically reduce JAK1 and JAK2 (but not JAK3 or TYK2 autophosphorylation and this function required both the conserved JIR and additional sequences within the long SOCS5 N-terminal region. We further demonstrate that SOCS5 can directly inhibit JAK1 kinase activity, although its mechanism of action appears distinct from that of SOCS1 and SOCS3. In addition, we identify phosphoTyr317 in Shc-1 as a high-affinity substrate for the SOCS5-SH2 domain and suggest that SOCS5 may negatively regulate EGF and growth factor-driven Shc-1 signaling by binding to this site. These findings suggest that different domains in SOCS5 contribute to two distinct mechanisms for regulation of cytokine and growth factor signaling.

  6. Adiponectin deficiency suppresses lymphoma growth in mice by modulating NK cells, CD8 T cells, and myeloid-derived suppressor cells.

    Science.gov (United States)

    Han, Sora; Jeong, Ae Lee; Lee, Sunyi; Park, Jeong Su; Kim, Kwang Dong; Choi, Inpyo; Yoon, Suk Ran; Lee, Myung Sok; Lim, Jong-Seok; Han, Seung Hyun; Yoon, Do Young; Yang, Young

    2013-05-01

    Previously, we found that adiponectin (APN) suppresses IL-2-induced NK cell activation by downregulating the expression of the IFN-γ-inducible TNF-related apoptosis-inducing ligand and Fas ligand. Although the antitumor function of APN has been reported in several types of solid tumors, with few controversial results, no lymphoma studies have been conducted. In this study, we assessed the role of APN in immune cell function, including NK cells, CTLs, and myeloid-derived suppressor cells, in EL4 and B16F10 tumor-bearing APN knockout (KO) mice. We observed attenuated EL4 growth in the APNKO mice. Increased numbers of splenic NK cells and splenic CTLs were identified under naive conditions and EL4-challenged conditions, respectively. In APNKO mice, splenic NK cells showed enhanced cytotoxicity with and without IL-2 stimulation. Additionally, there were decreased levels of myeloid-derived suppressor cell accumulation in the EL4-bearing APNKO mice. Enforced MHC class I expression on B16F10 cells led to attenuated growth of these tumors in APNKO mice. Thus, our results suggest that EL4 regression in APNKO mice is not only due to an enhanced antitumor immune response but also to a high level of MHC class I expression.

  7. Evolution and origin of merlin, the product of the Neurofibromatosis type 2 (NF2 tumor-suppressor gene

    Directory of Open Access Journals (Sweden)

    Omelyanchuk Leonid V

    2005-12-01

    Full Text Available Abstract Background Merlin, the product of the Neurofibromatosis type 2 (NF2 tumor suppressor gene, belongs to the ezrin-radixin-moesin (ERM subgroup of the protein 4.1 superfamily, which links cell surface glycoproteins to the actin cytoskeleton. While merlin's functional activity has been examined in mammalian and Drosophila models, little is understood about its evolution, diversity, and overall distribution among different taxa. Results By combining bioinformatic and phylogenetic approaches, we demonstrate that merlin homologs are present across a wide range of metazoan lineages. While the phylogenetic tree shows a monophyletic origin of the ERM family, the origin of the merlin proteins is robustly separated from that of the ERM proteins. The derivation of merlin is thought to be in early metazoa. We have also observed the expansion of the ERM-like proteins within the vertebrate clade, which occurred after its separation from Urochordata (Ciona intestinalis. Amino acid sequence alignment reveals the absence of an actin-binding site in the C-terminal region of all merlin proteins from various species but the presence of a conserved internal binding site in the N-terminal domain of the merlin and ERM proteins. In addition, a more conserved pattern of amino acid residues is found in the region containing the so-called "Blue Box," although some amino acid substitutions in this region exist in the merlin sequences of worms, fish, and Ciona. Examination of sequence variability at functionally significant sites, including the serine-518 residue, the phosphorylation of which modulates merlin's intra-molecular association and function as a tumor suppressor, identifies several potentially important sites that are conserved among all merlin proteins but divergent in the ERM proteins. Secondary structure prediction reveals the presence of a conserved α-helical domain in the central to C-terminal region of the merlin proteins of various species. The

  8. Cdh11 Acts as a Tumor Suppressor in a Murine Retinoblastoma Model by Facilitating Tumor Cell Death

    Science.gov (United States)

    Marchong, Mellone N.; Yurkowski, Christine; Ma, Clement; Spencer, Clarellen; Pajovic, Sanja; Gallie, Brenda L.

    2010-01-01

    CDH11 gene copy number and expression are frequently lost in human retinoblastomas and in retinoblastomas arising in TAg-RB mice. To determine the effect of Cdh11 loss in tumorigenesis, we crossed Cdh11 null mice with TAg-RB mice. Loss of Cdh11 had no gross morphological effect on the developing retina of Cdh11 knockout mice, but led to larger retinal volumes in mice crossed with TAg-RB mice (p = 0.01). Mice null for Cdh11 presented with fewer TAg-positive cells at postnatal day 8 (PND8) (p = 0.01) and had fewer multifocal tumors at PND28 (p = 0.016), compared to mice with normal Cdh11 alleles. However, tumor growth was faster in Cdh11-null mice between PND8 and PND84 (p = 0.003). In tumors of Cdh11-null mice, cell death was decreased 5- to 10-fold (p<0.03 for all markers), while proliferation in vivo remained unaffected (p = 0.121). Activated caspase-3 was significantly decreased and β-catenin expression increased in Cdh11 knockdown experiments in vitro. These data suggest that Cdh11 displays tumor suppressor properties in vivo and in vitro in murine retinoblastoma through promotion of cell death. PMID:20421947

  9. Expression of PML tumor suppressor in A 431 cells reduces cellular growth by inhibiting the epidermal growth factor receptor expression

    International Nuclear Information System (INIS)

    Vallian, S.; Chang, K.S.

    2004-01-01

    Our previous studies showed that the promyelocytic leukemia, PML, protein functions as a cellular and growth suppressor. Transient expression of PML was also found to repress the activity of the epidermal growth factor receptor gene promoter. In this study we have examined the effects of PML on A431 cells, which express a high level of + protein. The PML gene was introduced into the cells using the adenovirus-mediated gene transfer system. Western blot analysis on the extracts from the cells expressing PML showed a significant repression in the expression of the epidermal growth factor receptor protein. The cells were examined for growth and DNA synthesis. The data showed a marked reduction in both growth and DNA synthesis rate in the cells expressing PML compared with the control cells. Furthermore, in comparison with the controls, the cells expressing PML were found to be more in G1 phase, fewer in S and about the same number in the G2/M phase. This data clearly demonstrated that the repression of epidermal growth factor receptor expression in A 431 cells by PML was associated with inhibition of cell growth and alteration of the cell cycle distribution, suggesting a novel mechanism for the known growth inhibitory effects of PML

  10. Interleukin 33 in tumor microenvironment is crucial for the accumulation and function of myeloid-derived suppressor cells

    Science.gov (United States)

    Xiao, Peng; Wan, Xiaopeng; Cui, Bijun; Liu, Yang; Qiu, Chenyang; Rong, Jiabing; Zheng, Mingzhu; Song, Yinjing; Chen, Luoquan; He, Jia; Tan, Qinchun; Wang, Xiaojia; Shao, Xiying; Liu, Yuhua; Cao, Xuetao; Wang, Qingqing

    2016-01-01

    ABSTRACT Tumor-induced, myeloid-derived suppressor cells (MDSCs)-mediated immune dysfunction is an important mechanism that leads to tumor immune escape and the inefficacy of cancer immunotherapy. Importantly, tumor-infiltrating MDSCs have much stronger ability compared to MDSCs in the periphery. However, the mechanisms that tumor microenvironment induces the accumulation and function of MDSCs are poorly understood. Here, we report that Interleukin-33 (IL-33) – a cytokine which can be abundantly released in tumor tissues both in 4T1-bearing mice and breast cancer patients, is crucial for facilitating the expansion of MDSCs. IL-33 in tumor microenvironment reduces the apoptosis and sustains the survival of MDSCs through induction of autocrine secretion of GM-CSF, which forms a positive amplifying loop for MDSC accumulation. This is in conjunction with IL-33-driven induction of arginase-1 expression and activation of NF-κB and MAPK signaling in MDSCs which augments their immunosuppressive ability, and histone modifications were involved in IL-33 signaling in MDSCs. In ST2−/− mice, the defect of IL-33 signaling in MDSCs attenuates the immunosuppressive and pro-tumoral capacity of MDSCs. Our results identify IL-33 as a critical mediator that contributes to the abnormal expansion and enhanced immunosuppressive function of MDSCs within tumor microenvironment, which can be potentially targeted to reverse MDSC-mediated tumor immune evasion. PMID:26942079

