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  1. SOCS3 Silencing Attenuates Eosinophil Functions in Asthma Patients

    PubMed Central

    Zafra, Mª Paz; Cañas, Jose A.; Mazzeo, Carla; Gámez, Cristina; Sanz, Veronica; Fernández-Nieto, Mar; Quirce, Santiago; Barranco, Pilar; Ruiz-Hornillos, Javier; Sastre, Joaquín; del Pozo, Victoria

    2015-01-01

    Eosinophils are one of the key inflammatory cells in asthma. Eosinophils can exert a wide variety of actions through expression and secretion of multiple molecules. Previously, we have demonstrated that eosinophils purified from peripheral blood from asthma patients express high levels of suppressor of cytokine signaling 3 (SOCS3). In this article, SOCS3 gene silencing in eosinophils from asthmatics has been carried out to achieve a better understanding of the suppressor function in eosinophils. SOCS3 siRNA treatment drastically reduced SOCS3 expression in eosinophils, leading to an inhibition of the regulatory transcription factors GATA-3 and FoxP3, also interleukin (IL)-10; in turn, an increased STAT3 phosphorilation was observed. Moreover, SOCS3 abrogation in eosinophils produced impaired migration, adhesion and degranulation. Therefore, SOCS3 might be regarded as an important regulator implicated in eosinophil mobilization from the bone marrow to the lungs during the asthmatic process. PMID:25764157

  2. SOCS3 silencing attenuates eosinophil functions in asthma patients.

    PubMed

    Zafra, Ma Paz; Cañas, Jose A; Mazzeo, Carla; Gámez, Cristina; Sanz, Veronica; Fernández-Nieto, Mar; Quirce, Santiago; Barranco, Pilar; Ruiz-Hornillos, Javier; Sastre, Joaquín; del Pozo, Victoria

    2015-01-01

    Eosinophils are one of the key inflammatory cells in asthma. Eosinophils can exert a wide variety of actions through expression and secretion of multiple molecules. Previously, we have demonstrated that eosinophils purified from peripheral blood from asthma patients express high levels of suppressor of cytokine signaling 3 (SOCS3). In this article, SOCS3 gene silencing in eosinophils from asthmatics has been carried out to achieve a better understanding of the suppressor function in eosinophils. SOCS3 siRNA treatment drastically reduced SOCS3 expression in eosinophils, leading to an inhibition of the regulatory transcription factors GATA-3 and FoxP3, also interleukin (IL)-10; in turn, an increased STAT3 phosphorilation was observed. Moreover, SOCS3 abrogation in eosinophils produced impaired migration, adhesion and degranulation. Therefore, SOCS3 might be regarded as an important regulator implicated in eosinophil mobilization from the bone marrow to the lungs during the asthmatic process. PMID:25764157

  3. Zingerone silences quorum sensing and attenuates virulence of Pseudomonas aeruginosa.

    PubMed

    Kumar, Lokender; Chhibber, Sanjay; Kumar, Rajnish; Kumar, Manoj; Harjai, Kusum

    2015-04-01

    Quorum sensing in Pseudomonas aeruginosa plays an imperative role in virulence factor, biofilm formation and antimicrobial resistance. Blocking quorum sensing pathways are viewed as viable anti-virulent therapy in association with traditional antimicrobial therapy. Anti-quorum sensing dietary phytochemicals with may prove to be a safe and viable choice as anti-virulent drug candidates. Previously, our lab proved zingerone as potent anti-biofilm agent hence; further its anti-virulent and anti-quorum activities were evaluated. Zingerone, besides decreasing swimming, swarming and twitching phenotypes of P. aeruginosa PAO1, reduced biofilm forming capacity and production of virulence factors including rhamnolipid, elastase, protease, pyocyanin, cell free and cell bound hemolysin (p<0.001) indicating anti-virulent property attributing towards attenuation of virulence of P. aeruginosa. Further zingerone not only had marked effect on the production of quorum sensing signal molecules by clinical isolates of P. aeruginosa but also showed significant interference with the activation of QS reporter strains. To study the mechanism of blocking quorum sensing cascade, in silico analysis was carried out. Anti-QS activity was attributed to interference with the ligand receptor interaction of zingerone with QS receptors (TraR, LasR, RhlR and PqsR). Zingerone showed a good comparative docking score to respective autoinducer molecules which was even higher than that of vanillin, a proven anti-quorum sensing phytochemical. The results of the present study revealed the anti-quorum sensing activity of zingerone targeting ligand-receptor interaction, hence proposing zingerone as a suitable anti-virulent drug candidate against P. aeruginosa infections. PMID:25704369

  4. RNA-induced silencing attenuates G protein-mediated calcium signals.

    PubMed

    Philip, Finly; Sahu, Shriya; Golebiewska, Urszula; Scarlata, Suzanne

    2016-05-01

    Phospholipase Cβ (PLCβ) is activated by G protein subunits in response to environmental stimuli to increase intracellular calcium. In cells, a significant portion of PLCβ is cytosolic, where it binds a protein complex required for efficient RNA-induced silencing called C3PO (component 3 promoter of RISC). Binding between C3PO and PLCβ raises the possibility that RNA silencing activity can affect the ability of PLCβ to mediate calcium signals. By use of human and rat neuronal cell lines (SK-N-SH and PC12), we show that overexpression of one of the main components of C3PO diminishes Ca(2+) release in response to Gαq/PLCβ stimulation by 30 to 40%. In untransfected SK-N-SH or PC12 cells, the introduction of siRNA(GAPDH) [small interfering RNA(glyceraldehyde 3-phosphate dehydrogenase)] reduces PLCβ-mediated calcium signals by ∼30%, but addition of siRNA(Hsp90) (heat shock protein 90) had little effect. Fluorescence imaging studies suggest an increase in PLCβ-C3PO association in cells treated with siRNA(GAPDH) but not siRNA(Hsp90). Taken together, our studies raise the possibility that Ca(2+) responses to extracellular stimuli can be modulated by components of the RNA silencing machinery.-Philip, F., Sahu, S., Golebiewska, U., Scarlata, S. RNA-induced silencing attenuates G protein-mediated calcium signals. PMID:26862135

  5. Silencing MR-1 attenuates inflammatory damage in mice heart induced by AngII

    SciTech Connect

    Dai, Wenjian; Chen, Haiyang; Jiang, Jiandong; Kong, Weijia; Wang, Yiguang

    2010-01-15

    Myofibrillogenesis regulator-1(MR-1) can aggravate cardiac hypertrophy induced by angiotensin(Ang) II in mice through activation of NF-{kappa}B signaling pathway, and nuclear transcription factor (NF)-{kappa}B and activator protein-1(AP-1) regulate inflammatory and immune responses by increasing the expression of specific inflammatory genes in various tissues including heart. Whether inhibition of MR-1 expression will attenuate AngII-induced inflammatory injury in mice heart has not been explored. Herein, we monitored the activation of NF-{kappa}B and AP-1, together with expression of pro-inflammatory of interleukin(IL)-6, tumor necrosis factor(TNF)-{alpha}, vascular-cell adhesion molecule (VCAM)-1, platelet endothelial cell adhesion molecule (PECAM), and inflammatory cell infiltration in heart of mice which are induced firstly by AngII (PBS),then received MR-1-siRNA or control-siRNA injecting. We found that the activation of NF-{kappa}B and AP-1 was inhibited significantly, together with the decreased expression of IL-6, TNF-{alpha}, VCAM-1, and PECAM in AngII-induced mice myocardium in MR-1-siRNA injection groups compared with control-siRNA injecting groups. However, the expression level of MR-1 was not an apparent change in PBS-infused groups than in unoperation groups, and MR-1-siRNA do not affect the expression of MR-1 in PBS-infused mice. Our findings suggest that silencing MR-1 protected mice myocardium against inflammatory injury induced by AngII by suppression of pro-inflammatory transcription factors NF-{kappa}B and AP-1 signaling pathway.

  6. Gene silencing of endothelial von Willebrand Factor attenuates angiotensin II-induced endothelin-1 expression in porcine aortic endothelial cells.

    PubMed

    Dushpanova, Anar; Agostini, Silvia; Ciofini, Enrica; Cabiati, Manuela; Casieri, Valentina; Matteucci, Marco; Del Ry, Silvia; Clerico, Aldo; Berti, Sergio; Lionetti, Vincenzo

    2016-01-01

    Expression of endothelin (ET)-1 is increased in endothelial cells exposed to angiotensin II (Ang II), leading to endothelial dysfunction and cardiovascular disorders. Since von Willebrand Factor (vWF) blockade improves endothelial function in coronary patients, we hypothesized that targeting endothelial vWF with short interference RNA (siRNA) prevents Ang II-induced ET-1 upregulation. Nearly 65 ± 2% silencing of vWF in porcine aortic endothelial cells (PAOECs) was achieved with vWF-specific siRNA without affecting cell viability and growth. While showing ET-1 similar to wild type cells at rest, vWF-silenced cells did not present ET-1 upregulation during exposure to Ang II (100 nM/24 h), preserving levels of endothelial nitric oxide synthase activity similar to wild type. vWF silencing prevented AngII-induced increase in nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) activity and superoxide anion (O2-) levels, known triggers of ET-1 expression. Moreover, no increase in O2- or ET-1 levels was found in silenced cells treated with AngII or NOX-agonist phorbol ester (PMA 5 nM/48 h). Finally, vWF was required for overexpression of NOX4 and NOX2 in response to AngII and PMA. In conclusion, endothelial vWF knockdown prevented Ang II-induced ET-1 upregulation through attenuation of NOX-mediated O2- production. Our findings reveal a new role of vWF in preventing of Ang II-induced endothelial dysfunction. PMID:27443965

  7. Gene silencing of endothelial von Willebrand Factor attenuates angiotensin II-induced endothelin-1 expression in porcine aortic endothelial cells

    PubMed Central

    Dushpanova, Anar; Agostini, Silvia; Ciofini, Enrica; Cabiati, Manuela; Casieri, Valentina; Matteucci, Marco; Del Ry, Silvia; Clerico, Aldo; Berti, Sergio; Lionetti, Vincenzo

    2016-01-01

    Expression of endothelin (ET)-1 is increased in endothelial cells exposed to angiotensin II (Ang II), leading to endothelial dysfunction and cardiovascular disorders. Since von Willebrand Factor (vWF) blockade improves endothelial function in coronary patients, we hypothesized that targeting endothelial vWF with short interference RNA (siRNA) prevents Ang II-induced ET-1 upregulation. Nearly 65 ± 2% silencing of vWF in porcine aortic endothelial cells (PAOECs) was achieved with vWF-specific siRNA without affecting cell viability and growth. While showing ET-1 similar to wild type cells at rest, vWF-silenced cells did not present ET-1 upregulation during exposure to Ang II (100 nM/24 h), preserving levels of endothelial nitric oxide synthase activity similar to wild type. vWF silencing prevented AngII-induced increase in nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) activity and superoxide anion (O2−) levels, known triggers of ET-1 expression. Moreover, no increase in O2− or ET-1 levels was found in silenced cells treated with AngII or NOX-agonist phorbol ester (PMA 5 nM/48 h). Finally, vWF was required for overexpression of NOX4 and NOX2 in response to AngII and PMA. In conclusion, endothelial vWF knockdown prevented Ang II-induced ET-1 upregulation through attenuation of NOX-mediated O2− production. Our findings reveal a new role of vWF in preventing of Ang II-induced endothelial dysfunction. PMID:27443965

  8. Attenuation of Histone Methyltransferase KRYPTONITE-mediated transcriptional gene silencing by Geminivirus

    PubMed Central

    Sun, Yan-Wei; Tee, Chuan-Sia; Ma, Yong-Huan; Wang, Gang; Yao, Xiang-Mei; Ye, Jian

    2015-01-01

    Although histone H3K9 methylation has been intensively studied in animals and a model plant Arabidopsis thaliana, little is known about the evolution of the histone methyltransferase and its roles in plant biotic stress response. Here we identified a Nicotiana benthamiana homolog of H3K9 histone methyltransferase KRYPTONITE (NbKYP) and demonstrated its fundamental roles on methylation of plant and virus, beside of leading to the suppression of endogenous gene expression and virus replication. NbKYP and another gene encoding DNA methyltransferase CHROMOMETHYLTRANSFERASE 3 (NbCMT3-1) were further identified as the key components of maintenance of transcriptional gene silencing, a DNA methylation involved anti-virus machinery. All three types of DNA methylations (asymmetric CHH and symmetric CHG/CG) were severely affected in NbKYP-silenced plants, but only severe reduction of CHG methylation found in NbCMT3-1-silenced plants. Attesting to the importance of plant histone H3K9 methylation immunity to virus, the virulence of geminiviruses requires virus-encoded trans-activator AC2 which inhibits the expression of KYP via activation of an EAR-motif-containing transcription repressor RAV2 (RELATED TO ABI3 and VP1). The reduction of KYP was correlated with virulence of various similar geminiviruses. These findings provide a novel mechanism of how virus trans-activates a plant endogenous anti-silencing machinery to gain high virulence. PMID:26602265

  9. Silencing of ILK attenuates the abnormal proliferation and migration of human Tenon's capsule fibroblasts induced by TGF-β2.

    PubMed

    Xing, Yao; Cui, Lijun; Kang, Qianyan

    2016-08-01

    The cytokine, transforming growth factor-β (TGF‑β), plays a key role in wound healing and tissue repair. Integrin‑linked kinase (ILK) is a downstream factor of the TGF-β signaling pathway. Research on ILK has mainly focused on its role in the invasion and metastasis of cancer cells. However, little has been reported on the effects of ILK in human Tenon's capsule fibroblasts (HTFs). In this study, we investigated the role of ILK in the proliferation and migration of HTFs exposed to TGF-β2. A lentiviral vector targeting ILK was screened from three candidates and the experimental result indicated that RNA interference can be used to inhibit ILK expression at both the mRNA and protein level. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was used to assess ILK mRNA expression. Cell proliferation was quantified by MTT assay and cell cycle progression was detected by flow cytometric analysis. Migration was measured by wound healing assay. It was observed that the silencing of ILK suppressed the TGF-β2-induced proliferation of HTFs and led to G1 phase cell cycle arrest and the significant downregulation of cyclin D1 expression. The migration ability of the HTFs decreased following the silencing of ILK, while the downregulation of α-smooth muscle actin expression and the upregulation of E-cadherin expression were observed. The findings of our study indicate that the silencing of ILK attenuates the abnormal proliferation and migration of HTFs induced by TGF-β2, which reveals the therapeutic potential of ILK inhibition in the prevention of scarring following glaucoma filtration surgery. PMID:27315599

  10. Silencing of ILK attenuates the abnormal proliferation and migration of human Tenon's capsule fibroblasts induced by TGF-β2

    PubMed Central

    Xing, Yao; Cui, Lijun; Kang, Qianyan

    2016-01-01

    The cytokine, transforming growth factor-β (TGF-β), plays a key role in wound healing and tissue repair. Integrin-linked kinase (ILK) is a downstream factor of the TGF-β signaling pathway. Research on ILK has mainly focused on its role in the invasion and metastasis of cancer cells. However, little has been reported on the effects of ILK in human Tenon's capsule fibroblasts (HTFs). In this study, we investigated the role of ILK in the proliferation and migration of HTFs exposed to TGF-β2. A lentiviral vector targeting ILK was screened from three candidates and the experimental result indicated that RNA interference can be used to inhibit ILK expression at both the mRNA and protein level. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was used to assess ILK mRNA expression. Cell proliferation was quantified by MTT assay and cell cycle progression was detected by flow cytometric analysis. Migration was measured by wound healing assay. It was observed that the silencing of ILK suppressed the TGF-β2-induced proliferation of HTFs and led to G1 phase cell cycle arrest and the significant downregulation of cyclin D1 expression. The migration ability of the HTFs decreased following the silencing of ILK, while the downregulation of α-smooth muscle actin expression and the upregulation of E-cadherin expression were observed. The findings of our study indicate that the silencing of ILK attenuates the abnormal proliferation and migration of HTFs induced by TGF-β2, which reveals the therapeutic potential of ILK inhibition in the prevention of scarring following glaucoma filtration surgery. PMID:27315599

  11. BSCTV C2 attenuates the degradation of SAMDC1 to suppress DNA methylation-mediated gene silencing in Arabidopsis.

    PubMed

    Zhang, Zhonghui; Chen, Hao; Huang, Xiahe; Xia, Ran; Zhao, Qingzhen; Lai, Jianbin; Teng, Kunling; Li, Yin; Liang, Liming; Du, Quansheng; Zhou, Xueping; Guo, Huishan; Xie, Qi

    2011-01-01

    Plant viruses are excellent tools for studying microbial-plant interactions as well as the complexities of host activities. Our study focuses on the role of C2 encoded by Beet severe curly top virus (BSCTV) in the virus-plant interaction. Using BSCTV C2 as bait in a yeast two-hybrid screen, a C2-interacting protein, S-adenosyl-methionine decarboxylase 1 (SAMDC1), was identified from an Arabidopsis thaliana cDNA library. The interaction was confirmed by an in vitro pull-down assay and a firefly luciferase complemention imaging assay in planta. Biochemical analysis further showed that the degradation of the SAMDC1 protein was inhibited by MG132, a 26S proteasome inhibitor, and that C2 could attenuate the degradation of the SAMDC1 protein. Genetic analysis showed that loss of function of SAMDC1 resulted in reduced susceptibility to BSCTV infection and reduced viral DNA accumulation, similar to the effect of BSCTV C2 deficiency. Bisulfite sequencing analysis further showed that C2 deficiency caused enhanced DNA methylation of the viral genome in infected plants. We also showed that C2 can suppress de novo methylation in the FWA transgenic assay in the C2 transgene background. Overexpression of SAMDC1 can mimic the suppressive activity of C2 against green fluorescent protein-directed silencing. These results suggest that C2 interferes with the host defense mechanism of DNA methylation-mediated gene silencing by attenuating the 26S proteasome-mediated degradation of SAMDC1. PMID:21245466

  12. CA3 Synaptic Silencing Attenuates Kainic Acid-Induced Seizures and Hippocampal Network Oscillations123

    PubMed Central

    Yu, Lily M. Y.; Wintzer, Marie E.

    2016-01-01

    Abstract Epilepsy is a neurological disorder defined by the presence of seizure activity, manifest both behaviorally and as abnormal activity in neuronal networks. An established model to study the disorder in rodents is the systemic injection of kainic acid, an excitatory neurotoxin that at low doses quickly induces behavioral and electrophysiological seizures. Although the CA3 region of the hippocampus has been suggested to be crucial for kainic acid-induced seizure, because of its strong expression of kainate glutamate receptors and its high degree of recurrent connectivity, the precise role of excitatory transmission in CA3 in the generation of seizure and the accompanying increase in neuronal oscillations remains largely untested. Here we use transgenic mice in which CA3 pyramidal cell synaptic transmission can be inducibly silenced in the adult to demonstrate CA3 excitatory output is required for both the generation of epileptiform oscillatory activity and the progression of behavioral seizures. PMID:27022627

  13. Silencing of miR-34a Attenuates Cardiac Dysfunction in a Setting of Moderate, but Not Severe, Hypertrophic Cardiomyopathy

    PubMed Central

    Bernardo, Bianca C.; Gao, Xiao-Ming; Tham, Yow Keat; Kiriazis, Helen; Winbanks, Catherine E.; Ooi, Jenny Y. Y.; Boey, Esther J. H.; Obad, Susanna; Kauppinen, Sakari; Gregorevic, Paul; Du, Xiao-Jun; Lin, Ruby C. Y.; McMullen, Julie R.

    2014-01-01

    Therapeutic inhibition of the miR-34 family (miR-34a,-b,-c), or miR-34a alone, have emerged as promising strategies for the treatment of cardiac pathology. However, before advancing these approaches further for potential entry into the clinic, a more comprehensive assessment of the therapeutic potential of inhibiting miR-34a is required for two key reasons. First, miR-34a has ∼40% fewer predicted targets than the miR-34 family. Hence, in cardiac stress settings in which inhibition of miR-34a provides adequate protection, this approach is likely to result in less potential off-target effects. Secondly, silencing of miR-34a alone may be insufficient in settings of established cardiac pathology. We recently demonstrated that inhibition of the miR-34 family, but not miR-34a alone, provided benefit in a chronic model of myocardial infarction. Inhibition of miR-34 also attenuated cardiac remodeling and improved heart function following pressure overload, however, silencing of miR-34a alone was not examined. The aim of this study was to assess whether inhibition of miR-34a could attenuate cardiac remodeling in a mouse model with pre-existing pathological hypertrophy. Mice were subjected to pressure overload via constriction of the transverse aorta for four weeks and echocardiography was performed to confirm left ventricular hypertrophy and systolic dysfunction. After four weeks of pressure overload (before treatment), two distinct groups of animals became apparent: (1) mice with moderate pathology (fractional shortening decreased ∼20%) and (2) mice with severe pathology (fractional shortening decreased ∼37%). Mice were administered locked nucleic acid (LNA)-antimiR-34a or LNA-control with an eight week follow-up. Inhibition of miR-34a in mice with moderate cardiac pathology attenuated atrial enlargement and maintained cardiac function, but had no significant effect on fetal gene expression or cardiac fibrosis. Inhibition of miR-34a in mice with severe pathology

  14. Load-induced inattentional deafness.

    PubMed

    Raveh, Dana; Lavie, Nilli

    2015-02-01

    High perceptual load in a task is known to reduce the visual perception of unattended items (e.g., Lavie, Beck, & Konstantinou, 2014). However, it remains an open question whether perceptual load in one modality (e.g., vision) can affect the detection of stimuli in another modality (e.g., hearing). We report four experiments that establish that high visual perceptual load leads to reduced detection sensitivity in hearing. Participants were requested to detect a tone that was presented during performance of a visual search task of either low or high perceptual load (varied through item similarity). The findings revealed that auditory detection sensitivity was consistently reduced with higher load, and that this effect persisted even when the auditory detection response was made first (before the search response) and when the auditory stimulus was highly expected (50 % present). These findings demonstrate a phenomenon of load-induced deafness and provide evidence for shared attentional capacity across vision and hearing. PMID:25287617

  15. Caspase-12 Silencing Attenuates Inhibitory Effects of Cigarette Smoke Extract on NOD1 Signaling and hBDs Expression in Human Oral Mucosal Epithelial Cells

    PubMed Central

    Wang, Xiang; Qian, Ya-jie; Zhou, Qian; Ye, Pei; Duan, Ning; Huang, Xiao-feng; Zhu, Ya-nan; Li, Jing-jing; Hu, Li-ping; Zhang, Wei-yun; Han, Xiao-dong; Wang, Wen-mei

    2014-01-01

    Cigarette smoke exposure is associated with increased risk of various diseases. Epithelial cells-mediated innate immune responses to infectious pathogens are compromised by cigarette smoke. Although many studies have established that cigarette smoke exposure affects the expression of Toll-liked receptor (TLR), it remains unknown whether the nucleotide-binding oligomerization domain-containing protein 1 (NOD1) expression is affected by cigarette smoke exposure. In the study, we investigated effects of cigarette smoke extract (CSE) on NOD1 signaling in an immortalized human oral mucosal epithelial (Leuk-1) cell line. We first found that CSE inhibited NOD1 expression in a dose-dependent manner. Moreover, CSE modulated the expression of other crucial molecules in NOD1 signaling and human β defensin (hBD) 1, 2 and 3. We found that RNA interference-induced Caspase-12 silencing increased NOD1 and phospho-NF-κB (p-NF-κB) expression and down-regulated RIP2 expression. The inhibitory effects of CSE on NOD1 signaling can be attenuated partially through Caspase-12 silencing. Intriguingly, Caspase-12 silencing abrogated inhibitory effects of CSE on hBD1, 3 expression and augmented induced effect of CSE on hBD2 expression. Caspase-12 could play a vital role in the inhibitory effects of cigarette smoke on NOD1 signaling and hBDs expression in oral mucosal epithelial cells. PMID:25503380

  16. Caspase-12 silencing attenuates inhibitory effects of cigarette smoke extract on NOD1 signaling and hBDs expression in human oral mucosal epithelial cells.

    PubMed

    Wang, Xiang; Qian, Ya-jie; Zhou, Qian; Ye, Pei; Duan, Ning; Huang, Xiao-feng; Zhu, Ya-nan; Li, Jing-jing; Hu, Li-ping; Zhang, Wei-yun; Han, Xiao-dong; Wang, Wen-mei

    2014-01-01

    Cigarette smoke exposure is associated with increased risk of various diseases. Epithelial cells-mediated innate immune responses to infectious pathogens are compromised by cigarette smoke. Although many studies have established that cigarette smoke exposure affects the expression of Toll-liked receptor (TLR), it remains unknown whether the nucleotide-binding oligomerization domain-containing protein 1 (NOD1) expression is affected by cigarette smoke exposure. In the study, we investigated effects of cigarette smoke extract (CSE) on NOD1 signaling in an immortalized human oral mucosal epithelial (Leuk-1) cell line. We first found that CSE inhibited NOD1 expression in a dose-dependent manner. Moreover, CSE modulated the expression of other crucial molecules in NOD1 signaling and human β defensin (hBD) 1, 2 and 3. We found that RNA interference-induced Caspase-12 silencing increased NOD1 and phospho-NF-κB (p-NF-κB) expression and down-regulated RIP2 expression. The inhibitory effects of CSE on NOD1 signaling can be attenuated partially through Caspase-12 silencing. Intriguingly, Caspase-12 silencing abrogated inhibitory effects of CSE on hBD1, 3 expression and augmented induced effect of CSE on hBD2 expression. Caspase-12 could play a vital role in the inhibitory effects of cigarette smoke on NOD1 signaling and hBDs expression in oral mucosal epithelial cells. PMID:25503380

  17. Silencing MicroRNA-155 Attenuates Cardiac Injury and Dysfunction in Viral Myocarditis via Promotion of M2 Phenotype Polarization of Macrophages.