  11. Cdh11 acts as a tumor suppressor in a murine retinoblastoma model by facilitating tumor cell death.

    Directory of Open Access Journals (Sweden)

    Mellone N Marchong

    2010-04-01

    Full Text Available CDH11 gene copy number and expression are frequently lost in human retinoblastomas and in retinoblastomas arising in TAg-RB mice. To determine the effect of Cdh11 loss in tumorigenesis, we crossed Cdh11 null mice with TAg-RB mice. Loss of Cdh11 had no gross morphological effect on the developing retina of Cdh11 knockout mice, but led to larger retinal volumes in mice crossed with TAg-RB mice (p = 0.01. Mice null for Cdh11 presented with fewer TAg-positive cells at postnatal day 8 (PND8 (p = 0.01 and had fewer multifocal tumors at PND28 (p = 0.016, compared to mice with normal Cdh11 alleles. However, tumor growth was faster in Cdh11-null mice between PND8 and PND84 (p = 0.003. In tumors of Cdh11-null mice, cell death was decreased 5- to 10-fold (p<0.03 for all markers, while proliferation in vivo remained unaffected (p = 0.121. Activated caspase-3 was significantly decreased and beta-catenin expression increased in Cdh11 knockdown experiments in vitro. These data suggest that Cdh11 displays tumor suppressor properties in vivo and in vitro in murine retinoblastoma through promotion of cell death.

  12. [Compound K suppresses myeloid-derived suppressor cells in a mouse model bearing CT26 colorectal cancer xenograft].

    Science.gov (United States)

    Wang, Rong; Li, Yalin; Wang, Wuzhou; Zhou, Meijuan; Cao, Zhaohui

    2015-05-01

    To investigate the effect of ginseng-derived compound K (C-K) on apoptosis, immunosuppressive activity, and pro-inflammatory cytokine production of myeloid-derived suppressor cells (MDSCs) from mice bearing colorectal cancer xenograft. Flow-sorted bone marrow MDSCs from Balb/c mice bearing CT26 tumor xenograft were treated with either C-K or PBS for 96 h and examined for apoptosis with Annexin V/7-AAD, Cox-2 and Arg-1 expressions using qRT-PCR, and supernatant IL-1β, IL-6, and IL-17 levels with ELISA. C-K- or PBS-treated MDSCs were subcutaneously implanted along with CT26 tumor cells in WT Balb/c mice, and the tumor size and morphology were evaluated 21 days later. C-K treatment significantly increased the percentages of early and late apoptotic MDSCs in vitro (Pimmunosuppresive effect of MDSCs to inhibit tumor cell proliferation in mice, which suggests a new strategy of tumor therapy by targeting MDSCs.

  13. Functional identification of an Arabidopsis snf4 ortholog by screening for heterologous multicopy suppressors of snf4 deficiency in yeast

    DEFF Research Database (Denmark)

    Kleinow, T.; Bhalerao, R.; Breuer, F.

    2000-01-01

    Yeast Snf4 is a prototype of activating gamma-subunits of conserved Snf1/AMPK-related protein kinases (SnRKs) controlling glucose and stress signaling in eukaryotes. The catalytic subunits of Arabidopsis SnRKs, AKIN10 and AKIN11, interact with Snf4 and suppress the snf1 and snf4 mutations in yeast....... By expression of an Arabidopsis cDNA library in yeast, heterologous multicopy snf4 suppressors were isolated. In addition to AKIN10 and AKIN11, the deficiency of yeast snf4 mutant to grown on non-fermentable carbon source was suppressed by Arabidopsis Myb30, CAAT-binding factor Hap3b, casein kinase I, zinc......-finger factors AZF2 and ZAT10, as well as orthologs of hexose/UDP-hexose transporters, calmodulin, SMC1-cohesin and Snf4. Here we describe the characterization of AtSNF4, a functional Arabidopsis Snf4 ortholog, that interacts with yeast Snf1 and specifically binds to the C-terminal regulatory domain...

  14. Expression of metastasis suppressor BRMS1 in breast cancer cells results in a marked delay in cellular adhesion to matrix.

    Science.gov (United States)

    Khotskaya, Yekaterina B; Beck, Benjamin H; Hurst, Douglas R; Han, Zhenbo; Xia, Weiya; Hung, Mien-Chie; Welch, Danny R

    2014-12-01

    Metastatic dissemination is a multi-step process that depends on cancer cells' ability to respond to microenvironmental cues by adapting adhesion abilities and undergoing cytoskeletal rearrangement. Breast Cancer Metastasis Suppressor 1 (BRMS1) affects several steps of the metastatic cascade: it decreases survival in circulation, increases susceptibility to anoikis, and reduces capacity to colonize secondary organs. In this report, BRMS1 expression is shown to not significantly alter expression levels of integrin monomers, while time-lapse and confocal microscopy revealed that BRMS1-expressing cells exhibited reduced activation of both β1 integrin and focal adhesion kinase, and decreased localization of these molecules to sites of focal adhesions. Short-term plating of BRMS1-expressing cells onto collagen or fibronectin markedly decreased cytoskeletal reorganization and formation of cellular adhesion projections. Under 3D culture conditions, BRMS1-expressing cells remained rounded and failed to reorganize their cytoskeleton and form invasive colonies. Taken together, BRMS1-expressing breast cancer cells are greatly attenuated in their ability to respond to microenvironment changes. © 2013 Wiley Periodicals, Inc. © 2013 Wiley Periodicals, Inc.

  15. NLRX1 Acts as an Epithelial-Intrinsic Tumor Suppressor through the Modulation of TNF-Mediated Proliferation

    Directory of Open Access Journals (Sweden)

    Ivan Tattoli

    2016-03-01

    Full Text Available The mitochondrial Nod-like receptor protein NLRX1 protects against colorectal tumorigenesis through mechanisms that remain unclear. Using mice with an intestinal epithelial cells (IEC-specific deletion of Nlrx1, we find that NLRX1 provides an IEC-intrinsic protection against colitis-associated carcinogenesis in the colon. These Nlrx1 mutant mice have increased expression of Tnf, Egf, and Tgfb1, three factors essential for wound healing, as well as increased epithelial proliferation during the epithelial regeneration phase following injury triggered by dextran sodium sulfate. In primary intestinal organoids lacking Nlrx1, stimulation with TNF resulted in exacerbated proliferation and expression of the intestinal stem cell markers Olfm4 and Myb. This hyper-proliferation response was associated with increased activation of Akt and NF-κB pathways in response to TNF stimulation. Together, these results identify NLRX1 as a suppressor of colonic tumorigenesis that acts by controlling epithelial proliferation in the intestine during the regeneration phase following mucosal injury.

  16. Senescence-Associated Secretory Phenotypes Reveal Cell-Nonautonomous Functions of Oncogenic RAS and the p53 Tumor Suppressor

    Energy Technology Data Exchange (ETDEWEB)

    Copp& #233; , Jean-Philippe; Patil, Christopher; Rodier, Francis; Sun, Yu; Munoz, Denise; Goldstein, Joshua; Nelson, Peter; Desprez, Pierre-Yves; Campisi, Judith

    2008-10-24

    Cellular senescence suppresses cancer by arresting cell proliferation, essentially permanently, in response to oncogenic stimuli, including genotoxic stress. We modified the use of antibody arrays to provide a quantitative assessment of factors secreted by senescent cells. We show that human cells induced to senesce by genotoxic stress secrete myriad factors associated with inflammation and malignancy. This senescence-associated secretory phenotype (SASP) developed slowly over several days and only after DNA damage of sufficient magnitude to induce senescence. Remarkably similar SASPs developed in normal fibroblasts, normal epithelial cells, and epithelial tumor cells after genotoxic stress in culture, and in epithelial tumor cells in vivo after treatment of prostate cancer patients with DNA-damaging chemotherapy. In cultured premalignant epithelial cells, SASPs induced an epithelial-mesenchyme transition and invasiveness, hallmarks of malignancy, by a paracrine mechanism that depended largely on the SASP factors interleukin (IL)-6 and IL-8. Strikingly, two manipulations markedly amplified, and accelerated development of, the SASPs: oncogenic RAS expression, which causes genotoxic stress and senescence in normal cells, and functional loss of the p53 tumor suppressor protein. Both loss of p53 and gain of oncogenic RAS also exacerbated the promalignant paracrine activities of the SASPs. Our findings define a central feature of genotoxic stress-induced senescence. Moreover, they suggest a cell-nonautonomous mechanism by which p53 can restrain, and oncogenic RAS can promote, the development of age-related cancer by altering the tissue microenvironment.