    PubMed

    Zhang, Yingying; Zhang, Mengying; Li, Xueqin; Tang, Zongsheng; Wang, Xiangmin; Zhong, Min; Suo, Qifeng; Zhang, Yao; Lv, Kun

    2016-01-01

    Macrophage infiltration is a hallmark feature of viral myocarditis. As studies have shown that microRNA-155 regulates the differentiation of macrophages, we aimed to investigate the role of microRNA-155 in VM. We report that silencing microRNA-155 protects mice from coxsackievirus B3 induced myocarditis. We found that microRNA-155 expression was upregulated and localized primarily in heart-infiltrating macrophages and CD4(+) T lymphocytes during acute myocarditis. In contrast with wildtype (WT) mice, microRNA-155(-/-) mice developed attenuated viral myocarditis, which was characterized by decreased cardiac inflammation and decreased intracardiac CD45(+) leukocytes. Hearts of microRNA-155(-/-) mice expressed decreased levels of the IFN-γ and increased levels of the cytokines IL-4 and IL-13. Although total CD4(+) and regulatory T cells were unchanged in miR-155(-/-) spleen proportionally, the activation of T cells and CD4(+) T cell proliferation in miR-155(-/-) mice were significantly decreased. Beyond the acute phase, microRNA-15(5-/-) mice had reduced mortality and improved cardiac function during 5 weeks of follow-up. Moreover, silencing microRNA-155 led to increased levels of alternatively-activated macrophages (M2) and decreased levels of classically-activated macrophages (M1) in the heart. Combined, our studies suggest that microRNA-155 confers susceptibility to viral myocarditis by affecting macrophage polarization, and thus may be a potential therapeutic target for viral myocarditis. PMID:26931072

  18. Silencing MicroRNA-155 Attenuates Cardiac Injury and Dysfunction in Viral Myocarditis via Promotion of M2 Phenotype Polarization of Macrophages

    PubMed Central

    Zhang, Yingying; Zhang, Mengying; Li, Xueqin; Tang, Zongsheng; Wang, Xiangmin; Zhong, Min; Suo, Qifeng; Zhang, Yao; Lv, Kun

    2016-01-01

    Macrophage infiltration is a hallmark feature of viral myocarditis. As studies have shown that microRNA-155 regulates the differentiation of macrophages, we aimed to investigate the role of microRNA-155 in VM. We report that silencing microRNA-155 protects mice from coxsackievirus B3 induced myocarditis. We found that microRNA-155 expression was upregulated and localized primarily in heart-infiltrating macrophages and CD4+ T lymphocytes during acute myocarditis. In contrast with wildtype (WT) mice, microRNA-155−/− mice developed attenuated viral myocarditis, which was characterized by decreased cardiac inflammation and decreased intracardiac CD45+ leukocytes. Hearts of microRNA-155−/− mice expressed decreased levels of the IFN-γ and increased levels of the cytokines IL-4 and IL-13. Although total CD4+ and regulatory T cells were unchanged in miR-155−/− spleen proportionally, the activation of T cells and CD4+ T cell proliferation in miR-155−/− mice were significantly decreased. Beyond the acute phase, microRNA-155−/− mice had reduced mortality and improved cardiac function during 5 weeks of follow-up. Moreover, silencing microRNA-155 led to increased levels of alternatively-activated macrophages (M2) and decreased levels of classically-activated macrophages (M1) in the heart. Combined, our studies suggest that microRNA-155 confers susceptibility to viral myocarditis by affecting macrophage polarization, and thus may be a potential therapeutic target for viral myocarditis. PMID:26931072

  19. Allele-specific silencing of mutant p53 attenuates dominant-negative and gain-of-function activities

    PubMed Central

    Iyer, Swathi V.; Parrales, Alejandro; Begani, Priya; Narkar, Akshay; Adhikari, Amit S.; Martinez, Luis A.; Iwakuma, Tomoo

    2016-01-01

    Many p53 hotspot mutants not only lose the transcriptional activity, but also show dominant-negative (DN) and oncogenic gain-of-function (GOF) activities. Increasing evidence indicates that knockdown of mutant p53 (mutp53) in cancer cells reduces their aggressive properties, suggesting that survival and proliferation of cancer cells are, at least partially, dependent on the presence of mutp53. However, these p53 siRNAs can downregulate both wild-type p53 (wtp53) and mutp53, which limits their therapeutic applications. In order to specifically deplete mutp53, we have developed allele-specific siRNAs against p53 hotspot mutants and validated their biological effects in the absence or presence of wtp53. First, the mutp53-specific siRNAs selectively reduced protein levels of matched p53 mutants with minimal reduction in wtp53 levels. Second, downregulation of mutp53 in cancer cells expressing a mutp53 alone (p53mut) resulted in significantly decreased cell proliferation and migration. Third, transfection of mutp53-specific siRNAs in cancer cells expressing both wtp53 and mutp53 also reduced cell proliferation and migration with increased transcripts of p53 downstream target genes, which became further profound when cells were treated with an MDM2 inhibitor Nutlin-3a or a chemotherapeutic agent doxorubicin. These results indicate that depletion of mutp53 by its specific siRNA restored endogenous wtp53 activity in cells expressing both wtp53 and mutp53. This is the first study demonstrating biological effects and therapeutic potential of allele-specific silencing of mutp53 by mutp53-specific siRNAs in cancer cells expressing both wtp53 and mutp53, thus providing a novel strategy towards targeted cancer therapies. PMID:26700961

  20. Cystathionine-γ-lyase gene silencing with siRNA in monocytes/ macrophages attenuates inflammation in cecal ligation and puncture-induced sepsis in the mouse.

    PubMed

    Badiei, A; Chambers, S T; Gaddam, R R; Bhatia, M

    2016-03-01

    Hydrogen sulphide is an endogenous inflammatory mediator produced by cystathionine-γ-lyase (CSE) in macrophages. To determine the role of H2S and macrophages in sepsis, we used small interference RNA (siRNA) to target the CSE gene and investigated its effect in a mouse model of sepsis. Cecal ligation puncture (CLP)-induced sepsis is characterized by increased levels of myeloperoxidase (MPO) activity, morphological changes in liver and pro-inflammatory cytokines and chemokines in the liver and lung. SiRNA treatment attenuated inflammation in the liver and lungs of mice following CLP-induced sepsis. Liver MPO activity increased in CLP-induced sepsis and treatment with siRNA significantly reduced this. Similarly, lung MPO activity increased following induction of sepsis with CLP while siRNA treatment significantly reduced MPO activity. Liver and lung cytokine and chemokine levels in CLP-induced sepsis reduced following treatment with siRNA. These findings show a crucial pro-inflammatory role for H2S synthesized by CSE in macrophages in sepsis and suggest CSE gene silencing with siRNA as a potential therapeutic approach for this condition. PMID:26949091

  1. Attenuation of atherosclerotic lesions in diabetic apolipoprotein E-deficient mice using gene silencing of macrophage migration inhibitory factor

    PubMed Central

    Sun, Hui; Zhang, XianJun; Zhao, Lei; Zhen, Xi; Huang, ShanYing; Wang, ShaSha; He, Hong; Liu, ZiMo; Xu, NaNa; Yang, FaLin; Qu, ZhongHua; Ma, ZhiYong; Zhang, Cheng; Zhang, Yun; Hu, Qin

    2015-01-01

    Macrophage migration inhibitory factor (MIF) involves the pathogenesis of atherosclerosis (AS) and increased plasma MIF levels in diabetes mellitus (DM) patients are associated with AS. Here, we have been suggested that MIF could be a critical contributor for the pathological process of diabetes-associated AS by using adenovirus-mediated RNA interference. First, streptozotocin (STZ)-induced diabetic animal model was constructed in 114 apolipoprotein E-deficient mice (apoE−/− mice) fed on a regular chow diet. Then, the animals were randomly divided into three groups: Adenovirus-mediated MIF interference (Ad-MIFi), Ad-enhanced green fluorescent protein (EGFP) and normal saline (NS) group (n ≈ 33/group). Non-diabetic apoE−/− mice (n = 35) were served as controls. Ad-MIFi, Ad-EGFP and NS were, respectively, injected into the tail vein of mice from Ad-MIFi, Ad-EGFP and NS group, which were injected repeatedly 4 weeks later. Physical, biochemical, morphological and molecular parameters were measured. The results showed that diabetic apoE−/− mice had significantly aggravated atherosclerotic lesions. MIF gene interference attenuated atherosclerotic lesions and stabilized atheromatous plaque, accompanied by the decreased macrophages and lipids deposition and inflammatory cytokines production, improved glucose intolerance and plasma cholesterol level, the decreased ratio of matrix matalloproteinase-2/tissue inhibitor of metalloproteinase-1 and plaque instability index. An increased expression of MIF and its ligand CD74 was also detected in the diabetic patients with coronary artery disease. The results suggest that MIF gene interference is able to inhibit atherosclerotic lesions and increase plaque stability in diabetic apoE−/−mice. MIF inhibition could be a novel and promising approach to the treatment of DM-associated AS. PMID:25661015

  2. Transient silencing of Npr3 gene expression improved the circulatory levels of atrial natriuretic peptides and attenuated β-adrenoceptor activation- induced cardiac hypertrophic growth in experimental rats.

    PubMed

    Venkatesan, Balaji; Tumala, Anusha; Subramanian, Vimala; Vellaichamy, Elangovan

    2016-07-01

    Natriuretic peptide receptor-C (NPR-C) is considered as a clearance receptor that maintains the circulatory levels of natriuretic peptides. It has been suggested that augmented expression of NPR-C as a cause for the diminished anti-hypertrophic action of natriuretic peptides in the failing heart. Hence, we sought to determine the level of Npr3 gene (coding for NPR-C) expression in the Isoproterenol (ISO) treated Wistar rats. In addition, we studied the effect of Npr3 gene silencing on the hypertrophic growth. A significant increase in heart weight-to-body weight ratio (HW/BW-24%,P<0.01), an indicator of cardiac hypertrophic growth was observed in the ISO (10mg/kg BW/day,i.p for 7 days) treated rats. As expected, the cardiac NPR-C protein expression was significantly increased by 4 fold as compared to control rats. In parallel, the circulatory atrial natriuretic peptide (ANP) level was significantly decreased (2 fold) in ISO treated rats. Upon treatment with siRNA-Npr3, a significant decrease in the cardiac NPR-C protein expression (70%,P<0.01), HW/BW ratio (70%,P<0.01) and hypertrophic marker genes (α-Sk, β-MHC, c-fos, P<0.01, respectively) mRNA expression were observed. Interestingly, the circulatory ANP level was increased by 1.5 fold in the siRNA-Npr3 treated rats as compared to ISO treated rats. Moreover, the cardiac collagen content, matrixmetalloprotinases-2 (MMP-2) and enzymatic antioxidant status (P<0.01, respectively) were found to be restored back to near normal upon siRNA-Npr3 treatment. Taken together, the results of this study indicates that specific down-regulation of Npr3 gene improves the circulatory levels of ANP and antioxidant system and there by attenuates the β-adrenoceptor over-activation mediated cardiac hypertrophic growth in experimental rats. PMID:27108789

  3. Tissue transglutaminase is involved in mechanical load-induced osteogenic differentiation of human ligamentum flavum cells.

    PubMed

    Chao, Yuan-Hung; Huang, Shih-Yung; Yang, Ruei-Cheng; Sun, Jui-Sheng

    2016-07-01

    Mechanical load-induced osteogenic differentiation might be the key cellular event in the calcification and ossification of ligamentum flavum. The aim of this study was to investigate the influence of tissue transglutaminase (TGM2) on mechanical load-induced osteogenesis of ligamentum flavum cells. Human ligamentum flavum cells were obtained from 12 patients undergoing lumbar spine surgery. Osteogenic phenotypes of ligamentum flavum cells, such as alkaline phosphatase (ALP), Alizarin red-S stain, and gene expression of osteogenic makers were evaluated following the administration of mechanical load and BMP-2 treatment. The expression of TGM2 was evaluated by real-time PCR, Western blotting, and enzyme-linked immunosorbent assay (ELISA) analysis. Our results showed that mechanical load in combination with BMP-2 enhanced calcium deposition and ALP activity. Mechanical load significantly increased ALP and OC gene expression on day 3, whereas BMP-2 significantly increased ALP, OPN, and Runx2 on day 7. Mechanical load significantly induced TGM2 gene expression and enzyme activity in human ligamentum flavum cells. Exogenous TGM2 increased ALP and OC gene expression; while, inhibited TG activity significantly attenuated mechanical load-induced and TGM2-induced ALP activity. In summary, mechanical load-induced TGM2 expression and enzyme activity is involved in the progression of the calcification of ligamentum flavum. PMID:27115725

  4. AUTOPHAGY IN LOAD-INDUCED HEART DISEASE

    PubMed Central

    Zhu, Hongxin; Rothermel, Beverly A.; Hill, Joseph A.

    2009-01-01

    The heart is a highly plastic organ capable of remodeling in response to changes in physiological or pathological demand. When workload increases, the heart compensates through hypertrophic growth of individual cardiomyocytes to increase cardiac output. However, sustained stress, such as occurs with hypertension or following myocardial infarction, triggers changes in sarcomeric protein composition and energy metabolism, loss of cardiomyocytes, ventricular dilation, reduced pump function, and ultimately heart failure. It has been known for some time that autophagy is active in cardiomyocytes, occurring at increased levels in disease. Yet the potential contribution of cardiomyocyte autophagy to ventricular remodeling and disease pathogenesis has only recently been explored. This latter fact stems largely from the recent emergence of tools to probe molecular mechanisms governing cardiac plasticity and to define the role of autophagic flux in the context of heart disease. In this chapter, we briefly review prominent mouse models useful in the study of load-induced heart disease and standard techniques used to assess whether a molecular or cellular event is adaptive or maladaptive. We then outline methods available for monitoring autophagic activity in the heart, providing detailed protocols for several techniques unique to working with heart and other striated muscles. PMID:19216915

  5. Silencing Glycogen Synthase Kinase-3β Inhibits Acetaminophen Hepatotoxicity and Attenuates JNK Activation and Loss of Glutamate Cysteine Ligase and Myeloid Cell Leukemia Sequence 1*

    PubMed Central

    Shinohara, Mie; Ybanez, Maria D.; Win, Sanda; Than, Tin Aung; Jain, Shilpa; Gaarde, William A.; Han, Derick; Kaplowitz, Neil

    2010-01-01

    Previously we demonstrated that c-Jun N-terminal kinase (JNK) plays a central role in acetaminophen (APAP)-induced liver injury. In the current work, we examined other possible signaling pathways that may also contribute to APAP hepatotoxicity. APAP treatment to mice caused glycogen synthase kinase-3β (GSK-3β) activation and translocation to mitochondria during the initial phase of APAP-induced liver injury (∼1 h). The silencing of GSK-3β, but not Akt-2 (protein kinase B) or glycogen synthase kinase-3α (GSK-3α), using antisense significantly protected mice from APAP-induced liver injury. The silencing of GSK-3β affected several key pathways important in conferring protection against APAP-induced liver injury. APAP treatment was observed to promote the loss of glutamate cysteine ligase (GCL, rate-limiting enzyme in GSH synthesis) in liver. The silencing of GSK-3β decreased the loss of hepatic GCL, and promoted greater GSH recovery in liver following APAP treatment. Silencing JNK1 and -2 also prevented the loss of GCL. APAP treatment also resulted in GSK-3β translocation to mitochondria and the degradation of myeloid cell leukemia sequence 1 (Mcl-1) in mitochondrial membranes in liver. The silencing of GSK-3β reduced Mcl-1 degradation caused by APAP treatment. The silencing of GSK-3β also resulted in an inhibition of the early phase (0–2 h), and blunted the late phase (after 4 h) of JNK activation and translocation to mitochondria in liver following APAP treatment. Taken together our results suggest that activation of GSK-3β is a key mediator of the initial phase of APAP-induced liver injury through modulating GCL and Mcl-1 degradation, as well as JNK activation in liver. PMID:20061376

  6. RuBPCase activase mediates growth-defense tradeoffs: Silencing RCA redirects JA flux from JA-Ile to MeJA to attenuate induced defense responses in Nicotiana attenuata

    PubMed Central

    Mitra, Sirsha; Baldwin, Ian T.

    2014-01-01

    Summary RuBPCase activase (RCA), an abundant photosynthetic protein is strongly down-regulated in response to Manduca sexta’s oral secretion (OS) in Nicotiana attenuata. RCA-silenced plants are impaired not only in photosynthetic capacity and growth, but also in jasmonic acid (JA)-isoleucine (Ile) signaling, and herbivore resistance mediated by JA-Ile dependent defense traits. These responses are consistent with a resource-based growth-defense trade-off. Since JA+Ile-supplementation of OS restored WT levels of JA-Ile, defenses and resistance to M. sexta, but OS supplemented individually with JA- or Ile did not, the JA-Ile deficiency of RCA-silenced plants could not be attributed to lower JA or Ile pools or JAR4/6 conjugating activity. Similar levels of JA-Ile derivatives after OS elicitation indicated unaltered JA-Ile turnover and lower levels of other JA-conjugates ruled out competition from other conjugation reactions. RCA-silenced plants accumulated more methyl jasmonate (MeJA) after OS elicitation, which corresponded with increased jasmonate methyltransferase (JMT) activity. RCA-silencing phenocopies JMT over-expression, wherein elevated JMT activity redirects OS-elicited JA flux towards inactive MeJA, creating a JA sink which depletes JA-Ile and its associated defense responses. Hence RCA plays an additional non-photosynthetic role in attenuating JA-mediated defenses and their associated costs potentially allowing plants to anticipate resource-based constraints on growth before they actually occur. PMID:24491116

  7. Rescuing Loading Induced Bone Formation at Senescence

    PubMed Central

    Srinivasan, Sundar; Ausk, Brandon J.; Prasad, Jitendra; Threet, Dewayne; Bain, Steven D.; Richardson, Thomas S.; Gross, Ted S.

    2010-01-01

    The increasing incidence of osteoporosis worldwide requires anabolic treatments that are safe, effective, and, critically, inexpensive given the prevailing overburdened health care systems. While vigorous skeletal loading is anabolic and holds promise, deficits in mechanotransduction accrued with age markedly diminish the efficacy of readily complied, exercise-based strategies to combat osteoporosis in the elderly. Our approach to explore and counteract these age-related deficits was guided by cellular signaling patterns across hierarchical scales and by the insight that cell responses initiated during transient, rare events hold potential to exert high-fidelity control over temporally and spatially distant tissue adaptation. Here, we present an agent-based model of real-time Ca2+/NFAT signaling amongst bone cells that fully described periosteal bone formation induced by a wide variety of loading stimuli in young and aged animals. The model predicted age-related pathway alterations underlying the diminished bone formation at senescence, and hence identified critical deficits that were promising targets for therapy. Based upon model predictions, we implemented an in vivo intervention and show for the first time that supplementing mechanical stimuli with low-dose Cyclosporin A can completely rescue loading induced bone formation in the senescent skeleton. These pre-clinical data provide the rationale to consider this approved pharmaceutical alongside mild physical exercise as an inexpensive, yet potent therapy to augment bone mass in the elderly. Our analyses suggested that real-time cellular signaling strongly influences downstream bone adaptation to mechanical stimuli, and quantification of these otherwise inaccessible, transient events in silico yielded a novel intervention with clinical potential. PMID:20838577

  8. The silencing of Pokemon attenuates the proliferation of hepatocellular carcinoma cells in vitro and in vivo by inhibiting the PI3K/Akt pathway.

    PubMed

    Lin, Chan-Chan; Zhou, Jing-Ping; Liu, Yun-Peng; Liu, Jing-Jing; Yang, Xiao-Ning; Jazag, Amarsanaa; Zhang, Zhi-Ping; Guleng, Bayasi; Ren, Jian-Lin

    2012-01-01

    Pokemon (POK erythroid myeloid ontogenic factor), which belongs to the POK protein family, is also called LRF, OCZF and FBI-1. As a transcriptional repressor, Pokemon assumes a critical function in cellular differentiation and oncogenesis. Our study identified an oncogenic role for Pokemon in human hepatocellular carcinoma (HCC). We successfully established human HepG2 and Huh-7 cell lines in which Pokemon was stably knocked down. We demonstrated that Pokemon silencing inhibited cell proliferation and migration. Pokemon knockdown inhibited the PI3K/Akt and c-Raf/MEK/ERK pathways and modulated the expression of various cell cycle regulators in HepG2 and Huh-7 cells. Therefore, Pokemon may also be involved in cell cycle progression in these cells. We confirmed that Pokemon silencing suppresses hepatocellular carcinoma growth in tumor xenograft mice. These results suggest that Pokemon promotes cell proliferation and migration in hepatocellular carcinoma and accelerates tumor development in an Akt- and ERK-signaling-dependent manner. PMID:23300578

  9. Adenylyl cyclase 6 mediates loading-induced bone adaptation in vivo

    PubMed Central

    Lee, Kristen L.; Hoey, David A.; Spasic, Milos; Tang, Tong; Hammond, H. Kirk; Jacobs, Christopher R.

    2014-01-01

    Primary cilia are single, nonmotile, antenna-like structures extending from the apical membrane of most mammalian cells. They may mediate mechanotransduction, the conversion of external mechanical stimuli into biochemical intracellular signals. Previously we demonstrated that adenylyl cyclase 6 (AC6), a membrane-bound enzyme enriched in primary cilia of MLO-Y4 osteocyte-like cells, may play a role in a primary cilium-dependent mechanism of osteocyte mechanotransduction in vitro. In this study, we determined whether AC6 deletion impairs loading-induced bone formation in vivo. Skeletally mature mice with a global knockout of AC6 exhibited normal bone morphology and responded to osteogenic chemical stimuli similar to wild-type mice. Following ulnar loading over 3 consecutive days, bone formation parameters were assessed using dynamic histomorphometry. Mice lacking AC6 formed significantly less bone than control animals (41% lower bone formation rate). Furthermore, there was an attenuated flow-induced increase in COX-2 mRNA expression levels in primary bone cells isolated from AC6 knockout mice compared to controls (1.3±0.1- vs. 2.6±0.2-fold increase). Collectively, these data indicate that AC6 plays a role in loading-induced bone adaptation, and these findings are consistent with our previous studies implicating primary cilia and AC6 in a novel mechanism of osteocyte mechanotransduction.—Lee, K. L., Hoey, D. A., Spasic, M., Tang, T., Hammond, H. K., Jacobs, C. R. Adenylyl cyclase 6 mediates loading-induced bone adaptation in vivo. PMID:24277577

  10. Adenylyl cyclase 6 mediates loading-induced bone adaptation in vivo.

    PubMed

    Lee, Kristen L; Hoey, David A; Spasic, Milos; Tang, Tong; Hammond, H Kirk; Jacobs, Christopher R

    2014-03-01

    Primary cilia are single, nonmotile, antenna-like structures extending from the apical membrane of most mammalian cells. They may mediate mechanotransduction, the conversion of external mechanical stimuli into biochemical intracellular signals. Previously we demonstrated that adenylyl cyclase 6 (AC6), a membrane-bound enzyme enriched in primary cilia of MLO-Y4 osteocyte-like cells, may play a role in a primary cilium-dependent mechanism of osteocyte mechanotransduction in vitro. In this study, we determined whether AC6 deletion impairs loading-induced bone formation in vivo. Skeletally mature mice with a global knockout of AC6 exhibited normal bone morphology and responded to osteogenic chemical stimuli similar to wild-type mice. Following ulnar loading over 3 consecutive days, bone formation parameters were assessed using dynamic histomorphometry. Mice lacking AC6 formed significantly less bone than control animals (41% lower bone formation rate). Furthermore, there was an attenuated flow-induced increase in COX-2 mRNA expression levels in primary bone cells isolated from AC6 knockout mice compared to controls (1.3±0.1- vs. 2.6±0.2-fold increase). Collectively, these data indicate that AC6 plays a role in loading-induced bone adaptation, and these findings are consistent with our previous studies implicating primary cilia and AC6 in a novel mechanism of osteocyte mechanotransduction. PMID:24277577

  11. The caspase-1 inhibitor AC-YVAD-CMK attenuates acute gastric injury in mice: involvement of silencing NLRP3 inflammasome activities

    PubMed Central

    Zhang, Fang; Wang, Liang; Wang, Jun-jie; Luo, Peng-fei; Wang, Xing-tong; Xia, Zhao-fan

    2016-01-01

    This study evaluated the protective effects of inhibiting caspase-1 activity or gastric acid secretion on acute gastric injury in mice. AC-YVAD-CMK, omeprazole, or vehicle were administered to mice before cold-restraint stress- or ethanol-induced gastric injury. Survival rates and histological evidence of gastric injury of mice pretreated with AC-YVAD-CMK or omeprazole, and exposed to cold-restraint stress, improved significantly relative to the vehicle group. The increased levels of tumour necrosis factor-α, interleukin (IL)-1β, IL-6, and IL-18 following cold-stress injury were decreased by AC-YVAD-CMK, but not omeprazole, pretreatment. The increased expression of CD68 in gastric tissues was inhibited significantly by AC-YVAD-CMK pretreatment. Inhibiting caspase-1 activity in the NLRP3 inflammasome decreased gastric cell apoptosis, and the expression of Bax and cleaved caspase-3. AC-YVAD-CMK pretreatment significantly inhibited cold-restraint stress-induced increases in the expression of phosphorylated IκB-alpha and P38. General anatomy and histological results showed the protective effect of AC-YVAD-CMK on ethanol-induced acute gastric injury. Overall, our results showed that the caspase-1 inhibitor AC-YVAD-CMK protected against acute gastric injury in mice by affecting the NLRP3 inflammasome and attenuating inflammatory processes and apoptosis. This was similar to the mechanism associated with NF-κB and P38 mitogen-activated protein kinase signalling pathways. PMID:27053298

  12. The Molecular Basis for Load-Induced Skeletal Muscle Hypertrophy

    PubMed Central

    Marcotte, George R.; West, Daniel W.D.; Baar, Keith

    2016-01-01

    In a mature (weight neutral) animal, an increase in muscle mass only occurs when the muscle is loaded sufficiently to cause an increase in myofibrillar protein balance. A tight relationship between muscle hypertrophy, acute increases in protein balance, and the activity of the mechanistic target of rapamycin complex 1 (mTORC1) was demonstrated 15 years ago. Since then, our understanding of the signals that regulate load-induced hypertrophy has evolved considerably. For example, we now know that mechanical load activates mTORC1 in the same way as growth factors, by moving TSC2 (a primary inhibitor of mTORC1) away from its target (the mTORC activator) Rheb. However, the kinase that phosphorylates and moves TSC2 is different in the two processes. Similarly, we have learned that a distinct pathway exists whereby amino acids activate mTORC1 by moving it to Rheb. While mTORC1 remains at the forefront of load-induced hypertrophy, the importance of other pathways that regulate muscle mass are becoming clearer. Myostatin, is best known for its control of developmental muscle size. However, new mechanisms to explain how loading regulates this process are suggesting that it could play an important role in hypertrophic muscle growth as well. Lastly, new mechanisms are highlighted for how β2 receptor agonists could be involved in load-induced muscle growth and why these agents are being developed as non-exercise-based therapies for muscle atrophy. Overall, the results highlight how studying the mechanism of load-induced skeletal muscle mass is leading the development of pharmaceutical interventions to promote muscle growth in those unwilling or unable to perform resistance exercise. PMID:25359125

  13. Practising Silence in Teaching

    ERIC Educational Resources Information Center

    Forrest, Michelle

    2013-01-01

    The concept "silence" has diametrically opposed meanings; it connotes peace and contemplation as well as death and oblivion. Silence can also be considered a practice. There is keeping the rule of silence to still the mind and find inner truth, as well as forcibly silencing in the sense of subjugating another to one's own purposes.…

  14. Load-Induced Confinement Activates Diamond Lubrication by Water

    NASA Astrophysics Data System (ADS)

    Zilibotti, G.; Corni, S.; Righi, M. C.

    2013-10-01

    Tribochemical reactions are chemical processes, usually involving lubricant or environment molecules, activated at the interface between two solids in relative motion. They are difficult to be monitored in situ, which leaves a gap in the atomistic understanding required for their control. Here we report the real-time atomistic description of the tribochemical reactions occurring at the interface between two diamond films in relative motion, by means of large scale ab initio molecular dynamics. We show that the load-induced confinement is able to catalyze diamond passivation by water dissociative adsorption. Such passivation decreases the energy of the contacting surfaces and increases their electronic repulsion. At sufficiently high coverages, the latter prevents surface sealing, thus lowering friction. Our findings elucidate effects of the nanoscale confinement on reaction kinetics and surface thermodynamics, which are important for the design of new lubricants.

  15. Noise suppression by flexible fan silencers

    SciTech Connect

    Partyka, J.; Kelly, T.R.J.

    1995-12-31

    This paper presents the results on noise testing of a fan only, as well as the results of a steel silencer and of flexible silencers that were connected directly to a fan. On-site facilities and free-field method set by the British Standards Institution were used to measure and then compare the fan only and different practical silencer configuration setups. In order to determine the fan-silencer combination that would give the maximum noise attenuation, total noise intensity, noise contributed to by the fan motor only, as well as aerodynamical noise created through air interacting with the fan parts were considered to obtain decibel readings for the octave bands. Subsequently, the optimal configuration found was the setup with flexible silencers on the fan inlet and the fan outlet. If only one silencer is used, it should be installed on the fan inlet. The aerodynamic noise affects the low frequencies. The flow noise is then overtaken at 1 kHz by the mechanical noise.

  16. RNA silencing: an antiviral mechanism.

    PubMed

    Csorba, T; Pantaleo, V; Burgyán, J

    2009-01-01

    RNA silencing is an evolutionarily conserved sequence-specific gene-inactivation system that also functions as an antiviral mechanism in higher plants and insects. To overcome antiviral RNA silencing, viruses express silencing-suppressor proteins which can counteract the host silencing-based antiviral process. After the discovery of virus-encoded silencing suppressors, it was shown that these viral proteins can target one or more key points in the silencing machinery. Here we review recent progress in our understanding of the mechanism and function of antiviral RNA silencing in plants, and on the virus's counterattack by expression of silencing-suppressor proteins. We also discuss emerging evidence that RNA silencing and expression of viral silencing-suppressor proteins are tools forged as a consequence of virus-host coevolution for fine-tuning host-pathogen coexistence. PMID:20109663

  17. Analysis and design of pod silencers

    NASA Astrophysics Data System (ADS)

    Munjal, M. L.

    2003-05-01

    Parallel baffle mufflers or split silencers are used extensively in heating, ventilation and air conditioning systems for increased attenuation of noise within a short or given length. Acoustic analysis of rectangular parallel baffle mufflers runs on the same lines as that of a rectangular duct lined on two sides. This simplification would not hold for circular configurations. Often, a cylindrical pod is inserted into a circular lined duct to increase its attenuation (or transmission loss), thereby making the flow passage annular and providing an additional absorptive layer on the inner side of this annular passage. This configuration, called a pod silencer, is analyzed here for the four-pole parameters as well as transmission loss, making use of the bulk reaction model. The effect of thin protective film or a highly perforated metallic plate is duly incorporated by means of a grazing-flow impedance. Use of appropriate boundary conditions leads to a set of linear homogeneous equations which in turn lead to a transcendental frequency equation in the unknown complex axial wave number. This is solved by means of the Newton-Raphson method, and the axial wave number is then used in the expressions for transmission loss as well as the transfer matrix parameters. Finally, results of a parametric study are reported to help the designer in optimization of a pod silencer configuration within a given overall size for minimal cost.

  18. The Fruit of Silence

    ERIC Educational Resources Information Center

    Nelson, Marilyn

    2006-01-01

    This presentation explores how contemplative practices, especially those anchored in an active listening to silence, are integrated into creative writing courses. It pays particular attention to a course taught at the United States Military Academy at West Point and to a course on the poetry of war and peace taught at the University of…

  19. The Gift of Silence

    ERIC Educational Resources Information Center

    Haskins, Cathleen

    2011-01-01

    Slowing down, quieting the mind and body, and experiencing silence nourishes the spirit. Montessori educators are mandated to cultivate not just the intellect but the whole child. They recognize that nurturing the spirit of the child is part of what makes this form of education work so well. This article discusses the benefits of stillness and…

  20. The Conspiracy Of Silence

    PubMed Central

    Cohen, Saul

    1980-01-01

    The issue of the impaired physician is compounded by not only mass denial of the problem, but also a 'conspiracy of silence' among many groups associated with the physician. The conspirators—including the physician himself, his family, community, professional colleagues and nurses as well as hospital boards and administration—are unable to reconcile deteriorating performance due to alcohol or drugs with an otherwise gifted professional who should know the dangers of substance abuse. They may also fear the effects of labels such as 'alcoholism', 'drug abuse' and 'psychiatric illness', and they may not know how to handle the problem. A Saskatchewan Medical Association committee was formed in 1976 to penetrate the shroud of silence by identifying and rehabilitating impaired physicians. However, the committee's experience since that time has been largely frustrating because its function has been viewed as more punitive than therapeutic. Imagesp849-a PMID:21293652

  1. Small interference RNA-mediated gene silencing of human biliverdin reductase, but not that of heme oxygenase-1, attenuates arsenite-mediated induction of the oxygenase and increases apoptosis in 293A kidney cells.

    PubMed

    Miralem, Tihomir; Hu, Zhenbo; Torno, Michael D; Lelli, Katherine M; Maines, Mahin D

    2005-04-29

    BVR reduces biliverdin, the HO-1 and HO-2 product, to bilirubin. Human biliverdin (BVR) is a serine/threonine kinase activated by free radicals. It is a leucine zipper (bZip) DNA-binding protein and a regulatory factor for 8/7-bp AP-1-regulated genes, including HO-1 and ATF-2/CREB. Presently, small interference (si) RNA constructs were used to investigate the role of human BVR in sodium arsenite (As)-mediated induction of HO-1 and in cytoprotection against apoptosis. Activation of BVR involved increased serine/threonine phosphorylation but not its protein or transcript levels. The peak activity at 1 h (4-5-fold) after treatment of 293A cells with 5 mum As preceded induction of HO-1 expression by 3 h. The following suggests BVR involvement in regulating oxidative stress response of HO-1: siBVR attenuated As-mediated increase in HO-1 expression; siBVR, but not siHO-1, inhibited As-dependent increased c-jun promoter activity; treatment of cells with As increased AP-1 binding of nuclear proteins; BVR was identified in the DNA-protein complex; and AP-1 binding of the in vitro translated BVR was phosphorylation-dependent and was attenuated by biliverdin. Most unexpectedly, cells transfected with siBVR, but not siHO-1, displayed a 4-fold increase in apoptotic cells when treated with 10 mum As as detected by flow cytometry. The presence of BVR small interference RNA augmented the effect of As on levels of cytochrome c, TRAIL, and DR-5 mRNA and cleavage of poly(ADP-ribose) polymerase. The findings describe the function of BVR in HO-1 oxidative response and, demonstrate, for the first time, not only that BVR advances the role of HO-1 in cytoprotection but also affords cytoprotection independent of heme degradation. PMID:15741166

  2. Hybrid silencers with micro-perforated panels and internal partitions.

    PubMed

    Yu, Xiang; Cheng, Li; You, Xiangyu

    2015-02-01

    A sub-structuring approach, along with a unit cell treatment, is proposed to model expansion chamber silencers with internal partitions and micro-perforated panels (MPPs) in the absence of internal flow. The side-branch of the silencer is treated as a combination of unit cells connected in series. It is shown that, by connecting multiple unit cells with varying parameters, the noise attenuation bandwidth can be enlarged. With MPPs, the hybrid noise attenuation mechanism of the silencer is revealed. Depending on the size of the perforation hole, noise attenuation can be dominated by dissipative, reactive, or combined effects together. For a broadband sound absorption, the hole size, together with the perforation ratio and other parameters, can be optimized to strike a balance between the dissipative and reactive effect, for ultimately achieving the desired noise attenuation performance within a prescribed frequency region. The modular nature of the proposed formulation allows doing this in a flexible, accurate, and cost effective manner. The accuracy of the proposed approach is validated through comparisons with finite element method and experiments. PMID:25698027

  3. Rethinking the Day of Silence

    ERIC Educational Resources Information Center

    Murphy, Adriana

    2013-01-01

    Back in 2006, 7th and 8th graders at Green Acres, the K-8 independent school where the author taught in suburban Maryland, participated in the Day of Silence. The Day of Silence is a national event: Students across the country take a one-day pledge of silence to show that they want to make schools safe for all students, regardless of their sexual…

  4. Epigenetic silencing in transgenic plants

    PubMed Central

    Rajeevkumar, Sarma; Anunanthini, Pushpanathan; Sathishkumar, Ramalingam

    2015-01-01

    Epigenetic silencing is a natural phenomenon in which the expression of genes is regulated through modifications of DNA, RNA, or histone proteins. It is a mechanism for defending host genomes against the effects of transposable elements and viral infection, and acts as a modulator of expression of duplicated gene family members and as a silencer of transgenes. A major breakthrough in understanding the mechanism of epigenetic silencing was the discovery of silencing in transgenic tobacco plants due to the interaction between two homologous promoters. The molecular mechanism of epigenetic mechanism is highly complicated and it is not completely understood yet. Two different molecular routes have been proposed for this, that is, transcriptional gene silencing, which is associated with heavy methylation of promoter regions and blocks the transcription of transgenes, and post-transcriptional gene silencing (PTGS), the basic mechanism is degradation of the cytosolic mRNA of transgenes or endogenous genes. Undesired transgene silencing is of major concern in the transgenic technologies used in crop improvement. A complete understanding of this phenomenon will be very useful for transgenic applications, where silencing of specific genes is required. The current status of epigenetic silencing in transgenic technology is discussed and summarized in this mini-review. PMID:26442010

  5. Edwin Hubble's Silence

    NASA Astrophysics Data System (ADS)

    Lago, D.

    2013-04-01

    In late 1928 Edwin Hubble was right in the middle of using V. M. Slipher's redshift data to prove that the universe is expanding, when Hubble's boss, George Hale, directed him to drop everything and rush to the Grand Canyon and test it as a possible site for Hale's planned 200-inch telescope. On his way, Hubble stopped at Lowell Observatory and met with V. M. Slipher. The letters both men wrote about this visit suggest that Hubble never said a word about his being in the middle of using Slipher's research to transform the universe. At the least, this silence is symbolic of the silence with which astronomical history has often treated Slipher's work. A survey of the historical literature suggests several reasons for this. Theorists and observers in astronomy (and other sciences) have long had different perspectives about how science works, and those who place more importance on theory have tended to credit the idea of the expanding universe to the theorists. Also, many sources indicate that Edwin Hubble was not a modest man or generous about sharing credit.

  6. The Search for Strategic Silence.

    ERIC Educational Resources Information Center

    Lentz, Richard

    Media content analysts seldom observe the principle that editorial omissions are as telling as what is published or broadcast; hence, the purpose of this paper is to explore, and thus stimulate debate about, editorial omissions or "strategic silence." It is observed that as a concept, strategic silence embraces both tact and strategy--the former…

  7. The rest is silence.

    PubMed Central

    Bernstein, E; Denli, A M; Hannon, G J

    2001-01-01

    Over the past several years, RNAi and its related phenomena have emerged not only as a powerful experimental tool but also as a new mode of gene regulation. Through a combination of genetic and biochemical approaches we have learned much about the mechanisms underlying dsRNA responses. However, many of the most intriguing aspects of dsRNA-induced gene silencing have yet to be illuminated. What has become abundantly clear is that the complex and highly conserved biology underlying RNA interference is critical both for genome maintenance and for the development of complex organisms. However, it seems probable that we have only begun to reveal the diversity of biological roles played by RNAi-related processes. PMID:11720281

  8. Silence Amenity Engineering

    NASA Astrophysics Data System (ADS)

    Fujita, Hajime

    Engineering civilization brought convenient and comfortable life to us. However, some environmental problems such as various pollutions have also been developed with it. Acoustical noise is one of the major problems in modern life. Noise is generated from a noise source and propagates through transmitting medium such as the air and eventually reaches a receiver, usually a human being. The noise problem can be avoided, therefore, if one of those three elements in the noise problem is removed completely. In actual case, engineers are looking for most efficient way combining the controls for these three elements. In this article, basic characteristics of noise is reviewed briefly at first, then sound field analysis to predict sound transmission is discussed Aerodynamic noise is one of the major problems in silence amenity engineering today. Basic concept of the aerodynamic noise generation mechanism is discussed in detail with applications to turbo-machinery and high speed train noise control technology.

  9. The eerie silence

    NASA Astrophysics Data System (ADS)

    Davies, Paul

    2010-03-01

    Whether or not we are alone in the universe is one of the great outstanding questions of existence. For thousands of years it was restricted to the realm of philosophy and theology, but 50 years ago it became part of science. In April 1960 a young US astronomer, Frank Drake, began using a radio telescope to investigate whether signals from an extraterrestrial community might be coming our way. Known as the Search for Extraterrestrial Intelligence, or SETI, it has grown into a major international enterprise, involving scientific institutions in several countries. Apart from a few oddities, however, all that the radio astronomers have encountered is an eerie silence. So is humankind the only technological civilization in the universe after all? Or might we be looking for the wrong thing in the wrong place at the wrong time?

  10. Probabilistic evaluation of initiation time in RC bridge beams with load-induced cracks exposed to de-icing salts

    SciTech Connect

    Lu Zhaohui; Zhao Yangang; Yu Zhiwu; Ding Faxing

    2011-03-15

    In this study, a reliability-based method for predicting the initiation time of reinforced concrete bridge beams with load-induced cracks exposed to de-icing salts is presented. A practical model for predicting the diffusion coefficient of chloride ingress into load-induced cracked concrete is proposed. Probabilistic information about uncertainties related to the surface chloride content and the threshold chloride concentration has been estimated from a wide review of previous experimental or statistical studies. Probabilistic analysis to estimate the time to corrosion initiation with/without considering the effect of the load-induced cracks on the chloride ingress into concrete has been carried out. Results of the analysis demonstrate the importance of considering the effect of the load-induced cracks for correct prediction of corrosion initiation in RC bridge beams exposed to chlorides.

  11. Silence and the Notion of the Commons.

    ERIC Educational Resources Information Center

    Franklin, Ursula

    1994-01-01

    Stresses the value of silence, the right to have silence, and how technology has manipulated the sound environment and therefore taken silence out of common availability. Discusses noise pollution and the manipulative use of sound for private gain. Suggests taking action to restore the right to silence. (LP)

  12. RNA silencing movement in plants.

    PubMed

    Mermigka, Glykeria; Verret, Frédéric; Kalantidis, Kriton

    2016-04-01

    Multicellular organisms, like higher plants, need to coordinate their growth and development and to cope with environmental cues. To achieve this, various signal molecules are transported between neighboring cells and distant organs to control the fate of the recipient cells and organs. RNA silencing produces cell non-autonomous signal molecules that can move over short or long distances leading to the sequence specific silencing of a target gene in a well defined area of cells or throughout the entire plant, respectively. The nature of these signal molecules, the route of silencing spread, and the genes involved in their production, movement and reception are discussed in this review. Additionally, a short section on features of silencing spread in animal models is presented at the end of this review. PMID:26297506

  13. Traffic into silence: endomembranes and post-transcriptional RNA silencing

    PubMed Central

    Kim, Yun Ju; Maizel, Alexis; Chen, Xuemei

    2014-01-01

    microRNAs (miRNAs) and small interfering RNAs (siRNAs) are small RNAs that repress gene expression at the post-transcriptional level in plants and animals. Small RNAs guide Argonaute-containing RNA-induced silencing complexes to target RNAs in a sequence-specific manner, resulting in mRNA deadenylation followed by exonucleolytic decay, mRNA endonucleolytic cleavage, or translational inhibition. Although our knowledge of small RNA biogenesis, turnover, and mechanisms of action has dramatically expanded in the past decade, the subcellular location of small RNA-mediated RNA silencing still needs to be defined. In contrast to the prevalent presumption that RNA silencing occurs in the cytosol, emerging evidence reveals connections between the endomembrane system and small RNA activities in plants and animals. Here, we summarize the work that uncovered this link between small RNAs and endomembrane compartments and present an overview of the involvement of the endomembrane system in various aspects of RNA silencing. We propose that the endomembrane system is an integral component of RNA silencing that has been long overlooked and predict that a marriage between cell biology and RNA biology holds the key to a full understanding of post-transcriptional gene regulation by small RNAs. PMID:24668229

  14. RNA Polymerase I and Fob1 contributions to transcriptional silencing at the yeast rDNA locus.

    PubMed

    Buck, Stephen W; Maqani, Nazif; Matecic, Mirela; Hontz, Robert D; Fine, Ryan D; Li, Mingguang; Smith, Jeffrey S

    2016-07-27

    RNA polymerase II (Pol II)-transcribed genes embedded within the yeast rDNA locus are repressed through a Sir2-dependent process called 'rDNA silencing'. Sir2 is recruited to the rDNA promoter through interactions with RNA polymerase I (Pol I), and to a pair of DNA replication fork block sites (Ter1 and Ter2) through interaction with Fob1. We utilized a reporter gene (mURA3) integrated adjacent to the leftmost rDNA gene to investigate localized Pol I and Fob1 functions in silencing. Silencing was attenuated by loss of Pol I subunits or insertion of an ectopic Pol I terminator within the adjacent rDNA gene. Silencing left of the rDNA array is naturally attenuated by the presence of only one intact Fob1 binding site (Ter2). Repair of the 2nd Fob1 binding site (Ter1) dramatically strengthens silencing such that it is no longer impacted by local Pol I transcription defects. Global loss of Pol I activity, however, negatively affects Fob1 association with the rDNA. Loss of Ter2 almost completely eliminates localized silencing, but is restored by artificially targeting Fob1 or Sir2 as Gal4 DNA binding domain fusions. We conclude that Fob1 and Pol I make independent contributions to establishment of silencing, though Pol I also reinforces Fob1-dependent silencing. PMID:27060141

  15. Ice-load induced tectonics controlled tunnel valley evolution - instances from the southwestern Baltic Sea

    NASA Astrophysics Data System (ADS)

    Al Hseinat, M.; Hübscher, C.

    2014-08-01

    Advancing ice sheets have a strong impact on the earth's topography. For example, they leave behind an erosional unconformity, bulldozer the underlying strata and form tunnel valleys, primarily by subglacial melt-water erosion and secondarily by direct glacial erosion. The conceptual models of the reactivation of faults within the upper crust, due to the ice sheets' load, are also established. However, this phenomenon is also rather under-explored. Here, we propose a causal link between ice-load induced tectonics, the generation of near-vertical faults in the upper crust above an inherited deep-rooted fault and the evolution of tunnel valleys. The Kossau tunnel valley in the southeastern Bay of Kiel has been surveyed by means of high-resolution multi-channel seismic and echosounder data. It strikes almost south to north and can be mapped over a distance of ca 50 km. It is 1200-8000 m wide with a valley of up to 200 m deep. Quaternary deposits fill the valley and cover the adjacent glaciogenic unconformity. A near-vertical fault system with an apparent dip angle of >80°, which reaches from the top Zechstein upwards into the Quaternary, underlies the valley. The fault partially pierces the seafloor and growth is observed within the uppermost Quaternary strata only. Consequently, the fault evolved in the Late Quaternary. The fault is associated with an anticline that is between 700 and 3000 m wide and about 20-40 m high. The fault-anticline assemblage neither resembles any typical extensional, compressional or strike-slip deformation pattern, nor is it related to salt tectonics. Based on the observed position and deformation pattern of the fault-anticline assemblage, we suggest that these structures formed as a consequence of the differential ice-load induced tectonics above an inherited deep-rooted sub-salt fault related to the Glückstadt Graben. Lateral variations in the ice-load during the ice sheet's advance caused differential subsidence, thus rejuvenating the

  16. Communicative Silences: Forms and Functions

    ERIC Educational Resources Information Center

    Bruneau, Thomas J.

    1973-01-01

    The nature of silence is discussed as an imposition of mind, as an interdependent signification ground for speech signs, as a relationship to mental time (as opposed to artificial time), and as it relates to sensation, perception and metaphorical movement. (Author)

  17. Teaching Note: Gaining Voice through Silence

    ERIC Educational Resources Information Center

    Kaufman, Peter

    2008-01-01

    For educators striving to create an egalitarian classroom based on open, reflexive, and honest dialogue, silence can be the ultimate obstacle. Not only does silence stifle individual expression; more importantly, silence also prevents the collective production of knowledge, understanding, compassion, and empathy. When learners and teachers feel…

  18. Silence as the Foundation of Learning

    ERIC Educational Resources Information Center

    Caranfa, Angelo

    2004-01-01

    Past and present discussions on education all too frequently neglect the role that silence plays in learning. In this article I set out to demonstrate that silence is the very foundation of learning. My claim is that we must find ways of freeing silence in our pedagogical practices so that our discourse does not denigrate into mere empty words,…

  19. Is silence killing your company?

    PubMed

    Perlow, Leslie; Williams, Stephanie

    2003-05-01

    Many times, often with the best of intentions, people at work decide it's more productive to remain silent about their differences than to air them. There's no time, they think, or no point in going against what the boss says. But as new research by the authors shows, silencing doesn't smooth things over or make people more productive. It merely pushes differences beneath the surface and can set in motion powerfully destructive forces. When people stay silent about important disagreements, they can begin to fill with anxiety, anger, and resentment. As long as the conflict is unresolved, their repressed feelings remain potent, making them increasingly distrustful, self-protective, and all the more fearful that if they speak up they will be embarrassed or rejected. Their sense of insecurity grows, leading to further acts of silence, more defensiveness, and more distrust, thereby setting into motion a destructive "spiral of silence." Sooner or later, they mentally opt out--sometimes merely doing what they're told but contributing nothing of their own, sometimes spreading discontent and frustration throughout the workplace that can lead them, and others, to leave without thinking it through. These vicious spirals of silence can be replaced with virtuous spirals of communication, but that requires individuals to find the courage to act differently and executives to create the conditions in which people will value the expression of differences. All too often, behind failed products, broken processes, and mistaken career decisions are people who chose to hold their tongues. Breaking the silence can bring an outpouring of fresh ideas from all levels of an organization--ideas that might just raise the organization's performance to a whole new level. PMID:12747162

  20. Load-induced debonding of FRP composites applied to reinforced concrete

    NASA Astrophysics Data System (ADS)

    Blok, Joel; Brown, Jeff

    2009-05-01

    Fiber-reinforced polymer (FRP) composites are widely used to increase the flexural and shear capacity of reinforced concrete (RC) elements. One potential disadvantage is that strengthened surfaces are no longer visible and cracks or delaminations that result from excessive loading or fatigue may go undetected. This research investigated thermal imaging techniques for monitoring and evaluating load-induced delamination of FRP composites applied to small scale RC beams. Two beams (3.5 in x 4.5 in x 58 in) were loaded monotonically to failure. Infrared thermography (IRT) inspections were performed at various load levels through failure using a composite phase imaging technique. Two similar beams were tested in fatigue and periodic IRT inspections were performed at 50,000-cycle intervals. Individual phase values for each pixel were designated as "well-bonded", "suspect" or "unbonded" to indicate the quality of FRP bond. Suspect areas included regions of excess thickened-epoxy tack-coat and smaller installation defects in the unloaded specimens. The long-term objective of this research is to develop a practical framework for conducting quantitative IRT inspections of FRP composites applied to RC and incorporating these results into acceptance criteria for new installations and predictions for the remaining service life of in-service FRP systems. This method may also offer insight into the necessity for repairs to in-service systems.