  17. Dysregulation of suppressor of cytokine signaling 3 in keratinocytes causes skin inflammation mediated by interleukin-20 receptor-related cytokines.

    Directory of Open Access Journals (Sweden)

    Ayako Uto-Konomi

    Full Text Available Homeostatic regulation of epidermal keratinocytes is controlled by the local cytokine milieu. However, a role for suppressor of cytokine signaling (SOCS, a negative feedback regulator of cytokine networks, in skin homeostasis remains unclear. Keratinocyte specific deletion of Socs3 (Socs3 cKO caused severe skin inflammation with hyper-production of IgE, epidermal hyperplasia, and S100A8/9 expression, although Socs1 deletion caused no inflammation. The inflamed skin showed constitutive STAT3 activation and up-regulation of IL-6 and IL-20 receptor (IL-20R related cytokines, IL-19, IL-20 and IL-24. Disease development was rescued by deletion of the Il6 gene, but not by the deletion of Il23, Il4r, or Rag1 genes. The expression of IL-6 in Socs3 cKO keratinocytes increased expression of IL-20R-related cytokines that further facilitated STAT3 hyperactivation, epidermal hyperplasia and neutrophilia. These results demonstrate that skin homeostasis is strictly regulated by the IL-6-STAT3-SOCS3 axis. Moreover, the SOCS3-mediated negative feedback loop in keratinocytes has a critical mechanistic role in the prevention of skin inflammation caused by hyperactivation of STAT3.

  18. Tumor suppressor BLU inhibits proliferation of nasopharyngeal carcinoma cells by regulation of cell cycle, c-Jun N-terminal kinase and the cyclin D1 promoter

    International Nuclear Information System (INIS)

    Zhang, Xiangning; Liu, Hui; Li, Binbin; Huang, Peichun; Shao, Jianyong; He, Zhiwei

    2012-01-01

    Tumor suppressor genes function to regulate and block tumor cell proliferation. To explore the mechanisms underlying the tumor suppression of BLU/ZMYND10 gene on a frequently lost human chromosomal region, an adenoviral vector with BLU cDNA insert was constructed. BLU was re-expressed in nasopharyngeal carcinoma cells by transfection or viral infection. Clonogenic growth was assayed; cell cycle was analyzed by flow cytometry-based DNA content detection; c-Jun N-terminal kinase (JNK) and cyclin D1 promoter activities were measured by reporter gene assay, and phosphorylation was measured by immunoblotting. The data for each pair of groups were compared with Student t tests. BLU inhibits clonogenic growth of nasopharyngeal carcinoma cells, arrests cell cycle at G1 phase, downregulates JNK and cyclin D1 promoter activities, and inhibits phosphorylation of c-Jun. BLU inhibits growth of nasopharyngeal carcinoma cells by regulation of the JNK-cyclin D1 axis to exert tumor suppression

  19. IL-1beta-induced chemokine and Fas expression are inhibited by suppressor of cytokine signalling-3 in insulin-producing cells

    DEFF Research Database (Denmark)

    Jacobsen, M L B; Rønn, S G; Bruun, C

    2008-01-01

    AIMS/HYPOTHESIS: Chemokines recruit activated immune cells to sites of inflammation and are important mediators of insulitis. Activation of the pro-apoptotic receptor Fas leads to apoptosis-mediated death of the Fas-expressing cell. The pro-inflammatory cytokines IL-1beta and IFN-gamma regulate...... the transcription of genes encoding the Fas receptor and several chemokines. We have previously shown that suppressor of cytokine signalling (SOCS)-3 inhibits IL-1beta- and IFN-gamma-induced nitric oxide production in a beta cell line. The aim of this study was to investigate whether SOCS-3 can influence cytokine......-induced Fas and chemokine expression in beta cells. METHODS: Using a beta cell line with inducible Socs3 expression or primary neonatal rat islet cells transduced with a Socs3-encoding adenovirus, we employed real-time RT-PCR analysis to investigate whether SOCS-3 affects cytokine-induced chemokine and Fas m...

  20. Suppressor cells in transplantation tolerance. I. Analysis of the suppressor status of neonatally and adoptively tolerized rats

    International Nuclear Information System (INIS)

    Dorsch, S.; Roser, B.

    1982-01-01

    The lymphocytes from neonatally tolerant rats which adoptively transfer tolerance to sublethally irradiated recipients do so by specifically suppressing the regeneration of alloreactivity which normally occurs after irradiation. Although tolerant cells will only partially suppress normal alloreactive cells when the two are mixed in near equivalent numbers, experiments in which the interval between injection of tolerant and normal cells into irradiated recipients was gradually extended, indicated that total suppression of normally alloreactive cells was achieved after 8 weeks of prior residence of tolerant cells in the adoptive host. Further evidence that tolerant cells would only suppress if present in excess of normal cells was obtained by reducing the tolerant cell population in tolerant donor rats by whole body irradiation. These animals then lost their ability to suppress normal alloreactive cells administered to them. The immune status of adoptively tolerized animals did not mimic that of the donors of the tolerant cells. Even where full tolerance, as measured by skin graft survival, failure to synthesize alloantibodies, and capacity to further transfer skin graft tolerance to secondary recipients, was evident the lymphocytes of these animals showed considerable graft-versus-host (GVH) reactivity. The persistence of tolerance through repeated adoptive transfers was correlated with the persistence of donor (chimeric) cells and the indicator skin graft on adoptive recipients only amplified tolerance expression where the inocula of tolerant cells given was weakly suppressive. Finally, removal of the minor population of chimeric cells from tolerant inocula using cytotoxic alloantisera abolished the capacity to transfer tolerance. These results imply an active role for chimeric cells which is best understood as an immune response involving proliferation driven by the idiotypes of the alloreceptors on host cells

  1. Regulation of the tumor suppressor FOXO3 by the thromboxane-A2 receptors in urothelial cancer.

    Directory of Open Access Journals (Sweden)

    Philip M Sobolesky

    Full Text Available The transcription factor FOXO3 is a well-established tumor suppressor whose activity, stability, and localization are regulated by phosphorylation and acetylation. Previous data by our laboratory demonstrated amplified thromboxane-A2 signaling was associated with poor prognoses in bladder cancer patients and overexpression of the thromboxane-A2 isoform-β receptor (TPβ, but not TPα, induced malignant transformation of immortalized bladder cells in vivo. Here, we describe a mechanism of TP mediated modulation of FOXO3 activity and localization by phosphorylation and deacetylation in a bladder cancer cell model. In vitro gain and loss of function studies performed in non-transformed cell lines, UROsta and SV-HUC, revealed knockdown of FOXO3 expression by shRNA increased cell migration and invasion, while exogenously overexpressing TPβ raised basal phosphorylated (pFOXO3-S294 levels. Conversely, overexpression of ERK-resistant, mutant FOXO3 reduced increases in UMUC3 cell migration and invasion, including that mediated by TP agonist (U46619. Additionally, stimulation of UMUC3 cells with U46619 increased pFOXO3-S294 expression, which could be attenuated by treatment with a TP antagonist (PTXA2 or ERK inhibitor (U0126. Initially U46619 caused nuclear accumulation of pFOXO3-S294; however, prolonged stimulation increased FOXO3 cytoplasmic localization. U46619 stimulation decreased overall FOXO3 transcriptional activity, but was associated with increased expression of its pro-survival target, manganese superoxide dismutase. The data also shows that TP stimulation increased the expression of the histone deacetylase, SIRT1, and corresponded with decreased acetylated-FOXO3. Collectively, the data suggest a role for TP signaling in the regulation of FOXO3 activity, mediated in part through phosphorylation and deacetylation.