  1. Loading-Induced Heat-Shock Response in Bovine Intervertebral Disc Organ Culture.

    PubMed

    Chooi, Wai Hon; Chan, Samantha Chun Wai; Gantenbein, Benjamin; Chan, Barbara Pui

    2016-01-01

    Mechanical loading has been shown to affect cell viability and matrix maintenance in the intervertebral disc (IVD) but there is no investigation on how cells survive mechanical stress and whether the IVD cells perceive mechanical loading as stress and respond by expression of heat shock proteins. This study investigates the stress response in the IVD in response to compressive loading. Bovine caudal disc organ culture was used to study the effect of physiological range static loading and dynamic loading. Cell activity, gene expression and immunofluorescence staining were used to analyze the cell response. Cell activity and cytoskeleton of the cells did not change significantly after loading. In gene expression analysis, significant up-regulation of heat shock protein-70 (HSP70) was observed in nucleus pulposus after two hours of loading. However, the expression of the matrix remodeling genes did not change significantly after loading. Similarly, expressions of stress response and matrix remodeling genes changed with application and removal of the dynamic loading. The results suggest that stress response was induced by physiological range loading without significantly changing cell activity and upregulating matrix remodeling. This study provides direct evidence on loading induced stress response in IVD cells and contributes to our understanding in the mechanoregulation of intervertebral disc cells. PMID:27580124

  2. Transcriptional gene silencing in humans.

    PubMed

    Weinberg, Marc S; Morris, Kevin V

    2016-08-19

    It has been over a decade since the first observation that small non-coding RNAs can functionally modulate epigenetic states in human cells to achieve functional transcriptional gene silencing (TGS). TGS is mechanistically distinct from the RNA interference (RNAi) gene-silencing pathway. TGS can result in long-term stable epigenetic modifications to gene expression that can be passed on to daughter cells during cell division, whereas RNAi does not. Early studies of TGS have been largely overlooked, overshadowed by subsequent discoveries of small RNA-directed post-TGS and RNAi. A reappraisal of early work has been brought about by recent findings in human cells where endogenous long non-coding RNAs function to regulate the epigenome. There are distinct and common overlaps between the proteins involved in small and long non-coding RNA transcriptional regulatory mechanisms, suggesting that the early studies using small non-coding RNAs to modulate transcription were making use of a previously unrecognized endogenous mechanism of RNA-directed gene regulation. Here we review how non-coding RNA plays a role in regulation of transcription and epigenetic gene silencing in human cells by revisiting these earlier studies and the mechanistic insights gained to date. We also provide a list of mammalian genes that have been shown to be transcriptionally regulated by non-coding RNAs. Lastly, we explore how TGS may serve as the basis for development of future therapeutic agents. PMID:27060137

  3. [A gun silencer of a special kind].

    PubMed

    Schyma, C; Schyma, P; Milbradt, H

    2000-01-01

    The authors report about a small bore pistol with silencer. In addition, the silencer can be sealed up by a rubber plate. This leads to a false estimation of the shooting distance. Also at close range shots the shot with the silencer and the rubber seal leaves hardly gunshot residues. The bullet wipe persists but microradiography shows his changed morphological composition. The use of the rubber seal leads predominantly to atypical bullet holes. PMID:10829239

  4. Silence Is Consent, or Curse Ye Meroz!

    ERIC Educational Resources Information Center

    Levin, Richard

    1997-01-01

    Examines assumptions of "oppositional" literary criticism, namely the assumption that older-style "objective" literary criticism must, in its political silence, be supportive of dominant ideologies. (TB)

  5. [E. M. Jellinek's silenced and silencing transgenerational story].

    PubMed

    Kelemen, Gábor; Márk, Mónika

    2013-01-01

    Jellinek is a kind of archetypal character for future generations in the field of addiction studies. His implosion in the arena of alcoholism around the age of 50 was an unexpected challenge to medical science. We know very little about his own role models giving an intellectual and moral compass to his pragmatic creativity. More than 30 years has passed since Jellinek's death when an American sociologist Ron Roizen started unearthing his silent story. Roizen discerned that there are a lot of unsaid and muted issues in his personal Hungarian past. Our paper, based on the authors' research in Hungarian archives and other sources reveals that not just Jellinek's personal but his transgenerational narrative has been not-yet-said. This silenced and silencing history appears an unfinished business of acculturation of the family, which started prior to four generations. Authors have been concluding that the issue of religious conversion is a critical point in the process of acculturation. They examine the counter move of loyalty to family values and driving force of assimilation making their story unspeakable. PMID:24443572

  6. "The Silence Itself Is Enough of a Statement": The Day of Silence and LGBTQ Awareness Raising

    ERIC Educational Resources Information Center

    Woolley, Susan W.

    2012-01-01

    This ethnographic study of a high school gay-straight alliance club examines unintended consequences of silence during the Day of Silence, a day of action aimed at addressing anti-LGBTQ bias in schools. While this strategy calls for students to engage in intentional silences to raise awareness of anti-LGBTQ bias, it does not necessarily lead…

  7. Quantification of Crack Interaction in Loading-Induced Rock Deformation Tests

    NASA Astrophysics Data System (ADS)

    Reyes-Montes, J. M.; Goodfellow, S. D.; Nasseri, M. H.; Young, R.

    2013-12-01

    The coalescence of microcracks into major fractures marks the onset of major damage in rock and induces significant changes in its geotechnical and sealing properties. The interaction between neighbouring fractures affects the deformation process leading to this major failure. This study presents the analysis of load-induced fracture using spatial and temporal patterns of recorded acoustic emission (AE) events to investigate the role of crack interaction during fracturing processes Two methods based on event separation are used to estimate correlation and interaction distances between events. The first method uses the spacing of sequential and non-sequential event pairs, thus, the distance bin where both distributions converge can be interpreted as an upper bound for the interaction distance of correlated events in each of the experiments. The second method compares the distribution of inter-event separation of recorded events with the distribution obtained for a random population of events, quantifying the divergence between both distributions and its variation with time. A positive deviation for a distance range including up to the interaction distance observed in the previous approach indicates spatial clustering and process dominated by interactive events. This deviation is used to quantify the degree of event interaction or degree of non-randomness (NR) during the process. Two load-stress induced fracturing tests, carried out at two different scales, are analysed in this study to investigate the role of crack interaction in the deformation and fracturing process. The first test registered a total of 15,198 AE events from a pillar created between two deposition holes at SKB's Aspö Pillar Stability Experiment. The results show a good correlation between maxima of NR and minima in b-value in the time period immediately preceding and during the development of the observed major damage in form of spalling. The result can be interpreted as an indication of spalling

  8. Systemic silencing signal(s).

    PubMed

    Fagard, M; Vaucheret, H

    2000-06-01

    Grafting experiments have revealed that transgenic plants that undergo co-suppression of homologous transgenes and endogenous genes or PTGS of exogenous transgenes produce a sequence-specific systemic silencing signal that is able to propagate from cell to cell and at long distance. Similarly, infection of transgenic plants by viruses that carry (part of) a transgene sequence results in global silencing (VIGS) of the integrated transgenes although viral infection is localized. Systemic PTGS and VIGS strongly resemble recovery from virus infection in non-transgenic plants, leading to protection against secondary infection in newly emerging leaves and PTGS of transiently expressed homologous transgenes. The sequence-specific PTGS signal is probably a transgene product (for example, aberrant RNA) or a secondary product (for example, RNA molecules produced by an RNA-dependent RNA polymerase with transgene RNA as a matrix) that mimics the type of viral RNA that is targeted for degradation by cellular defence. Whether some particular cases of transgene TGS could also rely on the production of such a mobile molecule is discussed. PMID:10999411

  9. SILENCING POLYGALACTURONASE EXPRESSION INHIBITS TOMATO PETIOLE ABSCISSION

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We used Virus Induced Gene Silencing (VIGS) as a tool for functional analysis of cell-wall associated genes that have been suggested to be involved in leaf abscission. Tobacco rattle virus (TRV) is an effective vector for VIGS in tomato (Lycopersicon esculentum). Silencing was more efficient when ...

  10. Classroom Silence: Voices from Japanese EFL Learners

    ERIC Educational Resources Information Center

    Harumi, Seiko

    2011-01-01

    This article explores Japanese EFL learners' classroom silence in a Japanese EFL context. The existence of silence in second language learning contexts can be a source of conflict between students and teachers and even among students themselves. It can also be an obstacle to acquiring the target language. In order to tackle this problem and to…

  11. Venturis as silencers in a BOF stack

    SciTech Connect

    Kelsall, T.; Gerritsen, T.; Landon, T.

    1995-06-01

    Installation of a venture in a BOF stack reduced the sound level in a local community by 10 to 15 db. This application resulted in the development of a new type of fan silencer, called the modal silencer, having the inherent advantages of low pressure drop and less maintenance compared with conventional types.

  12. Low temperature inhibits RNA silencing-mediated defence by the control of siRNA generation

    PubMed Central

    Szittya, György; Silhavy, Dániel; Molnár, Attila; Havelda, Zoltán; Lovas, Ágnes; Lakatos, Lóránt; Bánfalvi, Zsófia; Burgyán, József

    2003-01-01

    Temperature dramatically affects plant–virus interactions. Outbreaks of virus diseases are frequently associated with low temperature, while at high temperature viral symptoms are often attenuated (heat masking) and plants rapidly recover from virus diseases. However, the underlying mechanisms of these well-known observations are not yet understood. RNA silencing is a conserved defence system of eukaryotic cells, which operates against molecular parasites including viruses and transgenes. Here we show that at low temperature both virus and transgene triggered RNA silencing are inhibited. Therefore, in cold, plants become more susceptible to viruses, and RNA silencing-based phenotypes of transgenic plants are lost. Consistently, the levels of virus- and transgene-derived small (21–26 nucleotide) interfering (si) RNAs—the central molecules of RNA silencing-mediated defence pathways—are dramatically reduced at low temperature. In contrast, RNA silencing was activated and the amount of siRNAs gradually increased with rising temperature. However, temperature does not influence the accumulation of micro (mi) RNAs, which play a role in developmental regulation, suggesting that the two classes of small (si and mi) RNAs are generated by different nuclease complexes. PMID:12554663

  13. Inhibition of IκB phosphorylation prevents load-induced cardiac dysfunction in mice.

    PubMed

    Tanaka, Tetsu; Ogawa, Masahito; Suzuki, Jun-ichi; Sekinishi, Asuka; Itai, Akiko; Hirata, Yasunobu; Nagai, Ryozo; Isobe, Mitsuaki

    2012-12-15

    Pressure overload is known to be a cause of cardiac hypertrophy that often transits to heart failure. Although nuclear factor (NF)-κB is a key factor in the progression of cardiac hypertrophy, its pathophysiology is yet to be elucidated. Thus, we aimed to show that inhibition of NF-κB activation improves pressure overload-induced cardiac dysfunction. To assess the effect of inhibition on NF-κB activation in pressure overload cardiac hypertrophy, we used IMD-1041 in a murine thoracic aortic constriction (TAC) model. IMD-1041 inhibits the phosphorylation of IκB via inhibition of IκB kinase-β. IMD-1041 (100 mg·kg(-1)·day(-1)) or vehicle was administered orally into mice once a day, and mice were euthanized on day 42 after TAC. TAC resulted in left ventricular wall thickening, cardiac dysfunction, and increases of heart and lung weight, whereas IMD-1041 significantly suppressed the development of cardiac hypertropy 6 wk after TAC. Histologically, developed cardiac fibrosis and cardiomyocyte hypertrophy occurred in the vehicle-treated group, whereas IMD-1041 significantly attenuated these changes. IMD-1041 suppressed the expression of p65-positive cells and nuclear translocation of p65 induced by TAC compared with vehicle. Matrix metalloproteinase-2 activity increased in the vehicle + TAC-treated group; however, it was suppressed in the IMD-1041 + TAC-treated group. IMD-1041 treatment from day 28 to day 42 after TAC significantly attenuated the decrease in the percentage of fractional shortening and cardiac fibrosis without an antihypertrophic effect. In conclusion, IMD-1041 may be useful for preventing pressure overload-induced cardiac dysfunction and the transition of cardiac hypertrophy to contraction failure via suppression of NF-κB activation. PMID:23042949

  14. Mammalian meiotic silencing exhibits sexually dimorphic features.

    PubMed

    Cloutier, J M; Mahadevaiah, S K; ElInati, E; Tóth, A; Turner, James

    2016-06-01

    During mammalian meiotic prophase I, surveillance mechanisms exist to ensure that germ cells with defective synapsis or recombination are eliminated, thereby preventing the generation of aneuploid gametes and embryos. Meiosis in females is more error-prone than in males, and this is in part because the prophase I surveillance mechanisms are less efficient in females. A mechanistic understanding of this sexual dimorphism is currently lacking. In both sexes, asynapsed chromosomes are transcriptionally inactivated by ATR-dependent phosphorylation of histone H2AFX. This process, termed meiotic silencing, has been proposed to perform an important prophase I surveillance role. While the transcriptional effects of meiotic silencing at individual genes are well described in the male germ line, analogous studies in the female germ line have not been performed. Here we apply single- and multigene RNA fluorescence in situ hybridization (RNA FISH) to oocytes from chromosomally abnormal mouse models to uncover potential sex differences in the silencing response. Notably, we find that meiotic silencing in females is less efficient than in males. Within individual oocytes, genes located on the same asynapsed chromosome are silenced to differing extents, thereby generating mosaicism in gene expression profiles across oocyte populations. Analysis of sex-reversed XY female mice reveals that the sexual dimorphism in silencing is determined by gonadal sex rather than sex chromosome constitution. We propose that sex differences in meiotic silencing impact on the sexually dimorphic prophase I response to asynapsis. PMID:26712235

  15. The gifts of silence and solitude.

    PubMed

    Schmidt Bunkers, Sandra

    2008-01-01

    In this column the author describes the importance of finding silence and solitude amid the noise and technology present today in the teaching-learning academy. Three gifts of silence and solitude are identified: the gift of comforting aloneness, the gift of vision for new horizons, and the gift of a sense of freedom. A humanbecoming perspective is used to explore the implications of these gifts. This column introduces a column by Diana Vander Woude describing her teaching-learning experience in leadership focusing on silence and solitude. PMID:18096981

  16. Rotary antenna attenuator

    NASA Technical Reports Server (NTRS)

    Dickinson, R. M.; Hardy, J. C.

    1969-01-01

    Radio frequency attenuator, having negligible insertion loss at minimum attenuation, can be used for making precise antenna gain measurements. It is small in size compared to a rotary-vane attenuator.

  17. Tomato mosaic virus replication protein suppresses virus-targeted posttranscriptional gene silencing.

    PubMed

    Kubota, Kenji; Tsuda, Shinya; Tamai, Atsushi; Meshi, Tetsuo

    2003-10-01

    Posttranscriptional gene silencing (PTGS), a homology-dependent RNA degradation system, has a role in defending against virus infection in plants, but plant viruses encode a suppressor to combat PTGS. Using transgenic tobacco in which the expression of green fluorescent protein (GFP) is posttranscriptionally silenced, we investigated a tomato mosaic virus (ToMV)-encoded PTGS suppressor. Infection with wild-type ToMV (L strain) interrupted GFP silencing in tobacco, coincident with visible symptoms, whereas some attenuated strains of ToMV (L(11) and L(11)A strains) failed to suppress GFP silencing. Analyses of recombinant viruses containing the L and L(11)A strains revealed that a single base change in the replicase gene, which causes an amino acid substitution, is responsible for the symptomless and suppressor-defective phenotypes of the attenuated strains. An agroinfiltration assay indicated that the 130K replication protein acts as a PTGS suppressor. Small interfering RNAs (siRNAs) of 21 to 25 nucleotides accumulated during ToMV infection, suggesting that the major target of the ToMV-encoded suppressor is downstream from the production of siRNAs in the PTGS pathway. Analysis with GFP-tagged recombinant viruses revealed that the suppressor inhibits the establishment of the ToMV-targeted PTGS system in the inoculated leaves but does not detectably suppress the activity of the preexisting, sequence-specific PTGS machinery there. Taken together, these results indicate that it is likely that the ToMV-encoded suppressor, the 130K replication protein, blocks the utilization of silencing-associated small RNAs, so that a homology-dependent RNA degradation machinery is not newly formed. PMID:14512550

  18. Gene silencing of TACE enhances plaque stability and improves vascular remodeling in a rabbit model of atherosclerosis

    PubMed Central

    Zhao, Xueqiang; Kong, Jing; Zhao, Yuxia; Wang, Xuping; Bu, Peili; Zhang, Cheng; Zhang, Yun

    2015-01-01

    We aimed to test the hypothesis that gene silencing of tumor necrosis factor alpha converting enzyme (TACE) may attenuate lesion inflammation and positive vascular remodeling and enhance plaque stability in a rabbit model of atherosclerosis. Lentivirus-mediated TACE shRNA was injected into the abdominal aortic plaques of rabbits which effectively down-regulated TACE expression and activities from week 8 to week 16. TACE gene silencing reduced remodeling index and plaque burden, and diminished the content of macrophages and lipids while increased that of smooth muscle cells and collagen in the aortic plaques. In addition, TACE gene silencing attenuated the local expression of P65, iNOS, ICAM-1, VEGF and Flt-1 and activities of MMP9 and MMP2 while increased the local expression of TGF-β1 together with reduced number of neovessels in the aorta. TACE shRNA treatment resulted in down-regulated expression of TACE in macrophages and blunted ERK-P38 phosphorylation and tube formation of co-cultured mouse vascular smooth muscle cells or human umbilical vein endothelial cells. In conclusion, gene silencing of TACE enhanced plaque stability and improved vascular positive remodeling. The mechanisms may involve attenuated local inflammation, neovascularization and MMP activation, as well as enhanced collagen production probably via down-regulated ERK-NF-κB and up-regulated TGF-β1 signaling pathways. PMID:26655882

  19. Replication and transcription. Silence of the ORCs.

    PubMed

    Kelly, T J; Jallepalli, P V; Clyne, R K

    1994-03-01

    The origin recognition complex, a multi-protein complex known to bind to replication origins, has now been implicated in transcriptional silencing, providing another link between DNA replication and transcription. PMID:7857395

  20. Common themes in mechanisms of gene silencing.

    PubMed

    Moazed, D

    2001-09-01

    The assembly of DNA into regions of inaccessible chromatin, called silent chromatin, is involved in the regulation of gene expression and maintenance of chromosome stability in eukaryotes. Recent studies on Sir2-containing silencing complexes in budding yeast and HP1- and Swi6-containing silencing complexes in metazoans and fission yeast suggest a common mechanism for the assembly of these domains, which involves the physical coupling of histone modifying enzymes to histone binding proteins. PMID:11583612

  1. Design of steam silencers for geothermal applications

    SciTech Connect

    Lazalde-Crabtree, H.

    1985-01-01

    Steam silencers are a means of reducing the loud noise caused by venting steam into the atmosphere as a consequence of load-reductions in a geothermal power plant. For new plants, or for those in which noise measurements cannot be made, an analytical method is given to determine the unsilenced noise levels. Designs fo two types of steam silencers, based on experimental work and theoretical considerations, are presented.

  2. Titration and hysteresis in epigenetic chromatin silencing

    NASA Astrophysics Data System (ADS)

    Dayarian, Adel; Sengupta, Anirvan M.

    2013-06-01

    Epigenetic mechanisms of silencing via heritable chromatin modifications play a major role in gene regulation and cell fate specification. We consider a model of epigenetic chromatin silencing in budding yeast and study the bifurcation diagram and characterize the bistable and the monostable regimes. The main focus of this paper is to examine how the perturbations altering the activity of histone modifying enzymes affect the epigenetic states. We analyze the implications of having the total number of silencing proteins, given by the sum of proteins bound to the nucleosomes and the ones available in the ambient, to be constant. This constraint couples different regions of chromatin through the shared reservoir of ambient silencing proteins. We show that the response of the system to perturbations depends dramatically on the titration effect caused by the above constraint. In particular, for a certain range of overall abundance of silencing proteins, the hysteresis loop changes qualitatively with certain jump replaced by continuous merger of different states. In addition, we find a nonmonotonic dependence of gene expression on the rate of histone deacetylation activity of Sir2. We discuss how these qualitative predictions of our model could be compared with experimental studies of the yeast system under anti-silencing drugs.

  3. Satellite RNAs interfere with the function of viral RNA silencing suppressors

    PubMed Central

    Shen, Wan-Xia; Au, Phil Chi Khang; Shi, Bu-Jun; Smith, Neil A.; Dennis, Elizabeth S.; Guo, Hui-Shan; Zhou, Chang-Yong; Wang, Ming-Bo

    2015-01-01

    Viral satellite RNAs (satRNAs) are small subviral RNAs and depend on the helper virus for replication and spread. satRNAs can attenuate helper virus-induced symptoms, the mechanism of which remains unclear. Here, we show that two virus-encoded suppressors of RNA silencing (VSRs), Cucumber mosaic virus (CMV) 2b and Tombusvirus P19, suppress hairpin RNA (hpRNA)-induced silencing of a β-glucuronidase (GUS) gene in Nicotiana benthamiana. This suppression can be overcome by CMV Y-satellite RNA (Y-Sat) via the Y-Sat-derived small interfering RNAs (siRNAs), which bind to the VSRs and displace the bound hpGUS-derived siRNAs. We also show that microRNA target gene expression in N. tabacum was elevated by CMV infection, presumably due to function of the 2b VSR, but this upregulation of microRNA target genes was reversed in the presence of Y-Sat. These results suggest that satRNA infection minimizes the effect of VSRs on host siRNA and microRNA-directed silencing. Our results suggest that the high abundance of satRNA-derived siRNAs contributes to symptom attenuation by binding helper virus-encoded VSRs, minimizing the capacity of the VSRs to bind host siRNA and miRNA and interfere with their function. PMID:25964791

  4. Rescue of Mtp siRNA-induced hepatic steatosis by DGAT2 siRNA silencing.

    PubMed

    Tep, Samnang; Mihaila, Radu; Freeman, Alexander; Pickering, Victoria; Huynh, Felicia; Huyhn, Felicia; Tadin-Strapps, Marija; Stracks, Allison; Hubbard, Brian; Caldwell, Jeremy; Flanagan, W Michael; Kuklin, Nelly A; Ason, Brandon

    2012-05-01

    Microsomal triglyceride transfer protein (Mtp) inhibitors represent a novel therapeutic approach to lower circulating LDL cholesterol, although therapeutic development has been hindered by the observed increase in hepatic triglycerides and liver steatosis following treatment. Here, we used small interfering RNAs (siRNA) targeting Mtp to achieve target-specific silencing to study this phenomenon and to determine to what extent liver steatosis is induced by changes in Mtp expression. We observed that Mtp silencing led to a decrease in many genes involved in hepatic triglyceride synthesis. Given the role of diacylglycerol O-acyltransferase 2 (Dgat2) in regulating hepatic triglyceride synthesis, we then evaluated whether target-specific silencing of both Dgat2 and Mtp were sufficient to attenuate Mtp silencing-induced liver steatosis. We showed that the simultaneous inhibition of Dgat2 and Mtp led to a decrease in plasma cholesterol and a reduction in the accumulation of hepatic triglycerides caused by the inhibition of Mtp. Collectively, these findings provide a proof-of-principle for a triglyceride synthesis/Mtp inhibitor combination and represent a potentially novel approach for therapeutic development in which targeting multiple pathways can achieve the desired response. PMID:22355095

  5. Bicc1 Polymerization Regulates the Localization and Silencing of Bound mRNA

    PubMed Central

    Rothé, Benjamin; Leal-Esteban, Lucia; Bernet, Florian; Urfer, Séverine; Doerr, Nicholas; Weimbs, Thomas; Iwaszkiewicz, Justyna

    2015-01-01

    Loss of the RNA-binding protein Bicaudal-C (Bicc1) provokes renal and pancreatic cysts as well as ectopic Wnt/β-catenin signaling during visceral left-right patterning. Renal cysts are linked to defective silencing of Bicc1 target mRNAs, including adenylate cyclase 6 (AC6). RNA binding of Bicc1 is mediated by N-terminal KH domains, whereas a C-terminal sterile alpha motif (SAM) self-polymerizes in vitro and localizes Bicc1 in cytoplasmic foci in vivo. To assess a role for multimerization in silencing, we conducted structure modeling and then mutated the SAM domain residues which in this model were predicted to polymerize Bicc1 in a left-handed helix. We show that a SAM-SAM interface concentrates Bicc1 in cytoplasmic clusters to specifically localize and silence bound mRNA. In addition, defective polymerization decreases Bicc1 stability and thus indirectly attenuates inhibition of Dishevelled 2 in the Wnt/β-catenin pathway. Importantly, aberrant C-terminal extension of the SAM domain in bpk mutant Bicc1 phenocopied these defects. We conclude that polymerization is a novel disease-relevant mechanism both to stabilize Bicc1 and to present associated mRNAs in specific silencing platforms. PMID:26217012

  6. DC attenuation meter

    DOEpatents

    Hargrove, Douglas L.

    2004-09-14

    A portable, hand-held meter used to measure direct current (DC) attenuation in low impedance electrical signal cables and signal attenuators. A DC voltage is applied to the signal input of the cable and feedback to the control circuit through the signal cable and attenuators. The control circuit adjusts the applied voltage to the cable until the feedback voltage equals the reference voltage. The "units" of applied voltage required at the cable input is the system attenuation value of the cable and attenuators, which makes this meter unique. The meter may be used to calibrate data signal cables, attenuators, and cable-attenuator assemblies.

  7. Epigenetic chromatin silencing: bistability and front propagation

    NASA Astrophysics Data System (ADS)

    Sedighi, Mohammad; Sengupta, Anirvan M.

    2007-12-01

    The role of post-translational modification of histones in eukaryotic gene regulation is well recognized. Epigenetic silencing of genes via heritable chromatin modifications plays a major role in cell fate specification in higher organisms. We formulate a coarse-grained model of chromatin silencing in yeast and study the conditions under which the system becomes bistable, allowing for different epigenetic states. We also study the dynamics of the boundary between the two locally stable states of chromatin: silenced and unsilenced. The model could be of use in guiding the discussion on chromatin silencing in general. In the context of silencing in budding yeast, it helps us understand the phenotype of various mutants, some of which may be non-trivial to see without the help of a mathematical model. One such example is a mutation that reduces the rate of background acetylation of particular histone side chains that competes with the deacetylation by Sir2p. The resulting negative feedback due to a Sir protein depletion effect gives rise to interesting counter-intuitive consequences. Our mathematical analysis brings forth the different dynamical behaviors possible within the same molecular model and guides the formulation of more refined hypotheses that could be addressed experimentally.

  8. "Stop the noise!" From voice to silence.

    PubMed

    Newton, Lorelei; Storch, Janet L; Makaroff, Kara Schick; Pauly, Bernadette

    2012-03-01

    Nurses are frequently portrayed in the literature as being silent about ethical concerns that arise in their practice. This silence is often represented as a lack of voice. However, in our study, we found that nurses who responded to questions about moral distress were not so much silent as silenced. These nurses were enacting their moral agency by engaging in diverse, multiple and time-consuming actions in response to situations identified as morally distressing with families, colleagues, physicians, educators or managers. In many situations, they took action by contacting other healthcare team members, making referrals and coordinating care with other departments such as home care and hospice, as well as initiating contact with groups such as professional regulatory bodies or unions. Examining the relationship between ethical climate, moral distress and voice offers insights into both the meaning and impact of being silenced in the workplace. PMID:22469764

  9. Grafting on a Non-Transgenic Tolerant Tomato Variety Confers Resistance to the Infection of a Sw5-Breaking Strain of Tomato spotted wilt virus via RNA Silencing

    PubMed Central

    Spanò, Roberta; Mascia, Tiziana; Kormelink, Richard; Gallitelli, Donato

    2015-01-01

    RNA silencing controls endogenous gene expression and drives defensive reactions against invasive nucleic acids like viruses. In plants, it has been demonstrated that RNA silencing can be transmitted through grafting between scions and silenced rootstocks to attenuate virus and viroid accumulation in the scions. This has been obtained mostly using transgenic plants, which may be a drawback in current agriculture. In the present study, we examined the dynamics of infection of a resistance-breaking strain of Tomato spotted wilt virus (RB-TSWV) through the graft between an old Apulian (southern Italy) tomato variety, denoted Sl-Ma, used as a rootstock and commercial tomato varieties used as scions. In tests with non-grafted plants, Sl-Ma showed resistance to the RB-TSWV infection as viral RNA accumulated at low levels and plants recovered from disease symptoms by 21 days post inoculation. The resistance trait was transmitted to the otherwise highly susceptible tomato genotypes grafted onto Sl-Ma. The results from the analysis of small RNAs hallmark genes involved in RNA silencing and virus-induced gene silencing suggest that RNA silencing is involved in the resistance showed by Sl-Ma against RB-TSWV and in scions grafted on this rootstock. The results from self-grafted susceptible tomato varieties suggest also that RNA silencing is enhanced by the graft itself. We can foresee interesting practical implications of the approach described in this paper. PMID:26496695

  10. Grafting on a Non-Transgenic Tolerant Tomato Variety Confers Resistance to the Infection of a Sw5-Breaking Strain of Tomato spotted wilt virus via RNA Silencing.

    PubMed

    Spanò, Roberta; Mascia, Tiziana; Kormelink, Richard; Gallitelli, Donato

    2015-01-01

    RNA silencing controls endogenous gene expression and drives defensive reactions against invasive nucleic acids like viruses. In plants, it has been demonstrated that RNA silencing can be transmitted through grafting between scions and silenced rootstocks to attenuate virus and viroid accumulation in the scions. This has been obtained mostly using transgenic plants, which may be a drawback in current agriculture. In the present study, we examined the dynamics of infection of a resistance-breaking strain of Tomato spotted wilt virus (RB-TSWV) through the graft between an old Apulian (southern Italy) tomato variety, denoted Sl-Ma, used as a rootstock and commercial tomato varieties used as scions. In tests with non-grafted plants, Sl-Ma showed resistance to the RB-TSWV infection as viral RNA accumulated at low levels and plants recovered from disease symptoms by 21 days post inoculation. The resistance trait was transmitted to the otherwise highly susceptible tomato genotypes grafted onto Sl-Ma. The results from the analysis of small RNAs hallmark genes involved in RNA silencing and virus-induced gene silencing suggest that RNA silencing is involved in the resistance showed by Sl-Ma against RB-TSWV and in scions grafted on this rootstock. The results from self-grafted susceptible tomato varieties suggest also that RNA silencing is enhanced by the graft itself. We can foresee interesting practical implications of the approach described in this paper. PMID:26496695

  11. Chemogenetic silencing of the midline and intralaminar thalamus blocks amygdala-kindled seizures.

    PubMed

    Wicker, Evan; Forcelli, Patrick A

    2016-09-01

    Temporal lobe epilepsy is the most common form of medically-intractable epilepsy. While seizures in TLE originate in structures such as hippocampus, amygdala, and temporal cortex, they propagate through a crucial relay: the midline/intralaminar thalamus. Prior studies have shown that pharmacological inhibition of midline thalamus attenuates limbic seizures. Here, we examined a recently developed technology, Designer Receptors Exclusively Activated by Designer Drugs (DREADDs), as a means of chemogenetic silencing to attenuate limbic seizures. Adult, male rats were electrically kindled from the amygdala, and injected with virus coding for inhibitory (hM4Di) DREADDs into the midline/intralaminar thalamus. When treated with the otherwise inert ligand Clozapine-N-Oxide (CNO) at doses of 2.5, 5, and 10mg/kg, electrographic and behavioral seizure manifestations were suppressed in comparison to vehicle. At higher doses, we found complete blockade of seizure activity in a subset of subjects. CNO displayed a sharp time-response profile, with significant seizure attenuation seen 20-30min post injection, in comparison to 10 and 40min post injection. Seizures in animals injected with a control vector (i.e., no DREADD) were unaffected by CNO administration. These data underscore the crucial role of the midline/intralaminar thalamus in the propagation of seizures, specifically in the amygdala kindling model, and provide validation of chemogenetic silencing of limbic seizures. PMID:27404844

  12. Nanovector-based prolyl hydroxylase domain 2 silencing system enhances the efficiency of stem cell transplantation for infarcted myocardium repair

    PubMed Central

    Zhu, Kai; Lai, Hao; Guo, Changfa; Li, Jun; Wang, Yulin; Wang, Lingyan; Wang, Chunsheng

    2014-01-01

    Mesenchymal stem cell (MSC) transplantation has attracted much attention in myocardial infarction therapy. One of the limitations is the poor survival of grafted cells in the ischemic microenvironment. Small interfering RNA-mediated prolyl hydroxylase domain protein 2 (PHD2) silencing in MSCs holds tremendous potential to enhance their survival and paracrine effect after transplantation. However, an efficient and biocompatible PHD2 silencing system for clinical application is lacking. Herein, we developed a novel PHD2 silencing system based on arginine-terminated generation 4 poly(amidoamine) (Arg-G4) nanoparticles. The system exhibited effective and biocompatible small interfering RNA delivery and PHD2 silencing in MSCs in vitro. After genetically modified MSC transplantation in myocardial infarction models, MSC survival and paracrine function of IGF-1 were enhanced significantly in vivo. As a result, we observed decreased cardiomyocyte apoptosis, scar size, and interstitial fibrosis, and increased angiogenesis in the diseased myocardium, which ultimately attenuated ventricular remodeling and improved heart function. This work demonstrated that an Arg-G4 nanovector-based PHD2 silencing system could enhance the efficiency of MSC transplantation for infarcted myocardium repair. PMID:25429216

  13. Embracing Silence and the Emptiness between Unspoken Words

    ERIC Educational Resources Information Center

    VanSlyke-Briggs, Kjersti

    2014-01-01

    This article examines the use of silence as a constructive teaching tool in the classroom rather than as a punitive measure. The author offers suggestions for the inclusion of silence to benefit students specifically in a literature high school classroom.

  14. Pressure surge attenuator

    DOEpatents

    Christie, Alan M.; Snyder, Kurt I.

    1985-01-01

    A pressure surge attenuation system for pipes having a fluted region opposite crushable metal foam. As adapted for nuclear reactor vessels and heads, crushable metal foam is disposed to attenuate pressure surges.

  15. Effect of flow on the drumlike silencer

    NASA Astrophysics Data System (ADS)

    Choy, Y. S.; Huang, Lixi

    2005-11-01

    This study examines the effects of a mean flow and turbulent flow excitation on the performance of the recently conceived device which was tested under the no-flow condition [J. Acoust. Soc. Am. 112, 2014-2035 (2002)]. The silencer consists of two cavity-backed membranes lining part of the duct walls. When a certain optimal tension is applied, the silencer gives a broad stopband in the low-frequency regime. Similar performance is predicted for the condition with a mean flow, and tests conducted for flow speeds from 5 to 15 m/s validated these predictions. The spectrum of transmission loss without flow features three resonance peaks, and the mean flow is found to smooth out all peaks and shift two of them through cross-modal coupling. The silencer was tested in a wind tunnel, and no flow induced flexural instability was found on the membrane in the range of flow speeds tested. Insertion loss measurement was also conducted in a natural ventilation condition where a turbulence intensity of 3% was recorded, and the results were close to those without flow. It is concluded that no noticeable extra sound is produced by the turbulent excitation of the membrane under the optimal tension required by the silencer.

  16. Applying gene silencing technology to contraception.

    PubMed

    Dissen, G A; Lomniczi, A; Boudreau, R L; Chen, Y H; Davidson, B L; Ojeda, S R

    2012-12-01

    Population control of feral animals is often difficult, as it can be dangerous for the animals, labour intensive and expensive. Therefore, a useful tool for control of animal populations would be a non-surgical method to induce sterility. Our laboratories utilize methods aimed at targeting brain cells in vivo with vehicles that deliver a payload of either inhibitory RNAs or genes intended to correct cellular dysfunction. A useful framework for design of a new approach will be the combination of these methods with the intended goal to produce a technique that can be used to non-invasively sterilize cats and dogs. For this approach to succeed, it has to meet several conditions: the target gene must be essential for fertility; the method must include a mechanism to effectively and specifically silence the gene of interest; the method of delivering the silencing agent must be minimally invasive, and finally, the silencing effect must be sustained for the lifespan of the target species, so that expansion of the population can be effectively prevented. In this article, we discuss our work to develop gene silencing technology to induce sterility; we will use examples of our previous studies demonstrating that this approach is viable. These studies include (i) the use of viral vectors able to disrupt reproductive cyclicity when delivered to the regions of the brain involved in the control of reproduction and (ii) experiments with viral vectors that are able to ameliorate neuronal disease when delivered systemically using a novel approach of gene therapy. PMID:23279544

  17. Silence in the Context of "Child Voice"

    ERIC Educational Resources Information Center

    Lewis, Ann

    2010-01-01

    Recent decades have seen growing enthusiasm internationally for the concept and practice of "child voice". This was encapsulated in, and stimulated, by Article 12 of the 1989 United Nations Convention on the Rights of the Child. This article presents the case for incorporating the equally important concept of "child silence" in both research and…

  18. Parenting a Precocious Preschooler: Breaking the Silence

    ERIC Educational Resources Information Center

    Fish, Leigh Ann

    2016-01-01

    Precocity in the very young should be a valid topic of discussion in parental and educational circles, yet too frequently those conversations are slow to occur or are absent altogether. Many parents and educators remain silent about raising and nurturing precocious preschoolers, and author Leigh Ann Fish believe that the silence is due to a lack…

  19. Behold: Silence and Attention in Education

    ERIC Educational Resources Information Center

    Lewin, David

    2014-01-01

    Educators continually ask about the best means to engage students and how best to capture attention. These concerns often make the problematic assumption that students can directly govern their own attention. In order to address the role and limits of attention in education, some theorists have sought to recover the significance of silence or…

  20. Mainstream Television, Adolescent Homosexuality, and Significant Silence.

    ERIC Educational Resources Information Center

    Kielwasser, Alfred P.; Wolf, Michelle A.

    1992-01-01

    Argues that the symbolic annihilation of gay and lesbian youth exhibited by network television contributes to a dysfunctional isolation supported by the mutually reinforcing invisibility of homosexual adolescents on television and in the real world. Suggests that the spiral of silence also partially accounts for the inefficacy of oppositional…

  1. Muted Colors: Gender and Classroom Silence.

    ERIC Educational Resources Information Center

    Fredericksen, Elaine

    2000-01-01

    Highlights some causes for silence in schoolgirls and other marginalized students. Suggests ways teachers can help these students participate more fully as speakers and writers in language arts classes. Shows how language arts instruction can change students' attitudes about themselves as gendered subjects, agents, and communicators. (SR)

  2. Complete Genome Sequence of Bacillus megaterium Siphophage Silence

    PubMed Central

    Solis, Jonathan A.; Farmer, Nicholas G.; Cahill, Jesse L.; Rasche, Eric S.

    2015-01-01

    Silence is a newly isolated siphophage that infects Bacillus megaterium, a soil bacterium that is used readily in research and commercial applications. A study of B. megaterium phage Silence will enhance our knowledge of the diversity of Bacillus phages. Here, we describe the complete genome sequence and annotated features of Silence. PMID:26450722

  3. Complete Genome Sequence of Bacillus megaterium Siphophage Silence.

    PubMed

    Solis, Jonathan A; Farmer, Nicholas G; Cahill, Jesse L; Rasche, Eric S; Kuty Everett, Gabriel F

    2015-01-01

    Silence is a newly isolated siphophage that infects Bacillus megaterium, a soil bacterium that is used readily in research and commercial applications. A study of B. megaterium phage Silence will enhance our knowledge of the diversity of Bacillus phages. Here, we describe the complete genome sequence and annotated features of Silence. PMID:26450722

  4. The Sound of Silence: The Case of Virtual Team Organising

    ERIC Educational Resources Information Center

    Panteli, N.; Fineman, S.

    2005-01-01

    In this paper we discuss the role of silence within a virtual organising context. The paper raises issues related to the construction of silence in the virtual team context and the implications it has on team interactions. By drawing upon existing studies on virtual teams, we argue that members' silence may not always have negative effects on team…

  5. Mutuality, Self-Silencing, and Disordered Eating in College Women

    ERIC Educational Resources Information Center

    Wechsler, Lisa S.; Riggs, Shelley A.; Stabb, Sally D.; Marshall, David M.

    2006-01-01

    The current study examined patterns of association among mutuality, self-silencing, and disordered eating in an ethnically diverse sample of college women (N = 149). Partner mutuality and overall self-silencing were negatively correlated and together were associated with six disordered eating indices. All four self-silencing subscales were…

  6. Silenced Voices and Extraordinary Conversations... Re-Imagining Schools.

    ERIC Educational Resources Information Center

    Fine, Michelle; Weis, Lois

    This collection of papers examines the crisis in public education, focusing on poor and minority children. There are seven chapters in two parts. After "Introduction: Silenced Voices and Extraordinary Conversations" (Michelle Fine and Lois Weis), Part 1, "Scenes of Silencing," includes: (1) "Silencing and Nurturing Voice in an Improbable Context:…

  7. After the Blackbird Whistles: Listening to Silence in Classrooms

    ERIC Educational Resources Information Center

    Schultz, Katherine

    2010-01-01

    Background/Context: Students spend a large part of their time in schools in silence. However, teachers tend to spend most of their time attending to student talk. Anthropological and linguistic research has contributed to an understanding of silence in particular communities, offering explanations for students' silence in school. This research…

  8. The Coanda effect in gas-dynamic noise control. [pressure reduction by silencers

    NASA Technical Reports Server (NTRS)

    Vasilescu, G.

    1974-01-01

    The principle types of silencers are discussed for gas dynamic noise of free steam and gas expansions, as well as the results of research in gas dynamics of jets and applied acoustics. Gas dynamic noise attenuation by means of the Coanda effect is due to fluid decompression in a Coanda ejector of the external type, where a structural change takes place in the acoustic frequency spectrum and in its direction, as well as a substantial decrease in the fluid's velocity, temperature and concentration. This process is continued in the second phase with absorption of the acoustic waves by means of an active structure.

  9. Design and performance of resonant-cavity parallel baffles for duct silencing

    NASA Technical Reports Server (NTRS)

    Soderman, P. T.

    1981-01-01

    Resonant-cavity parallel baffles, either empty or with a thin absorbent lining, have been investigated as an alternative to fiberglass-filled baffles commonly used to control noise emission from large ducts. A method for predicting silencer attenuation is described, and it is shown that the new type of baffle is characterized by an acoustic performance similar to that of fibrous baffles, while being virtually immune to such problems as clogging, erosion, or settling. The emphasis of the study is on insertion loss measurements in a 7 by 10 ft wind tunnel.

  10. Vision-based estimation of vertical dynamic loading induced by jumping and bobbing crowds on civil structures

    NASA Astrophysics Data System (ADS)

    Mazzoleni, P.; Zappa, E.

    2012-11-01

    People's motion on civil structures induces dynamic loading that may lead to excessive vibrations. The complete characterization of this force distribution over a wide area due to a large number of people is still an unsolved issue. This work presents a measuring technique for the vertical load estimation in case of jumping and bobbing crowd, based on the evaluation of the vertical inertia of the human body. Laboratory experiments verify the proposed model on a single volunteer through standard inertial sensors and then extend it introducing the non-contact measuring technique. The method validation is carried out in a real environment: a stand of the G. Meazza stadium in Milan, dynamically characterized in terms of frequency response function. The load induced by groups of jumping people is estimated with the proposed method and the resulting structure accelerations are computed: the comparison between measured and estimated vibrations shows a very high correspondence in both time domain and main spectral components and, above all, the performances do not get worse as the number of volunteer increases.

  11. Tracer attenuation in groundwater

    NASA Astrophysics Data System (ADS)

    Cvetkovic, Vladimir

    2011-12-01

    The self-purifying capacity of aquifers strongly depends on the attenuation of waterborne contaminants, i.e., irreversible loss of contaminant mass on a given scale as a result of coupled transport and transformation processes. A general formulation of tracer attenuation in groundwater is presented. Basic sensitivities of attenuation to macrodispersion and retention are illustrated for a few typical retention mechanisms. Tracer recovery is suggested as an experimental proxy for attenuation. Unique experimental data of tracer recovery in crystalline rock compare favorably with the theoretical model that is based on diffusion-controlled retention. Non-Fickian hydrodynamic transport has potentially a large impact on field-scale attenuation of dissolved contaminants.

  12. Mild hypothermia combined with a scaffold of NgR-silenced neural stem cells/Schwann cells to treat spinal cord injury

    PubMed Central

    Wang, Dong; Liang, Jinhua; Zhang, Jianjun; Liu, Shuhong; Sun, Wenwen

    2014-01-01

    Because the inhibition of Nogo proteins can promote neurite growth and nerve cell differentiation, a cell-scaffold complex seeded with Nogo receptor (NgR)-silenced neural stem cells and Schwann cells may be able to improve the microenvironment for spinal cord injury repair. Previous studies have found that mild hypothermia helps to attenuate secondary damage in the spinal cord and exerts a neuroprotective effect. Here, we constructed a cell-scaffold complex consisting of a poly(D,L-lactide-co-glycolic acid) (PLGA) scaffold seeded with NgR-silenced neural stem cells and Schwann cells, and determined the effects of mild hypothermia combined with the cell-scaffold complexes on the spinal cord hemi-transection injury in the T9 segment in rats. Compared with the PLGA group and the NgR-silencing cells + PLGA group, hindlimb motor function and nerve electrophysiological function were clearly improved, pathological changes in the injured spinal cord were attenuated, and the number of surviving cells and nerve fibers were increased in the group treated with the NgR-silenced cell scaffold + mild hypothermia at 34°C for 6 hours. Furthermore, fewer pathological changes to the injured spinal cord and more surviving cells and nerve fibers were found after mild hypothermia therapy than in injuries not treated with mild hypothermia. These experimental results indicate that mild hypothermia combined with NgR gene-silenced cells in a PLGA scaffold may be an effective therapy for treating spinal cord injury. PMID:25657741

  13. Nuclear gene silencing directs reception of long-distance mRNA silencing in Arabidopsis.

    PubMed

    Brosnan, C A; Mitter, N; Christie, M; Smith, N A; Waterhouse, P M; Carroll, B J

    2007-09-11

    In plants, silencing of mRNA can be transmitted from cell to cell and also over longer distances from roots to shoots. To investigate the long-distance mechanism, WT and mutant shoots were grafted onto roots silenced for an mRNA. We show that three genes involved in a chromatin silencing pathway, NRPD1a encoding RNA polymerase IVa, RNA-dependent RNA polymerase 2 (RDR2), and DICER-like 3 (DCL3), are required for reception of long-distance mRNA silencing in the shoot. A mutant representing a fourth gene in the pathway, argonaute4 (ago4), was also partially compromised in the reception of silencing. This pathway produces 24-nt siRNAs and resulted in decapped RNA, a known substrate for amplification of dsRNA by RDR6. Activation of silencing in grafted shoots depended on RDR6, but no 24-nt siRNAs were detected in mutant rdr6 shoots, indicating that RDR6 also plays a role in initial signal perception. After amplification of decapped transcripts, DCL4 and DCL2 act hierarchically as they do in antiviral resistance to produce 21- and 22-nt siRNAs, respectively, and these guide mRNA degradation. Several dcl genotypes were also tested for their capacity to transmit the mobile silencing signal from the rootstock. dcl1-8 and a dcl2 dcl3 dcl4 triple mutant are compromised in micro-RNA and siRNA biogenesis, respectively, but were unaffected in signal transmission. PMID:17785412

  14. Systemic RNAi-mediated Gene Silencing in Nonhuman Primate and Rodent Myeloid Cells.

    PubMed

    Novobrantseva, Tatiana I; Borodovsky, Anna; Wong, Jamie; Klebanov, Boris; Zafari, Mohammad; Yucius, Kristina; Querbes, William; Ge, Pei; Ruda, Vera M; Milstein, Stuart; Speciner, Lauren; Duncan, Rick; Barros, Scott; Basha, Genc; Cullis, Pieter; Akinc, Akin; Donahoe, Jessica S; Narayanannair Jayaprakash, K; Jayaraman, Muthusamy; Bogorad, Roman L; Love, Kevin; Whitehead, Katie; Levins, Chris; Manoharan, Muthiah; Swirski, Filip K; Weissleder, Ralph; Langer, Robert; Anderson, Daniel G; de Fougerolles, Antonin; Nahrendorf, Matthias; Koteliansky, Victor

    2012-01-01

    Leukocytes are central regulators of inflammation and the target cells of therapies for key diseases, including autoimmune, cardiovascular, and malignant disorders. Efficient in vivo delivery of small interfering RNA (siRNA) to immune cells could thus enable novel treatment strategies with broad applicability. In this report, we develop systemic delivery methods of siRNA encapsulated in lipid nanoparticles (LNP) for durable and potent in vivo RNA interference (RNAi)-mediated silencing in myeloid cells. This work provides the first demonstration of siRNA-mediated silencing in myeloid cell types of nonhuman primates (NHPs) and establishes the feasibility of targeting multiple gene targets in rodent myeloid cells. The therapeutic potential of these formulations was demonstrated using siRNA targeting tumor necrosis factor-α (TNFα) which induced substantial attenuation of disease progression comparable to a potent antibody treatment in a mouse model of rheumatoid arthritis (RA). In summary, we demonstrate a broadly applicable and therapeutically relevant platform for silencing disease genes in immune cells. PMID:23344621

  15. Cutting Edge: EZH2 Promotes Osteoclastogenesis by Epigenetic Silencing of the Negative Regulator IRF8.

    PubMed

    Fang, Celestia; Qiao, Yu; Mun, Se Hwan; Lee, Min Joon; Murata, Koichi; Bae, Seyeon; Zhao, Baohong; Park-Min, Kyung-Hyun; Ivashkiv, Lionel B

    2016-06-01

    Osteoclasts are resorptive cells that are important for homeostatic bone remodeling and pathological bone resorption. Emerging evidence suggests an important role for epigenetic mechanisms in osteoclastogenesis. A recent study showed that epigenetic silencing of the negative regulator of osteoclastogenesis Irf8 by DNA methylation is required for osteoclast differentiation. In this study, we investigated the role of EZH2, which epigenetically silences gene expression by histone methylation, in osteoclastogenesis. Inhibition of EZH2 by the small molecule GSK126, or decreasing its expression using antisense oligonucleotides, impeded osteoclast differentiation. Mechanistically, EZH2 was recruited to the IRF8 promoter after RANKL stimulation to deposit the negative histone mark H3K27me3 and downregulate IRF8 expression. GSK126 attenuated bone loss in the ovariectomy mouse model of postmenopausal osteoporosis. Our findings provide evidence for an additional mechanism of epigenetic IRF8 silencing during osteoclastogenesis that likely works cooperatively with DNA methylation, further emphasizing the importance of IRF8 as a negative regulator of osteoclastogenesis. PMID:27183582

  16. An siRNA-based method for efficient silencing of gene expression in mature brown adipocytes.

    PubMed

    Isidor, Marie S; Winther, Sally; Basse, Astrid L; Petersen, M Christine H; Cannon, Barbara; Nedergaard, Jan; Hansen, Jacob B