  2. An innovative strategy to clone positive modifier genes of defects caused by mtDNA mutations: MRPS18C as suppressor gene of m.3946G>A mutation in MT-ND1 gene.

    Science.gov (United States)

    Rodríguez-García, María Elena; Cotrina-Vinagre, Francisco Javier; Carnicero-Rodríguez, Patricia; Martínez-Azorín, Francisco

    2017-07-01

    We have developed a new functional complementation approach to clone modifier genes which overexpression is able to suppress the biochemical defects caused by mtDNA mutations (suppressor genes). This strategy consists in transferring human genes into respiratory chain-deficient fibroblasts, followed by a metabolic selection in a highly selective medium. We used a normalized expression cDNA library in an episomal vector (pREP4) to transfect the fibroblasts, and a medium with glutamine and devoid of any carbohydrate source to select metabolically. Growing the patient's fibroblasts in this selective medium, the deficient cells rapidly disappear unless they are rescued by the cDNA of a suppressor gene. The use of an episomal vector allows us to carry out several rounds of transfection/selection (cyclical phenotypic rescue) to enrich the rescue with true clones of suppressor genes. Using fibroblasts from a patient with epileptic encephalopathy with the m.3946G>A (p.E214K) mutation in the MT-ND1 gene, several candidate genes were identified and one of them was characterized functionally. Thus, overexpression of MRPS18C gene (that encode for bS18m protein) suppressed the molecular defects produced by this mtDNA mutation, recovering the complex I activity and reducing the ROS produced by this complex to normal levels. We suggest that modulation of bS18m expression may be an effective therapeutic strategy for the patients with this mutation.

  3. Genetic and biochemical evidence that haploinsufficiency of the Nf1 tumor suppressor gene modulates melanocyte and mast cell fates in vivo.

    Science.gov (United States)

    Ingram, D A; Yang, F C; Travers, J B; Wenning, M J; Hiatt, K; New, S; Hood, A; Shannon, K; Williams, D A; Clapp, D W

    2000-01-03

    Neurofibromatosis type 1 (NF1) is a common autosomal-dominant disorder characterized by cutaneous neurofibromas infiltrated with large numbers of mast cells, melanocyte hyperplasia, and a predisposition to develop malignant neoplasms. NF1 encodes a GTPase activating protein (GAP) for Ras. Consistent with Knudson's "two hit" model of tumor suppressor genes, leukemias and malignant solid tumors in NF1 patients frequently demonstrate somatic loss of the normal NF1 allele. However, the phenotypic and biochemical consequences of heterozygous inactivation of Nf1 are largely unknown. Recently neurofibromin, the protein encoded by NF1, was shown to negatively regulate Ras activity in Nf1-/- murine myeloid hematopoietic cells in vitro through the c-kit receptor tyrosine kinase (dominant white spotting, W). Since the W and Nf1 locus appear to function along a common developmental pathway, we generated mice with mutations at both loci to examine potential interactions in vivo. Here, we show that haploinsufficiency at Nf1 perturbs cell fates in mast cells in vivo, and partially rescues coat color and mast cell defects in W(41) mice. Haploinsufficiency at Nf1 also increased mast cell proliferation, survival, and colony formation in response to Steel factor, the ligand for c-kit. Furthermore, haploinsufficiency was associated with enhanced Ras-mitogen-activated protein kinase activity, a major downstream effector of Ras, via wild-type and mutant (W(41)) c-kit receptors. These observations identify a novel interaction between c-kit and neurofibromin in vivo, and offer experimental evidence that haploinsufficiency of Nf1 alters both cellular and biochemical phenotypes in two cell lineages that are affected in individuals with NF1. Collectively, these data support the emerging concept that heterozygous inactivation of tumor suppressor genes may have profound biological effects in multiple cell types.

  4. Potential role of TRIM3 as a novel tumour suppressor in colorectal cancer (CRC) development.

    Science.gov (United States)

    Piao, Mei-Yu; Cao, Hai-Long; He, Na-Na; Xu, Meng-Que; Dong, Wen-Xiao; Wang, Wei-Qiang; Wang, Bang-Mao; Zhou, Bing

    2016-01-01

    Colorectal cancer (CRC) is the third leading cause of cancer-related mortality in the United States. Recent cancer genome-sequencing efforts and complementary functional studies have led to the identification of a collection of candidate 'driver' genes involved in CRC tumorigenesis. Tripartite motif (TRIM3) is recently identified as a tumour suppressor in glioblastoma but this tumour-suppressive function has not been investigated in CRC. In this study, we investigated the potential role of TRIM3 as a tumour suppressor in CRC development by manipulating the expression of TRIM3 in two authentic CRC cell lines, HCT116 and DLD1, followed by various functional assays, including cell proliferation, colony formation, scratch wound healing, soft agar, and invasion assays. Xenograft experiment was performed to examine in vivo tumour-suppressive properties of TRIM3. Small-interfering RNA (siRNA) mediated knockdown of TRIM3 conferred growth advantage in CRC cells. In contrast, overexpression of TRIM3 affected cell survival, cell migration, anchorage independent growth and invasive potential in CRC cells. In addition, TRIM3 was found to be down-regulated in human colon cancer tissues compared with matched normal colon tissues. Overexpression of TRIM3 significantly inhibited tumour growth in vivo using xenograft mouse models. Mechanistic investigation revealed that TRIM3 can regulate p53 protein level through its stabilisation. TRIM3 functions as a tumour suppressor in CRC progression. This tumour-suppressive function is exerted partially through regulation of p53 protein. Therefore, this protein may represent a novel therapeutic target for prevention or intervention of CRC.

  5. Syk Tyrosine Kinase Acts as a Pancreatic Adenocarcinoma Tumor Suppressor by Regulating Cellular Growth and Invasion

    OpenAIRE

    Layton, Tracy; Stalens, Cristel; Gunderson, Felizza; Goodison, Steve; Silletti, Steve

    2009-01-01

    We have identified the nonreceptor tyrosine kinase syk as a marker of differentiation/tumor suppressor in pancreatic ductal adenocarcinoma (PDAC). Syk expression is lost in poorly differentiated PDAC cells in vitro and in situ, and stable reexpression of syk in endogenously syk-negative Panc1 (Panc1/syk) cells retarded their growth in vitro and in vivo and reduced anchorage-independent growth in vitro. Panc1/syk cells exhibited a more differentiated morphology and down-regulated cyclin D1, ak...

  6. Purification of SOCS (Suppressor of Cytokine Signaling) SH2 Domains for Structural and Functional Studies.

    Science.gov (United States)

    Liau, Nicholas P D; Laktyushin, Artem; Babon, Jeffrey J

    2017-01-01

    Src Homology 2 (SH2) domains are protein domains which have a high binding affinity for specific amino acid sequences containing a phosphorylated tyrosine residue. The Suppressors of Cytokine Signaling (SOCS) proteins use an SH2 domain to bind to components of certain cytokine signaling pathways to downregulate the signaling cascade. The recombinantly produced SH2 domains of various SOCS proteins have been used to undertake structural and functional studies elucidating the method of how such targeting occurs. Here, we describe the protocol for the recombinant production and purification of SOCS SH2 domains, with an emphasis on SOCS3.

  7. The Role of Tumor Metastases Suppressor Gene, Drg-1, in Breast Cancer

    Science.gov (United States)

    2008-03-01

    evidence to validate 14 our data of breast cancer. However, these prostate cells and reagents were existing materials in our lab or purchased by using...J. Lab . Clin. Med. 133, 265–273. Sloane, B.F., Honn, K.V., 1984. Cysteine proteinases and metastasis. Cancer Metastasis Rev. 3, 249–263. Sridhar, S.C... Beest , P. Moerer, K. van der Horn, R. Goldschmeding, T. Logtenberg and H. Clevers: Synergy between tumor suppressor APC and the beta- catenin-Tcf4

  8. Expression of the p16{sup INK4a} tumor suppressor gene in rodent lung tumors

    Energy Technology Data Exchange (ETDEWEB)

    Swafford, D.S.; Tesfaigzi, J.; Belinsky, S.A.