    2016-01-01

    Brown adipose tissue is a promising therapeutic target for opposing obesity, glucose intolerance and insulin resistance. The ability to modulate gene expression in mature brown adipocytes is important to understand brown adipocyte function and delineate novel regulatory mechanisms of non-shivering thermogenesis. The aim of this study was to optimize a lipofection-based small interfering RNA (siRNA) transfection protocol for efficient silencing of gene expression in mature brown adipocytes. We determined that a critical parameter was to deliver the siRNA to mature adipocytes by reverse transfection, i.e. transfection of non-adherent cells. Using this protocol, we effectively knocked down both high- and low-abundance transcripts in a model of mature brown adipocytes (WT-1) as well as in primary mature mouse brown adipocytes. A functional consequence of the knockdown was confirmed by an attenuated increase in uncoupled respiration (thermogenesis) in response to β-adrenergic stimulation of mature WT-1 brown adipocytes transfected with uncoupling protein 1 siRNA. Efficient gene silencing was also obtained in various mouse and human white adipocyte models (3T3-L1, primary mouse white adipocytes, hMADS) with the ability to undergo "browning." In summary, we report an easy and versatile reverse siRNA transfection protocol to achieve specific silencing of gene expression in various models of mature brown and browning-competent white adipocytes, including primary cells. PMID:27386153

  17. Disruption of Rpp1-mediated soybean rust immunity by virus-induced gene silencing

    PubMed Central

    Cooper, Bret; Campbell, Kimberly B; McMahon, Michael B; Luster, Douglas G

    2013-01-01

    Phakopsora pachyrhizi, a fungus that causes rust disease on soybean, has potential to impart significant yield loss and disrupt food security and animal feed production. Rpp1 is a soybean gene that confers immunity to soybean rust, and it is important to understand how it regulates the soybean defense system and to use this knowledge to protect commercial crops. It was previously discovered that some soybean proteins resembling transcription factors accumulate in the nucleus of Rpp1 soybeans. To determine if they contribute to immunity, Bean pod mottle virus was used to attenuate or silence the expression of their genes. Rpp1 plants subjected to virus-induced gene silencing exhibited reduced amounts of RNA for 5 of the tested genes, and the plants developed rust-like symptoms after subsequent inoculation with fungal spores. Symptoms were associated with the accumulation of rust fungal RNA and protein. Silenced plants also had reduced amounts of RNA for the soybean Myb84 transcription factor and soybean isoflavone O-methyltransferase, both of which are important to phenylpropanoid biosynthesis and lignin formation, crucial components of rust resistance. These results help resolve some of the genes that contribute to Rpp1-mediated immunity and improve upon the knowledge of the soybean defense system. It is possible that these genes could be manipulated to enhance rust resistance in otherwise susceptible soybean cultivars. PMID:24401541

  18. Silencing Nociceptor Neurons Reduces Allergic Airway Inflammation.

    PubMed

    Talbot, Sébastien; Abdulnour, Raja-Elie E; Burkett, Patrick R; Lee, Seungkyu; Cronin, Shane J F; Pascal, Maud A; Laedermann, Cedric; Foster, Simmie L; Tran, Johnathan V; Lai, Nicole; Chiu, Isaac M; Ghasemlou, Nader; DiBiase, Matthew; Roberson, David; Von Hehn, Christian; Agac, Busranour; Haworth, Oliver; Seki, Hiroyuki; Penninger, Josef M; Kuchroo, Vijay K; Bean, Bruce P; Levy, Bruce D; Woolf, Clifford J

    2015-07-15

    Lung nociceptors initiate cough and bronchoconstriction. To elucidate if these fibers also contribute to allergic airway inflammation, we stimulated lung nociceptors with capsaicin and observed increased neuropeptide release and immune cell infiltration. In contrast, ablating Nav1.8(+) sensory neurons or silencing them with QX-314, a charged sodium channel inhibitor that enters via large-pore ion channels to specifically block nociceptors, substantially reduced ovalbumin- or house-dust-mite-induced airway inflammation and bronchial hyperresponsiveness. We also discovered that IL-5, a cytokine produced by activated immune cells, acts directly on nociceptors to induce the release of vasoactive intestinal peptide (VIP). VIP then stimulates CD4(+) and resident innate lymphoid type 2 cells, creating an inflammatory signaling loop that promotes allergic inflammation. Our results indicate that nociceptors amplify pathological adaptive immune responses and that silencing these neurons with QX-314 interrupts this neuro-immune interplay, revealing a potential new therapeutic strategy for asthma. PMID:26119026

  19. The Marijuana Phenomenon: Contradictions and Silence.

    PubMed

    Hall, Joanne M; Shattell, Mona M; McConnell, Elizabeth A

    2016-01-01

    The United States is trending toward more permissiveness regarding recreational and medicinal marijuana (MJ). Many conditions for which MJ is recommended, prescribed, or self-prescribed are symptoms that advanced practice nurses address daily. Yet, the silence of nursing scientists on ethics, practices, and policies regarding such clinical decisions is deafening. This is but one of many contradictions about MJ use that we discuss in this article. We do not propose to resolve these contradictions; that is left to the community of nurse scientists in interprofessional discourse. Collectively, we must explore these contradictions and, through evidence-based policy recommendations, overcome the silence about how providers view MJ, how it might be helpful, its risks, and cultural shifts that have accompanied a changed political/legal environment. Long term, we must close the gaps in the nursing knowledge base regarding MJ as it affects users and how it is used interventionally. PMID:26950836

  20. Homology-dependent Gene Silencing in Paramecium

    PubMed Central

    Ruiz, Françoise; Vayssié, Laurence; Klotz, Catherine; Sperling, Linda; Madeddu, Luisa

    1998-01-01

    Microinjection at high copy number of plasmids containing only the coding region of a gene into the Paramecium somatic macronucleus led to a marked reduction in the expression of the corresponding endogenous gene(s). The silencing effect, which is stably maintained throughout vegetative growth, has been observed for all Paramecium genes examined so far: a single-copy gene (ND7), as well as members of multigene families (centrin genes and trichocyst matrix protein genes) in which all closely related paralogous genes appeared to be affected. This phenomenon may be related to posttranscriptional gene silencing in transgenic plants and quelling in Neurospora and allows the efficient creation of specific mutant phenotypes thus providing a potentially powerful tool to study gene function in Paramecium. For the two multigene families that encode proteins that coassemble to build up complex subcellular structures the analysis presented herein provides the first experimental evidence that the members of these gene families are not functionally redundant. PMID:9529389

  1. Silencing FAT10 inhibits metastasis of osteosarcoma.

    PubMed

    Ma, Chengbin; Zhang, Zhiyu; Cui, Yan; Yuan, Hongmou; Wang, Feng

    2016-08-01

    Metastasis is the main challenge of osteosarcoma treatment. Herein, we first reveal the oncogenic role of FAT10 in metastasis of osteosarcoma. FAT10 was upregulated in osteosarcoma, especially in metastatic osteosarcoma. High level of FAT10 was associated with poorer prognosis of osteosarcoma patients. Moreover, Transwell and Matrigel assays revealed that silencing FAT10 significantly inhibited the invasive and migratory abilities of osteosarcoma cells. Metastasis assay in vivo showed that silencing FAT10 decreased the number of mice with distant metastasis. We also found that FAT10 may act its oncogenic functions through regulating HOXB9. Collectively, the results suggested that FAT10 may be a novel therapeutic target for osteosarcoma patients. PMID:27279480

  2. Silencing the Singer. Antibioethics in Germany.

    PubMed

    Schöne-Seifert, B; Rippe, K P

    1991-01-01

    "Die Gedanken sind frei," in the words of the old song. But in Germany, thoughts are no longer free. Peter Singer, the "death ethicist," has become a special target for activists attempting to silence bioethical debate in Germany. In the context of the trauma inflicted by National Socialism, a profound unease over issues at the end of life is accompanied by an insistence that these issues are not to be discussed. PMID:1837289

  3. Targeted gene silencing to induce permanent sterility.

    PubMed

    Dissen, G A; Lomniczi, A; Boudreau, R L; Chen, Y H; Davidson, B L; Ojeda, S R

    2012-08-01

    A non-surgical method to induce sterility would be a useful tool to control feral populations of animals. Our laboratories have experience with approaches aimed at targeting brain cells in vivo with vehicles that deliver a payload of either inhibitory RNAs or genes intended to correct cellular dysfunction. A combination/modification of these methods may provide a useful framework for the design of approaches that can be used to sterilize cats and dogs. For this approach to succeed, it has to meet several conditions: it needs to target a gene essential for fertility. It must involve a method that can selectively silence the gene of interest. It also needs to deliver the silencing agent via a minimally invasive method. Finally, the silencing effect needs to be sustained for many years, so that expansion of the targeted population can be effectively prevented. In this article, we discuss this subject and provide a succinct account of our previous experience with: (i) molecular reagents able to disrupt reproductive cyclicity when delivered to regions of the brain involved in the control of reproduction and (ii) molecular reagents able to ameliorate neuronal disease when delivered systemically using a novel approach of gene therapy. PMID:22827375

  4. Variable laser attenuator

    DOEpatents

    Foltyn, Stephen R.

    1988-01-01

    The disclosure relates to low loss, high power variable attenuators comprng one or more transmissive and/or reflective multilayer dielectric filters. The attenuator is particularly suitable to use with unpolarized lasers such as excimer lasers. Beam attenuation is a function of beam polarization and the angle of incidence between the beam and the filter and is controlled by adjusting the angle of incidence the beam makes to the filter or filters. Filters are selected in accordance with beam wavelength.

  5. Variable laser attenuator

    DOEpatents

    Foltyn, S.R.

    1987-05-29

    The disclosure relates to low loss, high power variable attenuators comprising one or more transmissive and/or reflective multilayer dielectric filters. The attenuator is particularly suitable to use with unpolarized lasers such as excimer lasers. Beam attenuation is a function of beam polarization and the angle of incidence between the beam and the filter and is controlled by adjusting the angle of incidence the beam makes to the filter or filters. Filters are selected in accordance with beam wavelength. 9 figs.

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

    PubMed

    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. PMID:25993336

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

    PubMed Central

    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) (109KFTMHNQ117), which is highly conserved in Asia type tospoviruses, and the benzene ring of Y398 at the C-terminal β-sheet motif (397IYFL400) 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. PMID:25993336

  8. Silencing of protein kinase D2 induces glioma cell senescence via p53-dependent and -independent pathways

    PubMed Central

    Bernhart, Eva; Damm, Sabine; Heffeter, Petra; Wintersperger, Andrea; Asslaber, Martin; Frank, Saša; Hammer, Astrid; Strohmaier, Heimo; DeVaney, Trevor; Mrfka, Manuel; Eder, Hans; Windpassinger, Christian; Ireson, Christopher R.; Mischel, Paul S.; Berger, Walter; Sattler, Wolfgang

    2014-01-01

    Background Glioblastoma multiforme (GBM) is a highly aggressive tumor of the central nervous system with a dismal prognosis for affected patients. Aberrant protein kinase C (PKC) signaling has been implicated in gliomagenesis, and a member of the PKC-activated protein kinase D (PRKD) family, PRKD2, was identified as mediator of GBM growth in vitro and in vivo. Methods The outcome of PRKD2 silencing and pharmacological inhibition on glioma cell proliferation was established with different glioma cell lines. Western blotting, senescence assays, co-immunoprecipitation, fluorescence activated cell sorting, quantitative PCR, and immunofluorescence microscopy were utilized to analyze downstream signaling. Results RNA-interference (21-mer siRNA) and pharmacological inhibition (CRT0066101) of PRKD2 profoundly inhibited proliferation of p53wt (U87MG, A172, and primary GBM2), and p53mut (GM133, T98G, U251, and primary Gli25) glioma cells. In a xenograft experiment, PRKD2 silencing significantly delayed tumor growth of U87MG cells. PRKD2 silencing in p53wt and p53mut cells was associated with typical hallmarks of senescence and cell cycle arrest in G1. Attenuated AKT/PKB phosphorylation in response to PRKD2 silencing was a common observation made in p53wt and p53mut GBM cells. PRKD2 knockdown in p53wt cells induced upregulation of p53, p21, and p27 expression, decreased phosphorylation of CDK2 and/or CDK4, hypophosphorylation of retinoblastoma protein (pRb), and reduced transcription of E2F1. In p53mut GM133 and primary Gli25 cells, PRKD2 silencing increased p27 and p15 and reduced E2F1 transcription but did not affect pRb phosphorylation. Conclusions PRKD2 silencing induces glioma cell senescence via p53-dependent and -independent pathways. PMID:24463355

  9. Mechanical regulation of transcription controls Polycomb-mediated gene silencing during lineage commitment.

    PubMed

    Le, Huy Quang; Ghatak, Sushmita; Yeung, Ching-Yan Chloé; Tellkamp, Frederik; Günschmann, Christian; Dieterich, Christoph; Yeroslaviz, Assa; Habermann, Bianca; Pombo, Ana; Niessen, Carien M; Wickström, Sara A

    2016-08-01

    Tissue mechanics drive morphogenesis, but how forces are sensed and transmitted to control stem cell fate and self-organization remains unclear. We show that a mechanosensory complex of emerin (Emd), non-muscle myosin IIA (NMIIA) and actin controls gene silencing and chromatin compaction, thereby regulating lineage commitment. Force-driven enrichment of Emd at the outer nuclear membrane of epidermal stem cells leads to defective heterochromatin anchoring to the nuclear lamina and a switch from H3K9me2,3 to H3K27me3 occupancy at constitutive heterochromatin. Emd enrichment is accompanied by the recruitment of NMIIA to promote local actin polymerization that reduces nuclear actin levels, resulting in attenuation of transcription and subsequent accumulation of H3K27me3 at facultative heterochromatin. Perturbing this mechanosensory pathway by deleting NMIIA in mouse epidermis leads to attenuated H3K27me3-mediated silencing and precocious lineage commitment, abrogating morphogenesis. Our results reveal how mechanics integrate nuclear architecture and chromatin organization to control lineage commitment and tissue morphogenesis. PMID:27398909

  10. Cohabitation of insulators and silencing elements in yeast subtelomeric regions.

    PubMed Central

    Fourel, G; Revardel, E; Koering, C E; Gilson, E

    1999-01-01

    In budding yeast, the telomeric DNA is flanked by a combination of two subtelomeric repetitive sequences, the X and Y' elements. We have investigated the influence of these sequences on telomeric silencing. The telomere-proximal portion of either X or Y' dampened silencing when located between the telomere and the reporter gene. These elements were named STARs, for subtelomeric anti-silencing regions. STARs can also counteract silencer-driven repression at the mating-type HML locus. When two STARs bracket a reporter gene, its expression is no longer influenced by surrounding silencing elements, although these are still active on a second reporter gene. In addition, an intervening STAR uncouples the silencing of neighboring genes. STARs thus display the hallmarks of insulators. Protection from silencing is recapitulated by multimerized oligonucleotides representing Tbf1p- and Reb1p-binding sites, as found in STARs. In contrast, sequences located more centromere proximal in X and Y' elements reinforce silencing. They can promote silencing downstream of an insulated expressed domain. Overall, our results suggest that the silencing emanating from telomeres can be propagated in a discontinuous manner via a series of subtelomeric relay elements. PMID:10228166

  11. RNA silencing and antiviral defense in plants.

    PubMed

    Agius, Claire; Eamens, Andrew L; Millar, Anthony A; Watson, John M; Wang, Ming-Bo

    2012-01-01

    Given the widespread impact of RNA silencing on the Arabidopsis thaliana genome, it is indeed remarkable that this means of gene regulation went undiscovered for so long. Since the publication of landmark papers in 1998 (Fire et al., Nature 391:806-811, 1998; Waterhouse et al., Proc Natl Acad Sci U S A 95:13959-13964, 1998), intense research efforts have resulted in much progress from the speculation of Mello and colleagues that "the mechanisms underlying RNA interference probably exist for a biological purpose" (Fire et al., Nature 391:806-811, 1998). Across the eukaryotic kingdom, with the notable exception of Saccharomyces cerevisiae (Moazed, Science 326:544-550, 2009), the importance of small RNA-driven gene regulation has been recognized and implicated in central developmental processes as well as in aberrant and diseased states. Plants have by far the most complex RNA-based control of gene expression (Wang et al., Floriculture, ornamental and plant biotechnology, vol. III, 2006). Four distinct RNA silencing pathways have been recognized in plants, albeit with considerable conservation of the molecular components. These pathways are directed by various small RNA species, including microRNAs (miRNAs), trans-acting small interfering RNAs (siRNA) (ta-siRNAs), repeat-associated siRNAs (ra-siRNAs), and natural antisense transcript siRNAs (nat-siRNAs). The effective functionality of each of these pathways appear to be fundamental to the integrity of A. thaliana. Furthermore, in response to viral invasion, plants synthesize viral sRNAs as a means of defense. This process may in fact reflect the ancient origins of RNA silencing: plants may have evolved RNA silencing pathways as a defense mechanism against foreign nucleic acid species in the absence of an immune system (Wang and Metzlaff, Curr Opin Plant Biol 8:216-222, 2005). The generation of viral siRNAs is a particularly interesting illustration of RNA silencing as it provides a context to explore the potential to

  12. Gender Differences in Self-Silencing and Psychological Distress in Informal Cancer Carers

    ERIC Educational Resources Information Center

    Ussher, Jane M.; Perz, Janette

    2010-01-01

    This study examined gender differences in self-silencing, the relationship between self-silencing and psychological distress, and reasons for self-silencing in informal cancer carers (329 women, 155 men), using a mixed-method design. Men reported greater self-silencing than women on the Silencing the Self Scale; however, women reported higher…

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

    PubMed

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

    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(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. PMID:22361475

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

    SciTech Connect

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

    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.

  15. Suppressing RNA silencing with small molecules and the viral suppressor of RNA silencing protein p19.

    PubMed

    Danielson, Dana C; Filip, Roxana; Powdrill, Megan H; O'Hara, Shifawn; Pezacki, John P

    2015-08-01

    RNA silencing is a gene regulatory and host defense mechanism whereby small RNA molecules are engaged by Argonaute (AGO) proteins, which facilitate gene knockdown of complementary mRNA targets. Small molecule inhibitors of AGO represent a convenient method for reversing this effect and have applications in human therapy and biotechnology. Viral suppressors of RNA silencing, such as p19, can also be used to suppress the pathway. Here we assess the compatibility of these two approaches, by examining whether synthetic inhibitors of AGO would inhibit p19-siRNA interactions. We observe that aurintricarboxylic acid (ATA) is a potent inhibitor of p19's ability to bind siRNA (IC50 = 0.43 μM), oxidopamine does not inhibit p19:siRNA interactions, and suramin is a mild inhibitor of p19:siRNA interactions (IC50 = 430 μM). We observe that p19 and suramin are compatible inhibitors of RNA silencing in human hepatoma cells. Our data suggests that at least some inhibitors of AGO may be used in combination with p19 to inhibit RNA silencing at different points in the pathway. PMID:26079891

  16. A Modular Plasmid Assembly Kit for Multigene Expression, Gene Silencing and Silencing Rescue in Plants

    PubMed Central

    Binder, Andreas; Lambert, Jayne; Morbitzer, Robert; Popp, Claudia; Ott, Thomas; Lahaye, Thomas; Parniske, Martin

    2014-01-01

    The Golden Gate (GG) modular assembly approach offers a standardized, inexpensive and reliable way to ligate multiple DNA fragments in a pre-defined order in a single-tube reaction. We developed a GG based toolkit for the flexible construction of binary plasmids for transgene expression in plants. Starting from a common set of modules, such as promoters, protein tags and transcribed regions of interest, synthetic genes are assembled, which can be further combined to multigene constructs. As an example, we created T-DNA constructs encoding multiple fluorescent proteins targeted to distinct cellular compartments (nucleus, cytosol, plastids) and demonstrated simultaneous expression of all genes in Nicotiana benthamiana, Lotus japonicus and Arabidopsis thaliana. We assembled an RNA interference (RNAi) module for the construction of intron-spliced hairpin RNA constructs and demonstrated silencing of GFP in N. benthamiana. By combination of the silencing construct together with a codon adapted rescue construct into one vector, our system facilitates genetic complementation and thus confirmation of the causative gene responsible for a given RNAi phenotype. As proof of principle, we silenced a destabilized GFP gene (dGFP) and restored GFP fluorescence by expression of a recoded version of dGFP, which was not targeted by the silencing construct. PMID:24551083

  17. Co-silencing the mirabilis antiviral protein permits virus-induced gene silencing in Mirabilis jalapa

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Virus-induced gene silencing (VIGS) is an attractive and rapid technique for loss of function assay that can reveal the phenotype of embryo-lethal sequences and avoids the need for time consuming transformation and regeneration processes. Among various VIGS vectors that have been explored, the tobac...

  18. Landing gear noise attenuation

    NASA Technical Reports Server (NTRS)

    Moe, Jeffrey W. (Inventor); Whitmire, Julia (Inventor); Kwan, Hwa-Wan (Inventor); Abeysinghe, Amal (Inventor)

    2011-01-01

    A landing gear noise attenuator mitigates noise generated by airframe deployable landing gear. The noise attenuator can have a first position when the landing gear is in its deployed or down position, and a second position when the landing gear is in its up or stowed position. The noise attenuator may be an inflatable fairing that does not compromise limited space constraints associated with landing gear retraction and stowage. A truck fairing mounted under a truck beam can have a compliant edge to allow for non-destructive impingement of a deflected fire during certain conditions.

  19. RADIO FREQUENCY ATTENUATOR

    DOEpatents

    Giordano, S.

    1963-11-12

    A high peak power level r-f attenuator that is readily and easily insertable along a coaxial cable having an inner conductor and an outer annular conductor without breaking the ends thereof is presented. Spaced first and second flares in the outer conductor face each other with a slidable cylindrical outer conductor portion therebetween. Dielectric means, such as water, contact the cable between the flares to attenuate the radio-frequency energy received thereby. The cylindrical outer conductor portion is slidable to adjust the voltage standing wave ratio to a low level, and one of the flares is slidable to adjust the attenuation level. An integral dielectric container is also provided. (AFC)

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

    PubMed Central

    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. PMID:26275304

  1. Attenuator And Conditioner

    DOEpatents

    Anderson, Gene R.; Armendariz, Marcelino G.; Carson, Richard F.; Bryan, Robert P.; Duckett, III, Edwin B.; Kemme, Shanalyn Adair; McCormick, Frederick B.; Peterson, David W.

    2006-04-04

    An apparatus and method of attenuating and/or conditioning optical energy for an optical transmitter, receiver or transceiver module is disclosed. An apparatus for attenuating the optical output of an optoelectronic connector including: a mounting surface; an array of optoelectronic devices having at least a first end; an array of optical elements having at least a first end; the first end of the array of optical elements optically aligned with the first end of the array of optoelectronic devices; an optical path extending from the first end of the array of optoelectronic devices and ending at a second end of the array of optical elements; and an attenuator in the optical path for attenuating the optical energy emitted from the array of optoelectronic devices. Alternatively, a conditioner may be adapted in the optical path for conditioning the optical energy emitted from the array of optoelectronic devices.

  2. Fiber Optic Attenuators

    NASA Technical Reports Server (NTRS)

    1995-01-01

    Mike Buzzetti designed a fiber optic attenuator while working at Jet Propulsion Laboratory, intended for use in NASA's Deep Space Network. Buzzetti subsequently patented and received an exclusive license to commercialize the device, and founded Nanometer Technologies to produce it. The attenuator functions without introducing measurable back-reflection or insertion loss, and is relatively insensitive to vibration and changes in temperature. Applications include cable television, telephone networks, other signal distribution networks, and laboratory instrumentation.

  3. Silence in the Second Language Classrooms of Japanese Universities

    ERIC Educational Resources Information Center

    King, Jim

    2013-01-01

    Japanese language learners' proclivity for silence has been alluded to by various writers (e.g. Anderson 1993; Korst 1997; Greer 2000) and is supported by plenty of anecdotal evidence, but large-scale, empirical studies aimed at measuring the extent of macro-level silence within Japanese university L2 classrooms are notably lacking. This article…

  4. Deriving Silence through Dependent Reference: Focus on Pronouns

    ERIC Educational Resources Information Center

    Livitz, Inna G.

    2014-01-01

    The starting point of this dissertation is the observation that pronouns that are obligatorily dependent on a sufficiently local antecedent are persistently silent. The classical hypothesis has been that silence is a lexical property of such elements. The central claim of this dissertation is that silence is instead a product of syntax--of the way…

  5. Self-Silencing and Rejection Sensitivity in Adolescent Romantic Relationships

    ERIC Educational Resources Information Center

    Harper, Melinda S.; Dickson, Joseph W.; Welsh, Deborah P.

    2006-01-01

    This study examined the link between rejection sensitivity, self-silencing behaviors, and depressive symptomatology among adolescent dating couples. Self-silencing was hypothesized to be the process mediating the association between rejection sensitivity and depressive symptoms. Our sample included 211 couples between 14 and 21 who were dating at…

  6. The Question of Silence: Techniques to Ensure Full Class Participation.

    ERIC Educational Resources Information Center

    Wildman, Stephanie M.

    1988-01-01

    Law school professors have a responsibility to monitor student participation and silence in the classroom, to consider why some students are silent, and to examine their role in contributing to that silence. A variety of techniques are available to encourage participation. (MSE)

  7. Silenced by Sex: Hard Truths & Taboos in Teaching Literature.

    ERIC Educational Resources Information Center

    Wolf, Shelby A.; Maniotes, Leslie K.

    2002-01-01

    Focuses on the silencing of young voices, both on and off the page, because of sexual trauma and even sexual curiosity. Considers the novel, "Speak," in which the protagonist's silence stems from the fact that she was raped. Explores two other literary texts that center on sexual issues for young teens. (SG)

  8. Virus-Induced Gene Silencing in Ornametal Plants

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Virus-Induced Gene Silencing (VIGS) provides an attractive tool for high throughput analysis of the functional effects of gene knock-down. Virus genomes are engineered to include fragments of target host genes, and the infected plant recognizes and silences the target genes as part of its viral defe...

  9. Echoes of Silence: Empathy and Making Connections through Writing Process

    ERIC Educational Resources Information Center

    Freedman, Joel M.

    2009-01-01

    On April 25, 2008, students on college and public school campuses collectively committed to a vow of silence commemorating an event known as the National Day of Silence. This student-generated, nationwide action theatrically "speaks out" in solidarity with lesbian, gay, bisexual, and transgender (LGBT) people who for one reason or another fear…

  10. A Time for Silence: Booker T. Washington in Atlanta.

    ERIC Educational Resources Information Center

    Heath, Robert L.

    1978-01-01

    Discusses the rhetoric of silence as an alternative to participation in a discussion of social issues, particularly when participation confirms the efforts of those who seek social control. Concludes that Booker T. Washington should have employed the strategy of silence when invited to address an exposition in Atlanta in 1895. (JMF)

  11. Hearing the Silence: Acknowledging the Voice of My Latina Sisters

    ERIC Educational Resources Information Center

    Martinez-Vogt, Emily

    2015-01-01

    Latina community college students experience a number of challenges during their transition to college. Findings from a larger study indicated that Latina community college students experienced racism and stereotyping on campus responding with silence. Silence occurred in two ways: (1) Latinas were forced to be silent, and/or (2) Latinas chose to…

  12. Discourses that Silence: Teachers and Anti-Lesbian Harassment

    ERIC Educational Resources Information Center

    Ferfolja, Tania

    2008-01-01

    This paper examines the way lesbian identities are silenced in schools particularly through anti-lesbian harassment. Based on research with 30 self-identified lesbian teachers working across high schools in New South Wales, Australia, the discussion illustrates how various responses to anti-lesbian harassment silence the recognition of such…

  13. The molecular topography of silenced chromatin in Saccharomyces cerevisiae

    PubMed Central

    Thurtle, Deborah M.; Rine, Jasper

    2014-01-01

    Heterochromatin imparts regional, promoter-independent repression of genes and is epigenetically heritable. Understanding how silencing achieves this regional repression is a fundamental problem in genetics and development. Current models of yeast silencing posit that Sir proteins, recruited by transcription factors bound to the silencers, spread throughout the silenced region. To test this model directly at high resolution, we probed the silenced chromatin architecture by chromatin immunoprecipitation (ChIP) followed by next-generation sequencing (ChIP-seq) of Sir proteins, histones, and a key histone modification, H4K16-acetyl. These analyses revealed that Sir proteins are strikingly concentrated at and immediately adjacent to the silencers, with lower levels of enrichment over the promoters at HML and HMR, the critical targets for transcriptional repression. The telomeres also showed discrete peaks of Sir enrichment yet a continuous domain of hypoacetylated histone H4K16. Surprisingly, ChIP-seq of cross-linked chromatin revealed a distribution of nucleosomes at silenced loci that was similar to Sir proteins, whereas native nucleosome maps showed a regular distribution throughout silenced loci, indicating that cross-linking captured a specialized chromatin organization imposed by Sir proteins. This specialized chromatin architecture observed in yeast informs the importance of a steric contribution to regional repression in other organisms. PMID:24493645

  14. Reflections on the Silencing the Self Scale and Its Origins

    ERIC Educational Resources Information Center

    Jack, Dana Crowley

    2011-01-01

    In this article, the author reflects on the Silencing the Self Scale (STSS) and blends her personal and professional thoughts about self-silencing, gender, and depression. For her, the despair of depression deeply involves questions of value and meaning, culture and freedom. The STSS grew from listening to depressed women's voices. From them, the…

  15. Virus-Induced gene silencing in ornamental plants

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Virus-Induced Gene Silencing (VIGS) provides an attractive tool for high throughput analysis of the functional effects of gene knock-down. Virus genomes are engineered to include fragments of target host genes, and the infected plant recognizes and silences the target genes as part of its viral defe...

  16. Postnatal β-catenin deletion from Dmp1-expressing osteocytes/osteoblasts reduces structural adaptation to loading, but not periosteal load-induced bone formation.

    PubMed

    Kang, Kyung Shin; Hong, Jung Min; Robling, Alexander G

    2016-07-01

    Mechanical signal transduction in bone tissue begins with load-induced activation of several cellular pathways in the osteocyte population. A key pathway that participates in mechanotransduction is Wnt/Lrp5 signaling. A putative downstream mediator of activated Lrp5 is the nucleocytoplasmic shuttling protein β-catenin (βcat), which migrates to the nucleus where it functions as a transcriptional co-activator. We investigated whether osteocytic βcat participates in Wnt/Lrp5-mediated mechanotransduction by conducting ulnar loading experiments in mice with or without chemically induced βcat deletion in osteocytes. Mice harboring βcat floxed loss-of-function alleles (βcat(f/f)) were bred to the inducible osteocyte Cre transgenic (10)(kb)Dmp1-CreERt2. Adult male mice were induced to recombine the βcat alleles using tamoxifen, and intermittent ulnar loading sessions were applied over the following week. Although adult-onset deletion of βcat from Dmp1-expressing cells reduced skeletal mass, the bone tissue was responsive to mechanical stimulation as indicated by increased relative periosteal bone formation rates in recombined mice. However, load-induced improvements in cross sectional geometric properties were compromised in recombined mice. The collective results indicate that the osteoanabolic response to loading can occur on the periosteal surface when β-cat levels are significantly reduced in Dmp1-expressing cells, suggesting that either (i) only low levels of β-cat are required for mechanically induced bone formation on the periosteal surface, or (ii) other additional downstream mediators of Lrp5 might participate in transducing load-induced Wnt signaling. PMID:27143110

  17. Transcriptional Silencing by Hairpin RNAs Complementary to a Gene Promoter

    PubMed Central

    Chu, Yongjun; Kalantari, Roya; Dodd, David W.

    2012-01-01

    Double-stranded RNAs can target gene promoters and inhibit transcription. To date, most research has focused on synthetic RNA duplexes. Transcriptional silencing by hairpin RNAs would facilitate a better understanding of endogenous RNA-mediated regulation of transcription within cells. Here we examine transcriptional silencing of progesterone receptor (PR) expression by hairpin RNAs. We identify the guide strand as the strand complementary to an antisense transcript at the PR promoter and that hairpin RNAs are active transcriptional silencing agents. The sequence of the hairpin loop affects activity, with the highest activity achieved when the loop has the potential for full complementarity to the antisense transcript target. Introduction of centrally mismatched bases relative to the target transcript does not prevent transcriptional silencing unless the mismatches are present on both the guide and passenger strands. These data demonstrate that hairpin RNAs can cause transcriptional silencing and offer insights into the mechanism of gene modulation by RNAs that target gene promoters. PMID:22703280

  18. An electronically tunable duct silencer using dielectric elastomer actuators.

    PubMed

    Lu, Zhenbo; Godaba, Hareesh; Cui, Yongdong; Foo, Choon Chiang; Debiasi, Marco; Zhu, Jian

    2015-09-01

    A duct silencer with tunable acoustic characteristics is presented in this paper. Dielectric elastomer, a smart material with lightweight, high elastic energy density and large deformation under high direct current/alternating current voltages, was used to fabricate this duct silencer. The acoustic performances and tunable mechanisms of this duct silencer were experimentally investigated. It was found that all the resonance peaks of this duct silencer could be adjusted using external control signals without any additional mechanical part. The physics of the tunable mechanism is further discussed based on the electro-mechanical interactions using finite element analysis. The present promising results also provide insight into the appropriateness of the duct silencer for possible use as next generation acoustic treatment device to replace the traditional acoustic treatment. PMID:26428819

  19. Silencing suppressors: viral weapons for countering host cell defenses.

    PubMed

    Song, Liping; Gao, Shijuan; Jiang, Wei; Chen, Shuai; Liu, Yanjun; Zhou, Ling; Huang, Wenlin

    2011-04-01

    RNA silencing is a conserved eukaryotic pathway involved in the suppression of gene expression via sequence-specific interactions that are mediated by 21-23 nt RNA molecules. During infection, RNAi can act as an innate immune system to defend against viruses. As a counter-defensive strategy, silencing suppressors are encoded by viruses to inhibit various stages of the silencing process. These suppressors are diverse in sequence and structure and act via different mechanisms. In this review, we discuss whether RNAi is a defensive strategy in mammalian host cells and whether silencing suppressors can be encoded by mammalian viruses. We also review the modes of action proposed for some silencing suppressors. PMID:21528352

  20. Evaluating the ability of the barley stripe mosaic virus-induced gene silencing system to simultaneously silence two wheat genes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Virus-induced gene silencing (VIGS) is an important tool for rapid assessment of gene function in plants. The ability of the Barley Stripe Mosaic Virus (BSMV) VIGS system to simultaneously silence two genes was assessed by comparing the extent of down-regulation of the wheat PDS and SGT1 genes afte...

  1. Evaluating the Ability of the Barley Stripe Mosaic Virus-Induced Gene Silencing System to Simultaneously Silence Two Wheat Genes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Virus-induced gene silencing (VIGS) is an important tool for rapid assessment of gene function in plants. The ability of the Barley stripe mosaic virus (BSMV) VIGS system to simultaneously silence two genes was assessed by comparing the extent of down-regulation of the wheat PDS and SGT1 genes afte...

  2. GENE SILENCING. Epigenetic silencing by the HUSH complex mediates position-effect variegation in human cells.

    PubMed

    Tchasovnikarova, Iva A; Timms, Richard T; Matheson, Nicholas J; Wals, Kim; Antrobus, Robin; Göttgens, Berthold; Dougan, Gordon; Dawson, Mark A; Lehner, Paul J

    2015-06-26

    Forward genetic screens in Drosophila melanogaster for modifiers of position-effect variegation have revealed the basis of much of our understanding of heterochromatin. We took an analogous approach to identify genes required for epigenetic repression in human cells. A nonlethal forward genetic screen in near-haploid KBM7 cells identified the HUSH (human silencing hub) complex, comprising three poorly characterized proteins, TASOR, MPP8, and periphilin; this complex is absent from Drosophila but is conserved from fish to humans. Loss of HUSH components resulted in decreased H3K9me3 both at endogenous genomic loci and at retroviruses integrated into heterochromatin. Our results suggest that the HUSH complex is recruited to genomic loci rich in H3K9me3, where subsequent recruitment of the methyltransferase SETDB1 is required for further H3K9me3 deposition to maintain transcriptional silencing. PMID:26022416

  3. Antisense Gene Silencing: Therapy for Neurodegenerative Disorders?

    PubMed Central

    Nielsen, Troels T.; Nielsen, Jørgen E.

    2013-01-01

    Since the first reports that double-stranded RNAs can efficiently silence gene expression in C. elegans, the technology of RNA interference (RNAi) has been intensively exploited as an experimental tool to study gene function. With the subsequent discovery that RNAi could also be applied to mammalian cells, the technology of RNAi expanded from being a valuable experimental tool to being an applicable method for gene-specific therapeutic regulation, and much effort has been put into further refinement of the technique. This review will focus on how RNAi has developed over the years and how the technique is exploited in a pre-clinical and clinical perspective in relation to neurodegenerative disorders. PMID:24705213

  4. Post-transcriptional gene silencing, transcriptional gene silencing and human immunodeficiency virus

    PubMed Central

    Méndez, Catalina; Ahlenstiel, Chantelle L; Kelleher, Anthony D

    2015-01-01

    While human immunodeficiency virus 1 (HIV-1) infection is controlled through continuous, life-long use of a combination of drugs targeting different steps of the virus cycle, HIV-1 is never completely eradicated from the body. Despite decades of research there is still no effective vaccine to prevent HIV-1 infection. Therefore, the possibility of an RNA interference (RNAi)-based cure has become an increasingly explored approach. Endogenous gene expression is controlled at both, transcriptional and post-transcriptional levels by non-coding RNAs, which act through diverse molecular mechanisms including RNAi. RNAi has the potential to control the turning on/off of specific genes through transcriptional gene silencing (TGS), as well as fine-tuning their expression through post-transcriptional gene silencing (PTGS). In this review we will describe in detail the canonical RNAi pathways for PTGS and TGS, the relationship of TGS with other silencing mechanisms and will discuss a variety of approaches developed to suppress HIV-1 via manipulation of RNAi. We will briefly compare RNAi strategies against other approaches developed to target the virus, highlighting their potential to overcome the major obstacle to finding a cure, which is the specific targeting of the HIV-1 reservoir within latently infected cells. PMID:26279984

  5. The effect of eccentricity and spatiotemporal energy on motion silencing.

    PubMed

    Choi, Lark Kwon; Bovik, Alan C; Cormack, Lawrence K

    2016-03-01

    The now well-known motion-silencing illusion has shown that salient changes among a group of objects' luminances, colors, shapes, or sizes may appear to cease when objects move rapidly (Suchow & Alvarez, 2011). It has been proposed that silencing derives from dot spacing that causes crowding, coherent changes in object color or size, and flicker frequencies combined with dot spacing (Choi, Bovik, & Cormack, 2014; Peirce, 2013; Turi & Burr, 2013). Motion silencing is a peripheral effect that does not occur near the point of fixation. To better understand the effect of eccentricity on motion silencing, we measured the amount of motion silencing as a function of eccentricity in human observers using traditional psychophysics. Fifteen observers reported whether dots in any of four concentric rings changed in luminance over a series of rotational velocities. The results in the human experiments showed that the threshold velocity for motion silencing almost linearly decreases as a function of log eccentricity. Further, we modeled the response of a population of simulated V1 neurons to our stimuli. We found strong matches between the threshold velocities on motion silencing observed in the human experiment and those seen in the energy model of Adelson and Bergen (1985). We suggest the plausible explanation that as eccentricity increases, the combined motion-flicker signal falls outside the narrow spatiotemporal frequency response regions of the modeled receptive fields, thereby reducing flicker visibility. PMID:27019052

  6. Bioinformatics tools for achieving better gene silencing in plants.

    PubMed

    Ahmed, Firoz; Dai, Xinbin; Zhao, Patrick Xuechun

    2015-01-01

    RNA interference (RNAi) is one of the most popular and effective molecular technologies for knocking down the expression of an individual gene of interest in living organisms. Yet the technology still faces the major issue of nonspecific gene silencing, which can compromise gene functional characterization and the interpretation of phenotypes associated with individual gene knockdown. Designing an effective and target-specific small interfering RNA (siRNA) for induction of RNAi is therefore the major challenge in RNAi-based gene silencing. A 'good' siRNA molecule must possess three key features: (a) the ability to specifically silence an individual gene of interest, (b) little or no effect on the expressions of unintended siRNA gene targets (off-target genes), and (c) no cell toxicity. Although several siRNA design and analysis algorithms have been developed, only a few of them are specifically focused on gene silencing in plants. Furthermore, current algorithms lack a comprehensive consideration of siRNA specificity, efficacy, and nontoxicity in siRNA design, mainly due to lack of integration of all known rules that govern different steps in the RNAi pathway. In this review, we first describe popular RNAi methods that have been used for gene silencing in plants and their serious limitations regarding gene-silencing potency and specificity. We then present novel, rationale-based strategies in combination with computational and experimental approaches to induce potent, specific, and nontoxic gene silencing in plants. PMID:25740355

  7. Silencing of Eag1 Gene Inhibits Osteosarcoma Proliferation and Migration by Targeting STAT3-VEGF Pathway

    PubMed Central

    Wu, Xinyu; Chen, Zhida; Zeng, Wengrong; Zhong, Yuanfu; Liu, Qingjun; Wu, Jin

    2015-01-01

    So far, the role of Ether à go-go 1 (Eag1) potassium channels in migration and invasion progression of cancers remains elusive. In the present study, the effects of Eag1 knockdown on osteosarcoma cell proliferation, growth, and apoptosis were examined. Then, we evaluated the effects of Eag1 silencing on osteosarcoma cell migration and invasion. In addition, we detected the expression of vascular endothelial growth factor (VEGF) and signal transducer and activator of transcription 3 (STAT3) in osteosarcoma cell treated with Eag1 small interfering RNAs (siRNAs). Finally, STAT3 siRNA was employed to determine the influence of downregulation of STAT3 on cell proliferation and migration. The results showed that knockdown of Eag1 significantly suppressed osteosarcoma cell proliferation and osteosarcoma xenografts growth. However, Eag1 silencing had little effect on cell apoptosis. Additionally, osteosarcoma cell adhesion, migration, and invasion were also potently attenuated. Notably, the expression levels of VEGF decreased evidently upon Eag1 siRNAs treatment, paralleled with reductions in the expression levels of STAT3. Moreover, a similar pattern was observed in osteosarcoma cell proliferation and migration suppression between STAT3 siRNA and Eag1 siRNAs groups. Our data indicated that Eag1 promotes osteosarcoma proliferation and migration, at least in part, by targeting STAT3-VEGF pathway. PMID:26783521

  8. Targeted silencing of CXCR4 inhibits epithelial-mesenchymal transition in oral squamous cell carcinoma

    PubMed Central

    Duan, Yuansheng; Zhang, Shu; Wang, Longlong; Zhou, Xuan; He, Qinghua; Liu, Su; Yue, Kai; Wang, Xudong

    2016-01-01

    Aberrant overexpression of C-X-C chemokine receptor type 4 (CXCR4) is a critical event during tumor metastasis. It has been previously reported that the expression of CXCR4 is linked with epithelial-mesenchymal transition (EMT) in oral squamous cell carcinoma (OSCC) tissues derived from patients. The present study addresses the role of CXCR4 in EMT in tongue squamous cell carcinoma (TSCCA) cells in vitro and in xenograft models. Small interfering (si) RNA sequences targeting the CXCR4 gene were transfected into TSCCA cells. Cell migration, invasion, apoptosis and EMT markers were determined in TSCCA cells using wound healing and Transwell assays, Annexin V/propdidum iodide double staining and western blot analysis, respectively. In vivo, tumor growth was assessed by subcutaneous inoculation of cells into BALB/c nude mice. Phenotypic EMT markers and regulatory factors were detected in the tumor tissues derived from the mice. In vitro, silencing of CXCR4 expression suppressed cell migration and invasion, and induced apoptosis. The protein expression of the EMT-associated markers N-cadherin and matrix metalloproteinases 2/9 were attenuated, while E-cadherin was increased. In vivo, CXCR4 siRNA inhibited tumor growth, and EMT-associated proteins had similar expression patterns to the experimental results observed in vitro. In conclusion, the present study demonstrated that CXCR4 silencing suppressed EMT in OSCC, thus affecting tumor metastasis. PMID:27602138

  9. Seismic attenuation in Florida

    SciTech Connect

    Bellini, J.J.; Bartolini, T.J.; Lord, K.M.; Smith, D.L. . Dept. of Geology)

    1993-03-01

    Seismic signals recorded by the expanded distribution of earthquake seismograph stations throughout Florida and data from a comprehensive review of record archives from stations GAI contribute to an initial seismic attenuation model for the Florida Plateau. Based on calculations of surface particle velocity, a pattern of attenuation exists that appears to deviate from that established for the remainder of the southeastern US. Most values suggest greater seismic attenuation within the Florida Plateau. However, a separate pattern may exist for those signals arising from the Gulf of Mexico. These results have important implications for seismic hazard assessments in Florida and may be indicative of the unique lithospheric identity of the Florida basement as an exotic terrane.

  10. Radiofrequency attenuator and method

    DOEpatents

    Warner, Benjamin P.; McCleskey, T. Mark; Burrell, Anthony K.; Agrawal, Anoop; Hall, Simon B.

    2009-01-20

    Radiofrequency attenuator and method. The attenuator includes a pair of transparent windows. A chamber between the windows is filled with molten salt. Preferred molten salts include quarternary ammonium cations and fluorine-containing anions such as tetrafluoroborate (BF.sub.4.sup.-), hexafluorophosphate (PF.sub.6.sup.-), hexafluoroarsenate (AsF.sub.6.sup.-), trifluoromethylsulfonate (CF.sub.3SO.sub.3.sup.-), bis(trifluoromethylsulfonyl)imide ((CF.sub.3SO.sub.2).sub.2N.sup.-), bis(perfluoroethylsulfonyl)imide ((CF.sub.3CF.sub.2SO.sub.2).sub.2N.sup.-) and tris(trifluoromethylsulfonyl)methide ((CF.sub.3SO.sub.2).sub.3C.sup.-). Radicals or radical cations may be added to or electrochemically generated in the molten salt to enhance the RF attenuation.

  11. Radiofrequency attenuator and method

    DOEpatents

    Warner, Benjamin P.; McCleskey, T. Mark; Burrell, Anthony K.; Agrawal, Anoop; Hall, Simon B.

    2009-11-10

    Radiofrequency attenuator and method. The attenuator includes a pair of transparent windows. A chamber between the windows is filled with molten salt. Preferred molten salts include quarternary ammonium cations and fluorine-containing anions such as tetrafluoroborate (BF.sub.4.sup.-), hexafluorophosphate (PF.sub.6.sup.-), hexafluoroarsenate (AsF.sub.6.sup.-), trifluoromethylsulfonate (CF.sub.3SO.sub.3.sup.-), bis(trifluoromethylsulfonyl)imide ((CF.sub.3SO.sub.2).sub.2N.sup.-), bis(perfluoroethylsulfonyl)imide ((CF.sub.3CF.sub.2SO.sub.2).sub.2N.sup.-) and tris(trifluoromethylsulfonyl)methide ((CF.sub.3SO.sub.2).sub.3 C.sup.-). Radicals or radical cations may be added to or electrochemically generated in the molten salt to enhance the RF attenuation.

  12. RNA binding proteins implicated in Xist-mediated chromosome silencing.

    PubMed

    Moindrot, Benoit; Brockdorff, Neil

    2016-08-01

    Chromosome silencing by Xist RNA occurs in two steps; localisation in cis within the nuclear matrix to form a domain that corresponds to the territory of the inactive X chromosome elect, and transduction of silencing signals from Xist RNA to the underlying chromatin. Key factors that mediate these processes have been identified in a series of recent studies that harnessed comprehensive proteomic or genetic screening strategies. In this review we discuss these findings in light of prior knowledge both of Xist-mediated silencing and known functions/properties of the novel factors. PMID:26816113

  13. Analysis of trans-silencing interactions using transcriptional silencers of varying strength and targets with and without flanking nuclear matrix attachment regions.

    PubMed

    Ascenzi, Robert; Ulker, Bekir; Todd, Joselyn J; Sowinski, Dolores A; Schimeneck, Carolyn R; Allen, George C; Weissinger, Arthur K; Thompson, William F

    2003-06-01

    We investigated the effect of the Rb7 matrix attachment region (MAR) on trans-silencing in tobacco plants, comparing the effects of three transgene silencer loci on ten target loci. Two of the silencer loci, C40 and C190, contain complex and rearranged transgene arrays consisting of 35S:GUS or NOS:NPTII containing plasmids. The third silencer locus, V271, was previously characterized as a complex locus containing rearranged 35S:RiN sequences. Each of these silencers can reduce 35S promoter-driven expression at other loci, albeit with varying efficiencies. The presence of MARs at a target locus does not prevent trans-silencing by the V271 silencer. However, four of seven MAR-containing loci were at least partially resistant to silencing by the C40 and C190 loci. One MAR locus was unaffected by C40, our weakest silencer, and three were silenced only when the silencer locus was maternally inherited. Silencing is progressive in the F1 and F2 generations; two days after germination there is little or no difference between seedlings derived from crosses to silencing or control lines, but seedlings containing silencer loci slowly lose expression during subsequent development. These observations are compatible with the hypothesis that a product of the silencer locus must accumulate before unlinked loci can be affected. However, our silencer loci are themselves silenced for GUS transcription, and coding region homology is not required for their effects on target loci. Our results are consistent with a model in which transcriptional silencing is triggered by transcription of sequences during the early stages of embryo or seedling development. PMID:12779119

  14. In Vivo Silencing of A20 via TLR9-Mediated Targeted SiRNA Delivery Potentiates Antitumor Immune Response.

    PubMed

    Braun, Floriane C M; van den Brandt, Jens; Thomas, Sören; Lange, Sandra; Schrank, Juliane; Gand, Claudia; Przybylski, Grzegorz K; Schmoeckel, Katrin; Bröker, Barbara M; Schmidt, Christian A; Grabarczyk, Piotr

    2015-01-01

    A20 is an ubiquitin-editing enzyme that ensures the transient nature of inflammatory signaling pathways induced by cytokines like TNF-α and IL-1 or pathogens via Toll-like receptor (TLR) pathways. It has been identified as a negative regulator of dendritic cell (DC) maturation and attenuator of their immunostimulatory properties. Ex vivo A20-depleted dendritic cells showed enhanced expression of pro-inflammatory cytokines and costimulatory molecules, which resulted in hyperactivation of tumor-infiltrating T lymphocytes and inhibition of regulatory T cells. In the present study, we demonstrate that a synthetic molecule consisting of a CpG oligonucleotide TLR9 agonist linked to A20-specific siRNAs silences its expression in TLR9+ mouse dendritic cells in vitro and in vivo. In the B16 mouse melanoma tumor model, silencing of A20 enhances the CpG-triggered induction of NFκB activity followed by elevated expression of IL-6, TNF-α and IL-12. This leads to potentiated antitumor immune responses manifested by increased numbers of tumor-specific cytotoxic T cells, high levels of tumor cell apoptosis and delayed tumor growth. Our findings confirm the central role of A20 in controlling the immunostimulatory potency of DCs and provide a strategy for simultaneous A20 silencing and TLR activation in vivo. PMID:26327508

  15. Obesity-Induced Colorectal Cancer Is Driven by Caloric Silencing of the Guanylin-GUCY2C Paracrine Signaling Axis.

    PubMed

    Lin, Jieru E; Colon-Gonzalez, Francheska; Blomain, Erik; Kim, Gilbert W; Aing, Amanda; Stoecker, Brian; Rock, Justin; Snook, Adam E; Zhan, Tingting; Hyslop, Terry M; Tomczak, Michal; Blumberg, Richard S; Waldman, Scott A

    2016-01-15

    Obesity is a well-known risk factor for colorectal cancer but precisely how it influences risks of malignancy remains unclear. During colon cancer development in humans or animals, attenuation of the colonic cell surface receptor guanylyl cyclase C (GUCY2C) that occurs due to loss of its paracrine hormone ligand guanylin contributes universally to malignant progression. In this study, we explored a link between obesity and GUCY2C silencing in colorectal cancer. Using genetically engineered mice on different diets, we found that diet-induced obesity caused a loss of guanylin expression in the colon with subsequent GUCY2C silencing, epithelial dysfunction, and tumorigenesis. Mechanistic investigations revealed that obesity reversibly silenced guanylin expression through calorie-dependent induction of endoplasmic reticulum stress and the unfolded protein response in intestinal epithelial cells. In transgenic mice, enforcing specific expression of guanylin in intestinal epithelial cells restored GUCY2C signaling, eliminating intestinal tumors associated with a high calorie diet. Our findings show how caloric suppression of the guanylin-GUCY2C signaling axis links obesity to negation of a universal tumor suppressor pathway in colorectal cancer, suggesting an opportunity to prevent colorectal cancer in obese patients through hormone replacement with the FDA-approved oral GUCY2C ligand linaclotide. PMID:26773096

  16. In Vivo Silencing of A20 via TLR9-Mediated Targeted SiRNA Delivery Potentiates Antitumor Immune Response

    PubMed Central

    Braun, Floriane C. M.; van den Brandt, Jens; Thomas, Sören; Lange, Sandra; Schrank, Juliane; Gand, Claudia; Przybylski, Grzegorz K.; Schmoeckel, Katrin; Bröker, Barbara M.; Schmidt, Christian A.; Grabarczyk, Piotr

    2015-01-01

    A20 is an ubiquitin-editing enzyme that ensures the transient nature of inflammatory signaling pathways induced by cytokines like TNF-α and IL-1 or pathogens via Toll-like receptor (TLR) pathways. It has been identified as a negative regulator of dendritic cell (DC) maturation and attenuator of their immunostimulatory properties. Ex vivo A20-depleted dendritic cells showed enhanced expression of pro-inflammatory cytokines and costimulatory molecules, which resulted in hyperactivation of tumor-infiltrating T lymphocytes and inhibition of regulatory T cells. In the present study, we demonstrate that a synthetic molecule consisting of a CpG oligonucleotide TLR9 agonist linked to A20-specific siRNAs silences its expression in TLR9+ mouse dendritic cells in vitro and in vivo. In the B16 mouse melanoma tumor model, silencing of A20 enhances the CpG-triggered induction of NFκB activity followed by elevated expression of IL-6, TNF-α and IL-12. This leads to potentiated antitumor immune responses manifested by increased numbers of tumor-specific cytotoxic T cells, high levels of tumor cell apoptosis and delayed tumor growth. Our findings confirm the central role of A20 in controlling the immunostimulatory potency of DCs and provide a strategy for simultaneous A20 silencing and TLR activation in vivo. PMID:26327508

  17. Tritium Attenuation by Distillation

    SciTech Connect

    Wittman, N.E.