    1995-12-01

    Aberrations on the short arm of chromosome 9 are among the earliest genetic changes in human cancer. p16{sup INK4a} is a candidate tumor suppressor gene that lies within human 9p21, a chromosome region associated with frequent loss of heterozygosity in human lung tumors. The p16{sup INK4a} protein functions as an inhibitor of cyclin D{sub 1}-dependent kinases that phosphorylate the retinoblastoma (Rb) tumor suppressor gene product enabling cell-cycle progression. Thus, overexpression of cyclin D{sub 1}, mutation of cyclin-dependent kinase genes, or loss of p16{sup INK4a} function, can all result in functional inactivation of Rb. Inactivation of Rb by mutation or deletion can result in an increase in p16{sup INK4a} transcription, suggesting that an increased p16{sup INK4a} expression in a tumor cell signals dysfunction of the pathway. The p16{sup (INK4a)} gene, unlike some tumor suppressor genes, is rarely inactivated by mutation. Instead, the expression of this gene is suppressed in some human cancers by hypermethylation of the CpG island within the first exon or by homozygous deletion: 686. Chromosome losses have been observed at 9p21 syntenic loci in tumors of the mouse and rat, two species often used as animal models for pulmonary carcinogenesis. Expression of p16{sup INK4a} is lost in some mouse tumor cell lines, often due to homozygous deletion. These observations indicate that p16{sup INK4a} dysfunction may play a role in the development of neoplasia in rodents as well as humans. The purpose of the current investigation was to define the extent to which p16{sup INK4a} dysfunction contributes to the development of rodent lung tumors and to determine the mechanism of inactivation of the gene. There is no evidence to suggest a loss of function of the p16{sup INK4a} tumor suppressor gene in these primary murine lung tumors by mutation, deletion, or methylation.

  9. P18 tumor suppressor gene and progression of oligodendrogliomas to anaplasia.

    Science.gov (United States)

    He, J; Hoang-Xuan, K; Marie, Y; Leuraud, P; Mokhtari, K; Kujas, M; Delattre, J Y; Sanson, M

    2000-09-26

    P18INK4C is a good candidate to be the tumor suppressor gene involved in oligodendrogliomas on 1p32. Loss of heterozygosity on 1p, mutation(s), homozygous deletion(s), and expression of p18 in 30 oligodendroglial tumors were investigated. Loss of heterozygosity on 1p was found in 15 tumors. A p18 mutation was found at an recurrence of an anaplastic oligodendroglioma, but not in the primary, low-grade tumor. No homozygous deletions were found and p18 was expressed in all cases. These results show that p18 alteration is involved in tumor progression in a subset of oligodendrogliomas.

  10. Inhibitor of differentiation 4 (Id4) is a potential tumor suppressor in prostate cancer

    International Nuclear Information System (INIS)

    Carey, Jason PW; Asirvatham, Ananthi J; Galm, Oliver; Ghogomu, Tandeih A; Chaudhary, Jaideep

    2009-01-01

    Inhibitor of differentiation 4 (Id4), a member of the Id gene family is also a dominant negative regulator of basic helix loop helix (bHLH) transcription factors. Some of the functions of Id4 appear to be unique as compared to its other family members Id1, Id2 and Id3. Loss of Id4 gene expression in many cancers in association with promoter hypermethylation has led to the proposal that Id4 may act as a tumor suppressor. In this study we provide functional evidence that Id4 indeed acts as a tumor suppressor and is part of a cancer associated epigenetic re-programming. Data mining was used to demonstrate Id4 expression in prostate cancer. Methylation specific polymerase chain reaction (MSP) analysis was performed to understand molecular mechanisms associated with Id4 expression in prostate cancer cell lines. The effect of ectopic Id4 expression in DU145 cells was determined by cell cycle analysis (3H thymidine incorporation and FACS), expression of androgen receptor, p53 and cyclin dependent kinase inhibitors p27 and p21 by a combination of RT-PCR, real time-PCR, western blot and immuno-cytochemical analysis. Id4 expression was down-regulated in prostate cancer. Id4 expression was also down-regulated in prostate cancer line DU145 due to promoter hyper-methylation. Ectopic Id4 expression in DU145 prostate cancer cell line led to increased apoptosis and decreased cell proliferation due in part by an S-phase arrest. In addition to S-phase arrest, ectopic Id4 expression in PC3 cells also resulted in prolonged G2/M phase. At the molecular level these changes were associated with increased androgen receptor (AR), p21, p27 and p53 expression in DU145 cells. The results suggest that Id4 acts directly as a tumor suppressor by influencing a hierarchy of cellular processes at multiple levels that leads to a decreased cell proliferation and change in morphology that is possibly mediated through induction of previously silenced tumor suppressors

  11. Inhibitor of differentiation 4 (Id4 is a potential tumor suppressor in prostate cancer

    Directory of Open Access Journals (Sweden)

    Carey Jason PW

    2009-06-01

    Full Text Available Abstract Background Inhibitor of differentiation 4 (Id4, a member of the Id gene family is also a dominant negative regulator of basic helix loop helix (bHLH transcription factors. Some of the functions of Id4 appear to be unique as compared to its other family members Id1, Id2 and Id3. Loss of Id4 gene expression in many cancers in association with promoter hypermethylation has led to the proposal that Id4 may act as a tumor suppressor. In this study we provide functional evidence that Id4 indeed acts as a tumor suppressor and is part of a cancer associated epigenetic re-programming. Methods Data mining was used to demonstrate Id4 expression in prostate cancer. Methylation specific polymerase chain reaction (MSP analysis was performed to understand molecular mechanisms associated with Id4 expression in prostate cancer cell lines. The effect of ectopic Id4 expression in DU145 cells was determined by cell cycle analysis (3H thymidine incorporation and FACS, expression of androgen receptor, p53 and cyclin dependent kinase inhibitors p27 and p21 by a combination of RT-PCR, real time-PCR, western blot and immuno-cytochemical analysis. Results Id4 expression was down-regulated in prostate cancer. Id4 expression was also down-regulated in prostate cancer line DU145 due to promoter hyper-methylation. Ectopic Id4 expression in DU145 prostate cancer cell line led to increased apoptosis and decreased cell proliferation due in part by an S-phase arrest. In addition to S-phase arrest, ectopic Id4 expression in PC3 cells also resulted in prolonged G2/M phase. At the molecular level these changes were associated with increased androgen receptor (AR, p21, p27 and p53 expression in DU145 cells. Conclusion The results suggest that Id4 acts directly as a tumor suppressor by influencing a hierarchy of cellular processes at multiple levels that leads to a decreased cell proliferation and change in morphology that is possibly mediated through induction of previously

  12. Repression of Akt3 gene transcription by the tumor suppressor RIZ1

    OpenAIRE

    Liu, Qingnan; Qu, Xiaotian; Xie, Xiaolei; He, Pei; Huang, Shi

    2018-01-01

    RIZ1 has been studied as a tumor suppressor and may play a role in metabolic diseases related to the Western style diet, such as cancer and obesity. The Akt pathway is known to play a role in both cancer and obesity, and a link between Akt and RIZ1 has also been found. To better understand the role of RIZ1 in obesity and cancer, we investigated how RIZ1 regulates the expression of Akt3. We found that overexpression of RIZ1 in HEK293 cells reduced the expression of Akt3 protein. Luciferase rep...

  13. Role of natural antisense transcripts pertaining to tumor suppressor genes in human carcinomas

    International Nuclear Information System (INIS)

    Pelicci, G.; Pierotti, M.

    2009-01-01

    Overlapping transcripts in opposite orientations can potentially form perfect sense-antisense duplex RNA. Recently, several studies have revealed the extent of natural antisense transcripts (NATs) and their role in important biological phenomena also in higher organisms. In order to test the hypothesis that the function of NATs in man might represent an essential element in the regulation of gene expression, especially at transcriptional level, in this study we planned to look for, systematically examine, and characterize NATs belonging in the human genome to the tumour suppressor class of genes, so to identify physiological (and potentially pathological) modulators in this gene class

  14. Estrogen receptor beta, a possible tumor suppressor involved in ovarian carcinogenesis

    Science.gov (United States)

    Lazennec, Gwendal

    2006-01-01

    Ovarian cancer is one of the leading cause of death from gynecological tumors in women. Several lines of evidence suggest that estrogens may play an important role in ovarian carcinogenesis, through their receptors, ERα and ERβ. Interestingly, malignant ovarian tumors originating from epithelial surface constitute about 90% of ovarian cancers and expressed low levels of ERβ, compared to normal tissues. In addition, restoration of ERβ in ovarian cancer cells, leads to strong inhibition of their proliferation and invasion, while apoptosis is enhanced. In this manuscript, recent data suggesting a possible tumor-suppressor role for ERβ in ovarian carcinogenesis are discussed. PMID:16399219

  15. Molecular studies on the function of tumor suppressor gene in gastrointestinal cancer

    International Nuclear Information System (INIS)

    Kim, You Cheoul

    1993-01-01

    Cancer of stomach, colon and liver are a group of the most common cancer in Korea. However, results with current therapeutic modalities are still unsatisfactory. The intensive efforts have been made to understand basic pathogenesis and to find better therapeutic tools for the treatment of this miserable disease. We studies the alteration of tumor suppressor gene in various Gastrointestinal cancer in Korea. Results showed that genetic alteration of Rb gene was in 83% of colorectal cancer. Our results suggest that genetic alteration of Rb gene is crucially involved in the tumorigenesis of colorectum in Korea. (Author)

  16. HLA-DR-specific suppressor cells after repeated allogeneic sensitizations of human lymphocytes in vitro

    International Nuclear Information System (INIS)

    Sasportes, M.; Fradelizi, D.; Dausset, J.

    1978-01-01

    In conclusion, DR-specific suppressor cells can be induced by repeated in vitro sensitizations. They were able to decrease a secondary proliferation, to suppress consistently, in a primary proliferative assay, when added as third cells (primed twice against a DR antigen [PLT II] and γ-irradiated), the response of unprimed cells towards stimulating cells, which share a DR specificity with the priming cell of the PLT II. The suppression follows the D part of the recombinant haplotype within an HLA-B/D recombinant family and is specific for the DR antigen used twice as stimulator for production of the PLT II

  17. Indomethacin sensitive suppressor cell activity in head and neck cancer patients pre- and postirradiation therapy

    International Nuclear Information System (INIS)

    Maca, R.D.; Panje, W.R.

    1982-01-01

    The effects of the addition of indomethacin to PHA or Con A stimulated lymphocytes from patients with untreated squamous cell carcinoma of the head and neck or from patients with the disease who have just finished irradiation therapy from the disease was quantitated and compared to those of the control group. Lymphocytes from eight of 26 patients with untreated carcinoma were significantly augmented by the addition of indomethacin. The remaining eighteen patients were equal to the controls. For all 17 patients who had just finished extensive field irradiation therapy, significant enhancement of PHA and Con A reactivity by indomethacin was found, which did not appear to be solely a function of low baseline mitogen reactivity. The results of this study support the hypothesis that perhaps patients receiving irradiation therapy may benefit by the oral administration of indomethacin, an approach that needs further consideration

  18. Tumor Suppressor Activity of the EphB2 Receptor in Prostate Cancer

    Science.gov (United States)

    2008-11-01

    H. (2006) Cancer Sci. 97, 1211–1216 34. Yao, V. J., Ozawa, M. G., Trepel, M., Arap, W., McDonald, D. M., and Pasqualini , R. (2005) Am. J. Pathol...and Lemay, S. (2006) Am. J. Physiol. 291, F960–F971 72. Ogawa, K., Pasqualini , R., Lindberg, R. A., Kain, R., Freeman, A. L., and Pasquale, E. B...Science 117: 2037-2049. 21. Ogawa K Pasqualini R Lindberg RA et al. (2000) The ephrin-A1 ligand and its receptor, EphA2, are expressed during tumor

  19. Evaluating the silencing suppressor activity of proteins encoded by maize rayado fino virus

    Science.gov (United States)

    Maize rayado fino virus (MRFV), the type member of the genus Marafivirus, family Tymoviridae, is transmitted in a persistent, circulative manner by leafhoppers of the genus Dalbulus. Symptoms of MRFV infection on leaves of its maize host are small chlorotic spots that coalesce into short stripes. T...

  20. Tumor suppressor function of Bruton tyrosine kinase is independent of its catalytic activity

    NARCIS (Netherlands)

    S. Middendorp; A.J.E. Zijlstra (Esther); R. Kersseboom (Rogier); G.M. Dingjan (Gemma); H. Jumaa; R.W. Hendriks (Rudi)

    2005-01-01

    textabstractDuring B-cell development in the mouse, Bruton tyrosine kinase (Btk) and the adaptor protein SLP-65 (Src homology 2 [SH2] domain-containing leukocyte protein of 65 kDa) limit the expansion and promote the differentiation of pre-B cells. Btk is thought to mainly function

  1. LncRNA TUG1 acts as a tumor suppressor in human glioma by promoting cell apoptosis.

    Science.gov (United States)

    Li, Jun; Zhang, Meng; An, Gang; Ma, Qingfang

    2016-03-01

    Previous studies have revealed multiple functional roles of long non-coding RNA taurine upregulated gene 1 in different types of malignant tumors, except for human glioma. Here, it was designed to study the potential function of taurine upregulated gene 1 in glioma pathogenesis focusing on its regulation on cell apoptosis. The expression of taurine upregulated gene 1 in glioma tissues was detected by quantitative RT-PCR and compared with that in adjacent normal tissues. Further correlation analysis was conducted to show the relationship between taurine upregulated gene 1 expression and different clinicopathologic parameters. Functional studies were performed to investigate the influence of taurine upregulated gene 1 on apoptosis and cell proliferation by using Annexin V/PI staining and cell counting kit-8 assays, respectively. And, caspase activation and Bcl-2 expression were analyzed to explore taurine upregulated gene 1-induced mechanism. taurine upregulated gene 1 expression was significantly inhibited in glioma and showed significant correlation with WHO Grade, tumor size and overall survival. Further experiments revealed that the dysregulation of taurine upregulated gene 1 affected the apoptosis and cell proliferation of glioma cells. Moreover, taurine upregulated gene 1 could induce the activation of caspase-3 and-9, with inhibited expression of Bcl-2, implying the mechanism in taurine upregulated gene 1-induced apoptosis. taurine upregulated gene 1 promoted cell apoptosis of glioma cells by activating caspase-3 and -9-mediated intrinsic pathways and inhibiting Bcl-2-mediated anti-apoptotic pathways, acting as a tumor suppressor in human glioma. This study provided new insights for the function of taurine upregulated gene 1 in cancer biology, and suggested a potent application of taurine upregulated gene 1 overexpression for glioma therapy. © 2016 by the Society for Experimental Biology and Medicine.

  2. Zyxin regulates migration of renal epithelial cells through activation of hepatocyte nuclear factor-1β.

    Science.gov (United States)

    Choi, Yun-Hee; McNally, Brian T; Igarashi, Peter

    2013-07-01

    Hepatocyte nuclear factor-1β (HNF-1β) is an epithelial tissue-specific transcription factor that regulates gene expression in the kidney, liver, pancreas, intestine, and other organs. Mutations of HNF-1β in humans produce renal cysts and congenital kidney anomalies. Here, we identify the LIM-domain protein zyxin as a novel binding partner of HNF-1β in renal epithelial cells. Zyxin shuttles to the nucleus where it colocalizes with HNF-1β. Immunoprecipitation of zyxin in leptomycin B-treated cells results in coprecipitation of HNF-1β. The protein interaction requires the second LIM domain of zyxin and two distinct domains of HNF-1β. Overexpression of zyxin stimulates the transcriptional activity of HNF-1β, whereas small interfering RNA silencing of zyxin inhibits HNF-1β-dependent transcription. Epidermal growth factor (EGF) induces translocation of zyxin into the nucleus and stimulates HNF-1β-dependent promoter activity. The EGF-mediated nuclear translocation of zyxin requires activation of Akt. Expression of dominant-negative mutant HNF-1β, knockdown of zyxin, or inhibition of Akt inhibits EGF-stimulated cell migration. These findings reveal a novel pathway by which extracellular signals are transmitted to the nucleus to regulate the activity of a transcription factor that is essential for renal epithelial differentiation.

  3. Axial-Symmetry Numerical Approaches for Noise Predicting and Attenuating of Rifle Shooting with Suppressors

    Directory of Open Access Journals (Sweden)

    Shi-Wei Lo

    2011-01-01

    Full Text Available The moving bullet out of a rifle barrel is propelled by a fired explosive charge. Subsequently, a disturbed muzzle blast wave is initiated which lasts several milliseconds. In this study, axially symmetric, unsteady, Large Eddy Simulation (LES, and Ffowcs Williams and Hawkins (FWH equations were solved by the implicit-time formulation. For the spatial discretization, second order upwind scheme was employed. In addition, dynamic mesh model was used to where the ballistic domain changed with time due to the motion of bullet. Results obtained for muzzle flow field and for noise recorded were compared with those obtained from experimental data; these two batches of results were in agreement. Five cases of gunshot including one model of an unsuppressed rifle and four models of suppressors were simulated. Besides, serial images of species distributions and velocity vectors-pressure contours in suppressors and near muzzle field were displayed. The sound pressure levels (dB in far field that were post-processed by the fast Fourier transform (FFT were compared. The proposed physical model and the numerical simulations used in the present work are expected to be extended to solve other shooting weapon problems with three-dimensional and complex geometries.

  4. Polysaccharide from Lentinus edodes inhibits the immunosuppressive function of myeloid-derived suppressor cells.

    Directory of Open Access Journals (Sweden)

    Hao Wu

    Full Text Available Reversing the function of immune suppressor cells may improve the efficacy of cancer therapy. Here, we have isolated a novel polysaccharide MPSSS (577.2 Kd from Lentinus edodes and examined its effects on differentiation and function of myeloid-derived suppressor cells (MDSCs. MPSSS is composed of glucose (75.0%, galactose (11.7%, mannose (7.8%, and xylose (0.4%. In vivo, it inhibits the growth of McgR32 tumor cells, which is correlated with a reduced percentage of MDSCs in peripheral blood. In vitro, it induces both morphological and biophysical changes in MDSCs. Importantly, MPSSS up-regulates MHC II and F4/80 expression on MDSCs, and reverses their inhibition effect on CD4(+ T cells in a dose-dependent manner. The mechanism study shows that MPSSS may stimulate MDSCs through a MyD88 dependent NF-κB signaling pathway. Together, we demonstrated for the first time that MPSSS stimulates the differentiation of MDSCs and reverses its immunosuppressive functions, shedding new light on developing novel anti-cancer strategies by targeting MDSCs.

  5. Non-Aqueous Titration Method for Determining Suppressor Concentration in the MCU Next Generation Solvent (NGS)

    Energy Technology Data Exchange (ETDEWEB)

    Taylor-Pashow, Kathryn M. L. [Savannah River Site (SRS), Aiken, SC (United States). Savannah River National Lab. (SRNL); Jones, Daniel H. [Savannah River Site (SRS), Aiken, SC (United States). Savannah River National Lab. (SRNL)

    2017-10-23

    A non-aqueous titration method has been used for quantifying the suppressor concentration in the MCU solvent hold tank (SHT) monthly samples since the Next Generation Solvent (NGS) was implemented in 2013. The titration method measures the concentration of the NGS suppressor (TiDG) as well as the residual tri-n-octylamine (TOA) that is a carryover from the previous solvent. As the TOA concentration has decreased over time, it has become difficult to resolve the TiDG equivalence point as the TOA equivalence point has moved closer. In recent samples, the TiDG equivalence point could not be resolved, and therefore, the TiDG concentration was determined by subtracting the TOA concentration as measured by semi-volatile organic analysis (SVOA) from the total base concentration as measured by titration. In order to improve the titration method so that the TiDG concentration can be measured directly, without the need for the SVOA data, a new method has been developed that involves spiking of the sample with additional TOA to further separate the two equivalence points in the titration. This method has been demonstrated on four recent SHT samples and comparison to results obtained using the SVOA TOA subtraction method shows good agreement. Therefore, it is recommended that the titration procedure be revised to include the TOA spike addition, and this to become the primary method for quantifying the TiDG.

  6. Non-Aqueous Titration Method for Determining Suppressor Concentration in the MCU Next Generation Solvent (NGS)

    International Nuclear Information System (INIS)

    Taylor-Pashow, Kathryn M. L.; Jones, Daniel H.

    2017-01-01

    A non-aqueous titration method has been used for quantifying the suppressor concentration in the MCU solvent hold tank (SHT) monthly samples since the Next Generation Solvent (NGS) was implemented in 2013. The titration method measures the concentration of the NGS suppressor (TiDG) as well as the residual tri-n-octylamine (TOA) that is a carryover from the previous solvent. As the TOA concentration has decreased over time, it has become difficult to resolve the TiDG equivalence point as the TOA equivalence point has moved closer. In recent samples, the TiDG equivalence point could not be resolved, and therefore, the TiDG concentration was determined by subtracting the TOA concentration as measured by semi-volatile organic analysis (SVOA) from the total base concentration as measured by titration. In order to improve the titration method so that the TiDG concentration can be measured directly, without the need for the SVOA data, a new method has been developed that involves spiking of the sample with additional TOA to further separate the two equivalence points in the titration. This method has been demonstrated on four recent SHT samples and comparison to results obtained using the SVOA TOA subtraction method shows good agreement. Therefore, it is recommended that the titration procedure be revised to include the TOA spike addition, and this to become the primary method for quantifying the TiDG.

  7. Sleep quality and methylation status of selected tumor suppressor genes among nurses and midwives.

    Science.gov (United States)

    Bukowska-Damska, Agnieszka; Reszka, Edyta; Kaluzny, Pawel; Wieczorek, Edyta; Przybek, Monika; Zienolddiny, Shanbeh; Peplonska, Beata

    2018-01-01

    Chronic sleep restriction may affect metabolism, hormone secretion patterns and inflammatory responses. Limited reports suggest also epigenetic effects, such as changes in DNA methylation profiles. The study aims to assess the potential association between poor sleep quality or sleep duration and the levels of 5-methylcytosine in the promoter regions of selected tumor suppressor genes. A cross-sectional study was conducted on 710 nurses and midwives aged 40-60 years. Data from interviews regarding sleep habits and potential confounders were used. The methylation status of tumor suppressor genes was determined via qMSP reactions using DNA samples derived from leucocytes. No significant findings were observed in the total study population or in the two subgroups of women stratified by the current system of work. A borderline significance association was observed between a shorter duration of sleep and an increased methylation level in CDKN2A among day working nurses and midwives. Further studies are warranted to explore this under-investigated topic.

  8. Tumor Suppressor RARRES1 Regulates DLG2, PP2A, VCP, EB1, and Ankrd26

    Directory of Open Access Journals (Sweden)

    Ziad J. Sahab, Michael D. Hall, Lihua Zhang, Amrita K. Cheema, Stephen W. Byers

    2010-01-01

    Full Text Available Retinoic Acid Receptor Responder (RARRES1 initially identified as a novel retinoic acid receptor regulated gene in the skin is a putative tumor suppressor of unknown function. RARRES1 was knocked down in immortalized human prostatic epithelial cell line PWR-1E cells and differential protein expression was identified using differential in-gel electrophoresis (DIGE followed by matrix-assisted laser desorption ionization (MALDI mass spectrometry and western Blot analysis excluding highly abundant proteins routinely identified in almost all proteomics projects. Knock-down of RARRES1: 1- down-regulates PP2A, an enzyme involved in the negative regulation of the growth hormone-stimulated signal transduction pathways; 2- down-regulates Valosin-containing protein causing impaired autophagy; 3- up-regulates the tumor suppressor disks large 2; 4- up-regulates Ankrd26 that belongs to the POTE family of genes that are highly expressed in cancer patients with poor outcome; and 5- down-regulates EB1, a protein that is involved in spindle dynamics and chromosome alignment during mitosis.

  9. Polymorphism of the p53 tumor suppressor gene is associated with susceptibility to uterine leiomyoma.

    Science.gov (United States)

    Denschlag, Dominik; Bettendorf, Herta; Watermann, Dirk; Keck, Christoph; Tempfer, Clemens; Pietrowski, Detlef

    2005-07-01

    To evaluate the association between the presence of uterine leiomyoma and two single nuclear polymorphisms of the p53 tumor suppressor and the angiopoietin-2 (ANGPT2) genes. Prospective case control study. Academic research institution. One hundred thirty-two women with clinically and surgically diagnosed uterine leiomyomas and 280 controls. Peripheral venous puncture. Genotyping was performed by polymerase chain reaction-based amplification of the Arg and Pro variants at codon 72 of the p53 gene and by restriction fragment length polymorphism analysis of the G/G and G/A alleles in exon 4 of the ANGPT2 gene. Comparing women with uterine leiomyomas and controls, no statistically significant difference with respect to allele frequency and genotype distribution were ascertained for the ANGPT2 polymorphism (P=.2 and P=.5, respectively). However, for the p53 tumor suppressor gene polymorphism, statistically significant differences in terms of a higher Pro allele frequency and a higher prevalence of the Pro/Pro genotype among women with uterine leiomyoma (32.0% vs. 16.0%, respectively, and 21.3% vs. 4.7%, respectively) were ascertained (P=.001, OR 1.74; 95% CI 1.24-2.45, P=.001; OR 3.84, 95% CI 1.81-8.14; respectively). Carriage of the p53 polymorphism at codon 72 predicts the susceptibility to leiomyoma in a Caucasian population and may contribute to the pathogenesis of uterine leiomyoma.

  10. Immunopurification of the suppressor tRNA dependent rabbit β-globin readthrough protein

    International Nuclear Information System (INIS)

    Hatfield, D.; Thorgeirsson, S.S.; Copeland, T.D.; Oroszlan, S.; Bustin, M.

    1988-01-01

    In mammalian cells, the rabbit β-globin readthrough protein is the only known example of a naturally occurring readthrough protein which does not involve a viral system. To provide an efficient means for its isolation, detection, and study, the authors elicited specific antibodies against this unique protein. The 22 amino acid peptide corresponding to the readthrough portion of this protein was synthesized, coupled to keyhole limpet hemocyanin, and injected into sheep. Specific antibodies to the peptide were produced as demonstrated by the enzyme-linked immunosorbent assay technique and by immunoblotting. The antibodies did not react with globin. The rabbit β-globin readthrough protein was separated from globin and other reticulocyte proteins by polyacrylamide gel electrophoresis and visualized by silver staining or by labeling with [ 35 S] methionine. Incorporation of [ 35 S] methionine into the readthrough protein was significantly enhanced upon addition of an opal suppressor tRNA to reticulocyte lysates. Immunoblotting revealed that the readthrough protein also occurs in lysates without added suppressor tRNA. The antibodies were purified on an affi-gel column which had been coupled with the peptide antigen. The readthrough protein was then purified from reticulocytes by immunoaffinity chromatography and by high-performance liquid chromatography. The results provide conclusive evidence that the β-globin readthrough protein is naturally occurring in rabbit reticulocytes

  11. Mapping the glaucousness suppressor Iw1 from wild emmer wheat “PI 481521”

    Institute of Scientific and Technical Information of China (English)

    Zongchang; Xu; Cuiling; Yuan; Jirui; Wang; Daolin; Fu; Jiajie; Wu

    2015-01-01

    Many species of Triticeae display a glaucous phenotype. In wheat, glaucousness/waxiness on spikes, leaves and shoots is controlled by wax production genes(W loci) and epistatic inhibitors(Iw loci). In this study, a suppressor of glaucousness from wild emmer wheat(Triticum turgidum ssp. dicoccoides) accession "PI 481521" was investigated in a pair of durum(T. turgidum ssp. durum cv. "Langdon", LDN)—wild emmer wheat chromosome substitution lines, LDN and "LDNDIC521-2B". Genetic analysis revealed that the non-glaucous phenotype of LDNDIC521-2Bwas controlled by the dominant glaucous suppressor Iw1 on the short arm of chromosome 2B. In total, 371 2B-specific marker differences were identified between LDN and LDNDIC521-2B. The location of the Iw1 gene was mapped using an F2 population that stemmed from LDN and LDNDIC521-2B, generating a partial linkage map that included 19 simple sequence repeats(SSR) and ten gene-based markers. On the current map, the Iw1 gene was located within the Xgwm614–BE498111 interval, and cosegregated with BQ788707,CD893659, CD927782, CD938589, and Xbarc35. Mapping of Iw1 in LDNDIC521-2B, a publically accessible and widely distributed line, will provide valuable information for marker-assisted selection of the agronomically important trait of glaucousness.

  12. Induction of CD4 suppressor T cells with anti-Leu-8 antibody

    International Nuclear Information System (INIS)

    Kanof, M.E.; Strober, W.; James, S.P.

    1987-01-01

    To characterize the conditions under which CD4 T cells suppress polyclonal immunoglobulin synthesis, we investigated the capacity of CD4 T cells that coexpress the surface antigen recognized by the monoclonal antibody anti-Leu-8 to mediate suppression. In an in vitro system devoid of CD8 T cells, CD4, Leu-8+ T cells suppressed pokeweed mitogen-induced immunoglobulin synthesis. Similarly, suppressor function was induced in unfractionated CD4 T cell populations after incubation with anti-Leu-8 antibody under cross-linking conditions. This induction of suppressor function by anti-Leu-8 antibody was not due to expansion of the CD4, Leu-8+ T cell population because CD4 T cells did not proliferate in response to anti-Leu-8 antibody. However, CD4, Leu-8+ T cell-mediated suppression was radiosensitive. Finally, CD4, Leu-8+ T cells do not inhibit immunoglobulin synthesis when T cell lymphokines were used in place of helper CD4 T cells (CD4, Leu-8- T cells), suggesting that CD4 T cell-mediated suppression occurs at the T cell level. We conclude that CD4 T cells can be induced to suppress immunoglobulin synthesis by modulation of the membrane antigen recognized by anti-Leu-8 antibody

  13. Tumor suppressor genes are frequently methylated in lymph node metastases of breast cancers

    Directory of Open Access Journals (Sweden)

    Xu Jia

    2010-07-01

    Full Text Available Abstract Introduction Metastasis represents a major adverse step in the progression of breast carcinoma. Lymph node invasion is the most relevant prognostic factor; however little is known on the molecular events associated with lymph node metastasis process. This study is to investigate the status and role of methylation in lymph node metastatic tumors. Materials and methods Bisulfite pyrosequencing is used to screen 6 putative tumor suppressor genes (HIN-1, RASSF1A, RIL, CDH13, RARβ2 and E-cadherin in 38 pairs of primary breast tumors and lymph node metastases. Results We found that HIN-1, CDH13, RIL, RASSF1A and RARβ2 were frequently methylated both in primary and metastatic tissues (range: 55.3%~89.5%. E-cadherin was not frequently methylated in either setting (range: 18.4%~23.7%. The methylation status of HIN-1, CDH13, RIL, and RARβ2 in lymph nodes metastasis were correlated with that in primary tumors. The Pearson correlation values ranged from 0.624 to 0.472 (p values HIN-1 methylation and hormone status in metastatic lymph nodes. Hypermethylation of HIN-1 in metastasis lymph nodes was significantly associated with expression of ER (odds ratio, 1.070; P = 0.024 and with PR (odds ratio, 1.046; P = 0.026. Conclusions This study suggests that hypermethylation of tumor suppressor genes is extended from primary to metastatic tumors during tumor progression.

  14. Unexpected functional similarities between gatekeeper tumour suppressor genes and proto-oncogenes revealed by systems biology.

    Science.gov (United States)

    Zhao, Yongzhong; Epstein, Richard J

    2011-05-01

    Familial tumor suppressor genes comprise two subgroups: caretaker genes (CTs) that repair DNA, and gatekeeper genes (GKs) that trigger cell death. Since GKs may also induce cell cycle delay and thus enhance cell survival by facilitating DNA repair, we hypothesized that the prosurvival phenotype of GKs could be selected during cancer progression, and we used a multivariable systems biology approach to test this. We performed multidimensional data analysis, non-negative matrix factorization and logistic regression to