    2001-07-31

    The objective of this study was to determine how a 100 Area distillation system could be used to reduce to a satisfactory low value the tritium content of the dilute moderator produced in the 100 Area stills, and whether such a tritium attenuator would have sufficient capacity to process all this material before it is sent to the 400 Area for reprocessing.

  18. The Effects of Vocational High School Teachers' Perceived Trust on Organizational Silence

    ERIC Educational Resources Information Center

    Saglam, Aycan Çiçek

    2016-01-01

    The objective of this research is to reveal the effects of vocational school teachers' perceived organizational trust on organizational silence. For this purpose, at first teachers' perception on sub-dimensions of organizational silence and organizational trust, which are respectively "acquiescent silence," "defensive silence,"…

  19. Post-transcriptional gene silencing activity of human GIGYF2.

    PubMed

    Kryszke, Marie-Hélène; Adjeriou, Badia; Liang, Feifei; Chen, Hong; Dautry, François

    2016-07-01

    In mammalian post-transcriptional gene silencing, the Argonaute protein AGO2 indirectly recruits translation inhibitors, deadenylase complexes, and decapping factors to microRNA-targeted mRNAs, thereby repressing mRNA translation and accelerating mRNA decay. However, the exact composition and assembly pathway of the microRNA-induced silencing complex are not completely elucidated. As the GYF domain of human GIGYF2 was shown to bind AGO2 in pulldown experiments, we wondered whether GIGYF2 could be a novel protein component of the microRNA-induced silencing complex. Here we show that full-length GIGYF2 coimmunoprecipitates with AGO2 in human cells, and demonstrate that, upon tethering to a reporter mRNA, GIGYF2 exhibits strong, dose-dependent silencing activity, involving both mRNA destabilization and translational repression. PMID:27157137

  20. Mutations in Ran system affected telomere silencing in Saccharomyces cerevisiae

    SciTech Connect

    Hayashi, Naoyuki Kobayashi, Masahiko; Shimizu, Hiroko; Yamamoto, Ken-ichi; Murakami, Seishi; Nishimoto, Takeharu

    2007-11-23

    The Ran GTPase system regulates the direction and timing of several cellular events, such as nuclear-cytosolic transport, centrosome formation, and nuclear envelope assembly in telophase. To gain insight into the Ran system's involvement in chromatin formation, we investigated gene silencing at the telomere in several mutants of the budding yeast Saccharomyces cerevisiae, which had defects in genes involved in the Ran system. A mutation of the RanGAP gene, rna1-1, caused reduced silencing at the telomere, and partial disruption of the nuclear Ran binding factor, yrb2-{delta}2, increased this silencing. The reduced telomere silencing in rna1-1 cells was suppressed by a high dosage of the SIR3 gene or the SIT4 gene. Furthermore, hyperphosphorylated Sir3 protein accumulated in the rna1-1 mutant. These results suggest that RanGAP is required for the heterochromatin structure at the telomere in budding yeast.

  1. Temperature-Responsive Gene Silencing by a Smart Polymer.

    PubMed

    Wang, Mingming; Cheng, Yiyun

    2016-03-16

    Intracellular siRNA release is a crucial step in efficient gene silencing mediated by cationic polymers. Here, we show an example of temperature change-induced intracellular siRNA release and silencing using a temperature-responsive polymer consisting of dendrimer, poly(N-isopropylacrylamide) and phenylboronic acid. The smart polymer can trigger the release of loaded siRNA in a controlled manner upon cooling the surrounding solution below its lower critical solution temperature. Gene silencing efficacy of the polymer was significantly increased by cool treatment after its cellular uptake. The polymer and the cool treatment cause minimal toxicity to the transfected cells. The results provide a facile and promising strategy to design stimuli-responsive polymers for efficient gene silencing. PMID:26783999

  2. New Construct Approaches for Efficient Gene Silencing in Plants

    PubMed Central

    Yan, Hua; Chretien, Robert; Ye, Jingsong; Rommens, Caius M.

    2006-01-01

    An important component of conventional sense, antisense, and double-strand RNA-based gene silencing constructs is the transcriptional terminator. Here, we show that this regulatory element becomes obsolete when gene fragments are positioned between two oppositely oriented and functionally active promoters. The resulting convergent transcription triggers gene silencing that is at least as effective as unidirectional promoter-to-terminator transcription. In addition to short, variably sized, and nonpolyadenylated RNAs, terminator-free cassette produced rare, longer transcripts that reach into the flanking promoter. These read-through products did not influence the efficacy and expression levels of the neighboring hygromycin phosphotransferase gene. Replacement of gene fragments by promoter-derived sequences further increased the extent of gene silencing. This finding indicates that genomic DNA may be a more efficient target for gene silencing than gene transcripts. PMID:16766670

  3. Variations in hypovirus interactions with the fungal-host RNA-silencing antiviral-defense response.

    PubMed

    Zhang, Xuemin; Shi, Diane; Nuss, Donald L

    2012-12-01

    Hypoviruses Cryphonectria hypovirus 1 (CHV-1)/EP713, CHV-1/Euro7, and CHV-1/EP721, which infect the chestnut blight fungus Cryphonectria parasitica, differ in their degrees of virulence attenuation (hypovirulence), symptom expression, and viral RNA accumulation, even though they share between 90% and 99% amino acid sequence identity. In this report we examine whether this variability is influenced by interactions with the C. parasitica Dicer gene dcl2-dependent RNA-silencing antiviral defense response. The mild symptoms exhibited by strains infected with CHV-1/Euro7 and CHV-1/EP721 relative to those with severe hypovirus CHV-1/EP713 did not correlate with a higher induction of the RNA-silencing pathway. Rather, dcl2 transcripts accumulated to a higher level (∼8-fold) following infection by CHV-1/EP713 than following infection by CHV-1/Euro7 (1.2-fold) or CHV-1/EP721 (1.4-fold). The differences in dcl2 transcript accumulation in response to CHV-1/EP713 and CHV-1/EP721 were unrelated to the suppressor of RNA silencing, p29, encoded by the two viruses. Moreover, the coding strand viral RNA levels increased by 33-, 32-, and 16-fold for CHV-1/EP713, CHV-1/Euro7, and CHV-1/EP721, respectively, in Δdcl2 mutant strains. This indicates that a very robust antiviral RNA-silencing response was induced against all three viruses, even though significant differences in the levels of dcl2 transcript accumulation were observed. Unexpectedly, the severe debilitation previously reported for CHV-1/EP713-infected Δdcl2 mutant strains, and observed here for the CHV-1/Euro7-infected Δdcl2 mutant strains, was not observed with infection by CHV-1/EP721. By constructing chimeric viruses containing portions of CHV-1/EP713 and CHV-1/EP721, it was possible to map the region that is associated with the severe debilitation of the Δdcl2 mutant hosts to a 4.1-kb coding domain located in the central part of the CHV-1/EP713 genome. PMID:22993160

  4. Improved design of PPRHs for gene silencing.

    PubMed

    Rodríguez, Laura; Villalobos, Xenia; Solé, Anna; Lliberós, Carolina; Ciudad, Carlos J; Noé, Véronique

    2015-03-01

    Nowadays, the modulation of gene expression by nucleic acids has become a routine tool in biomedical research for target validation and it is also used to develop new therapeutic approaches. Recently, we developed the so-called polypurine reverse Hoogsteen hairpins (PPRHs) that show high stability and a low immunogenic profile and we demonstrated their efficacy both in vitro and in vivo. In this work, we explored different characteristics of PPRHs to improve their usage as a tool for gene silencing. We studied the role of PPRH length in the range from 20 to 30 nucleotides. We also proved their higher affinity of binding and efficacy on cell viability compared to nonmodified TFOs. To overcome possible off-target effects, we tested wild-type PPRHs, which proved to be capable of binding to their target sequence with more affinity, displaying a higher stability of binding and a higher effect in terms of cell viability. Moreover, we developed a brand new molecule called Wedge-PPRH with the ability to lock the ds-DNA into the displaced structure and proved its efficacy in prostate and breast cancer cell lines. PMID:25615267

  5. A is for aphorism - the power of silence.

    PubMed

    Kreijkamp-Kaspers, Sanne; Glasziou, Paul

    2012-11-01

    'All you have to do is listen' is the title of Rob Kapilow's delightful book on classical music; but he could equally have been talking about general practice consultations. Listening requires several skills including attention, echoing and body language, but begins with silence. Well timed silences, used judiciously, can allow the patient adequate space to express symptoms and concerns, while allowing the general practitioner more time for attention, comprehension and synthesis. PMID:23145428

  6. RNA Silencing Is Resistant to Low-Temperature in Grapevine

    PubMed Central

    Romon, Marjorie; Soustre-Gacougnolle, Isabelle; Schmitt, Carine; Perrin, Mireille; Burdloff, Yannick; Chevalier, Elodie; Mutterer, Jérome; Himber, Christophe; Zervudacki, Jérôme; Montavon, Thomas; Zimmermann, Aude; Elmayan, Taline; Vaucheret, Hervé; Dunoyer, Patrice; Masson, Jean E.

    2013-01-01

    RNA silencing is a natural defence mechanism against viruses in plants, and transgenes expressing viral RNA-derived sequences were previously shown to confer silencing-based enhanced resistance against the cognate virus in several species. However, RNA silencing was shown to dysfunction at low temperatures in several species, questioning the relevance of this strategy in perennial plants such as grapevines, which are often exposed to low temperatures during the winter season. Here, we show that inverted-repeat (IR) constructs trigger a highly efficient silencing reaction in all somatic tissues in grapevines. Similarly to other plant species, IR-derived siRNAs trigger production of secondary transitive siRNAs. However, and in sharp contrast to other species tested to date where RNA silencing is hindered at low temperature, this process remained active in grapevine cultivated at 4°C. Consistently, siRNA levels remained steady in grapevines cultivated between 26°C and 4°C, whereas they are severely decreased in Arabidopsis grown at 15°C and almost undetectable at 4°C. Altogether, these results demonstrate that RNA silencing operates in grapevine in a conserved manner but is resistant to far lower temperatures than ever described in other species. PMID:24376561

  7. A compact rotary vane attenuator

    NASA Technical Reports Server (NTRS)

    Nixon, D. L.; Otosh, T. Y.; Stelzried, C. T.

    1969-01-01

    Rotary vane attenuator, when used as a front end attenuator, introduces an insertion loss that is proportional to the angle of rotation. New technique allows the construction of a shortened compact unit suitable for most installations.

  8. Scion on a Stock Producing siRNAs of Potato Spindle Tuber Viroid (PSTVd) Attenuates Accumulation of the Viroid

    PubMed Central

    Kasai, Atsushi; Sano, Teruo; Harada, Takeo

    2013-01-01

    Plants can attenuate the replication of plant viruses and viroids by RNA silencing induced by virus and viroid infection. In higher plants, silencing signals such as small interfering RNAs (siRNAs) produced by RNA silencing can be transported systemically through phloem, so it is anticipated that antiviral siRNA signals produced in a stock would have the potential to attenuate propagation of viruses or viroids in the scion. To test whether this is indeed the case, we prepared transgenic tobacco (Nicotiana benthamiana) expressing a hairpin RNA (hpRNA) of Potato spindle tuber viroid (PSTVd) in companion cells by using a strong companion cell-specific promoter. A grafting experiment of the wild type tobacco scion on the top of the transgenic tobacco stock revealed that accumulation of PSTVd challenge-inoculated into the scion was apparently attenuated compared to the control grafted plants. These results indicate that genetically modified rootstock expressing viroid-specific siRNAs can attenuate viroid accumulation in a non-genetically modified scion grafted on the stock. PMID:23469061

  9. The Treatment of Fibrosis of Joint Synovium and Frozen Shoulder by Smad4 Gene Silencing in Rats

    PubMed Central

    Xue, MingFeng; Gong, SuiLiang; Dai, JiaPing; Chen, Gang; Hu, JunYu

    2016-01-01

    Soft tissue fibrosis at the joint induced by inflammation is the pathological basis of frozen shoulder. In the present study, we utilized a lentiviral approach to silence the Smad4 gene in an in vitro fibrosis model of fibroblasts and an in vivo frozen shoulder model. We observed the change in the fibrosis process and the biological indicators of frozen shoulder. The in vitro fibrosis models (Rat myoblasts L6, Rat synovial cell RSC-364 and Rat chondrocytes RCs) were established using TGF-β1 induction, and the effect of Smad4 gene silencing on fibrosis was analyzed. The method of Kanno A was employed to establish a rat model of frozen shoulder, and Smad4 in the relevant part was knocked down with the lentiviral approach. We then examined the abduction and rotation angles and the length of synovial intima and measured the inflammatory factors in effusion and the fibrotic markers of tissues. We found that Smad4 knockdown suppressed the proliferation and expression of fibrotic markers in L6, RSC-364 and RCs cells induced by TGF-β1. MMP activity measurements showed that Smad4 knockdown significantly reversed the decrease in MMP activity in these three cell lines that were induced by TGF-β1. Furthermore, using lentivirus in the rat frozen shoulder model, we found that Smad4 silencing attenuated the inflammatory response and fibrosis. It significantly inhibited the increase of the Vimentin, α-SMA, collagen I and III, Lama1 and Timp1 proteins in synovial tissue as well as the inflammatory factors of TNF-a, IL-1α/β, IL-6 and IL-10 in effusion. MMP acidity assays revealed that Smad4 silencing inhibited MMP activity in the synovial, cartilage and ligament tissues in the model animals. The assessment of the phosphorylated Smad2/3 in the nuclei isolated from the synovial tissues showed that Smad4 silencing significantly inhibited the phosphorylation and subsequent nuclear translocation of Smad2/3 proteins. Moreover, Smad4-shRNA lentivirus inhibited the decrease in both

  10. Inhibition of T-Type Voltage Sensitive Calcium Channel Reduces Load-Induced OA in Mice and Suppresses the Catabolic Effect of Bone Mechanical Stress on Chondrocytes

    PubMed Central

    Srinivasan, Padma P.; Parajuli, Ashutosh; Price, Christopher; Wang, Liyun; Duncan, Randall L.; Kirn-Safran, Catherine B.

    2015-01-01

    Voltage-sensitive calcium channels (VSCC) regulate cellular calcium influx, one of the earliest responses to mechanical stimulation in osteoblasts. Here, we postulate that T-type VSCCs play an essential role in bone mechanical response to load and participate in events leading to the pathology of load-induced OA. Repetitive mechanical insult was used to induce OA in Cav3.2 T-VSCC null and wild-type control mouse knees. Osteoblasts (MC3T3-E1) and chondrocytes were treated with a selective T-VSCC inhibitor and subjected to fluid shear stress to determine how blocking of T-VSCCs alters the expression profile of each cell type upon mechanical stimulation. Conditioned-media (CM) obtained from static and sheared MC3T3-E1 was used to assess the effect of osteoblast-derived factors on the chondrocyte phenotype. T-VSCC null knees exhibited significantly lower focal articular cartilage damage than age-matched controls. In vitro inhibition of T-VSCC significantly reduced the expression of both early and late mechanoresponsive genes in osteoblasts but had no effect on gene expression in chondrocytes. Furthermore, treatment of chondrocytes with CM obtained from sheared osteoblasts induced expression of markers of hypertrophy in chondrocytes and this was nearly abolished when osteoblasts were pre-treated with the T-VSCC-specific inhibitor. These results indicate that T-VSCC plays a role in signaling events associated with induction of OA and is essential to the release of osteoblast-derived factors that promote an early OA phenotype in chondrocytes. Further, these findings suggest that local inhibition of T-VSCC may serve as a therapy for blocking load-induced bone formation that results in cartilage degeneration. PMID:26011709

  11. Silencing PP2A inhibitor by lenti-shRNA interference ameliorates neuropathologies and memory deficits in tg2576 mice.

    PubMed

    Liu, Gong-Ping; Wei, Wei; Zhou, Xin; Shi, Hai-Rong; Liu, Xing-Hua; Chai, Gao-Shang; Yao, Xiu-Qing; Zhang, Jia-Yu; Peng, Cai-Xia; Hu, Juan; Li, Xia-Chun; Wang, Qun; Wang, Jian-Zhi

    2013-12-01

    Deficits of protein phosphatase-2A (PP2A) play a crucial role in tau hyperphosphorylation, amyloid overproduction, and synaptic suppression of Alzheimer's disease (AD), in which PP2A is inactivated by the endogenously increased inhibitory protein, namely inhibitor-2 of PP2A (I2(PP2A)). Therefore, in vivo silencing I2(PP2A) may rescue PP2A and mitigate AD neurodegeneration. By infusion of lentivirus-shRNA targeting I2(PP2A) (LV-siI2(PP2A)) into hippocampus and frontal cortex of 11-month-old tg2576 mice, we demonstrated that expression of LV-siI2(PP2A) decreased remarkably the elevated I2(PP2A) in both mRNA and protein levels. Simultaneously, the PP2A activity was restored with the mechanisms involving reduction of the inhibitory binding of I2(PP2A) to PP2A catalytic subunit (PP2AC), repression of the inhibitory Leu309-demethylation and elevation of PP2AC. Silencing I2(PP2A) induced a long-lasting attenuation of amyloidogenesis in tg2576 mice with inhibition of amyloid precursor protein hyperphosphorylation and β-secretase activity, whereas simultaneous inhibition of PP2A abolished the antiamyloidogenic effects of I2(PP2A) silencing. Finally, silencing I2(PP2A) could improve learning and memory of tg2576 mice with preservation of several memory-associated components. Our data reveal that targeting I2(PP2A) can efficiently rescue Aβ toxicities and improve the memory deficits in tg2576 mice, suggesting that I2(PP2A) could be a promising target for potential AD therapies. PMID:23922015

  12. Silencing of a Germin-Like Gene in Nicotiana attenuata Improves Performance of Native Herbivores1[W

    PubMed Central

    Lou, Yonggen; Baldwin, Ian T.

    2006-01-01

    Germins and germin-like proteins (GLPs) are known to function in pathogen resistance, but their involvement in defense against insect herbivores is poorly understood. In the native tobacco Nicotiana attenuata, attack from the specialist herbivore Manduca sexta or elicitation by adding larval oral secretions (OS) to wounds up-regulates transcripts of a GLP. To understand the function of this gene, which occurs as a single copy, we cloned the full-length NaGLP and silenced its expression in N. attenuata by expressing a 250-bp fragment in an antisense orientation with an Agrobacterium-based transformation system and by virus-induced gene silencing (VIGS). Homozygous lines harboring a single insert and VIGS plants had significantly reduced constitutive (measured in roots) and elicited NaGLP transcript levels (in leaves). Silencing NaGLP improved M. sexta larval performance and Tupiocoris notatus preference, two native herbivores of N. attenuata. Silencing NaGLP also attenuated the OS-induced hydrogen peroxide (H2O2), diterpene glycosides, and trypsin proteinase inhibitor responses, which may explain the observed susceptibility of antisense or VIGS plants to herbivore attack and increased nicotine contents, but did not influence the OS-elicited jasmonate and salicylate bursts, or the release of the volatile organic compounds (limonene, cis-α-bergamotene, and germacrene-A) that function as an indirect defense. This suggests that NaGLP is involved in H2O2 production and might also be related to ethylene production and/or perception, which in turn influences the defense responses of N. attenuata via H2O2 and ethylene-signaling pathways. PMID:16461381

  13. Functional characterization of neural-restrictive silencer element in mouse pituitary adenylate cyclase-activating polypeptide (PACAP) gene expression.

    PubMed

    Sugawara, Hideki; Tominaga, Aiko; Inoue, Kazuhiko; Takeda, Yasuo; Yamada, Katsushi; Miyata, Atsuro

    2014-11-01

    Pituitary adenylate cyclase-activating polypeptide (PACAP) is predominantly localized in the nervous system, but the underlying mechanism in its neuron-specific expression remains unclear. In addition to two neural-restrictive silencer-like element (NRSLE1 and 2), as reported previously, we have identified the third element in -1,601 to -1,581 bp from the translational initiation site of mouse PACAP gene and termed it as NRSLE3, of which, the sequence and location were highly conserved among mouse, rat, and human PACAP genes. In luciferase reporter assay, the deletion or site-directed mutagenesis of NRSLE3 in the reporter gene construct, driven by heterologous SV40 promoter, cancelled the repression of luciferase activity in non-neuronal Swiss-3T3 cells. Furthermore, its promoter activity was significantly repressed in Swiss-3T3 cells, but not in neuronal-differentiated PC12 cells. The electrophoretic mobility shift assay (EMSA) with nuclear extracts of Swiss-3T3 cells demonstrated a specific complex with NRSLE3 probe that exhibited the same migration with the neural-restrictive silencer element (NRSE) probe of rat type II sodium channel gene. During neuronal differentiation of PC12 cells, the increment of PACAP mRNA exhibited the correlation with that of REST4 mRNA, which is a neuron-specific variant form of neural-restrictive silencer factor (NRSF). In undifferentiated PC12 cells, trichostatin A (TSA), a histone deacetylase (HDAC) inhibitor, which indirectly inhibits NRSF-mediated gene silencing, increased PACAP mRNA level and attenuated the repression of promoter activity of 5' flanking region of mouse PACAP gene containing NRSLEs. These suggest that the NRSE-NRSF system implicates in the regulatory mechanism of neuron-specific expression of PACAP gene. PMID:24939248

  14. A role for the Saccharomyces cerevisiae RENT complex protein Net1 in HMR silencing.

    PubMed Central

    Kasulke, Daniela; Seitz, Stefanie; Ehrenhofer-Murray, Ann E

    2002-01-01

    Silencing in the yeast Saccharomyces cerevisiae is known in three classes of loci: in the silent mating-type loci HML and HMR, in subtelomeric regions, and in the highly repetitive rDNA locus, which resides in the nucleolus. rDNA silencing differs markedly from the other two classes of silencing in that it requires a DNA-associated protein complex termed RENT. The Net1 protein, a central component of RENT, is required for nucleolar integrity and the control of exit from mitosis. Another RENT component is the NAD(+)-dependent histone deacetylase Sir2, which is the only silencing factor known to be shared among the three classes of silencing. Here, we investigated the role of Net1 in HMR silencing. The mutation net1-1, as well as NET1 expression from a 2micro-plasmid, restored repression at silencing-defective HMR loci. Both effects were strictly dependent on the Sir proteins. We found overexpressed Net1 protein to be directly associated with the HMR-E silencer, suggesting that Net1 could interact with silencer binding proteins and recruit other silencing factors to the silencer. In agreement with this, Net1 provided ORC-dependent, Sir1-independent silencing when artificially tethered to the silencer. In contrast, our data suggested that net1-1 acted indirectly in HMR silencing by releasing Sir2 from the nucleolus, thus shifting the internal competition for Sir2 from the silenced loci toward HMR. PMID:12196389

  15. Fluid dynamic bowtie attenuators

    NASA Astrophysics Data System (ADS)

    Szczykutowicz, Timothy P.; Hermus, James

    2015-03-01

    Fluence field modulated CT allows for improvements in image quality and dose reduction. To date, only 1-D modulators have been proposed, the extension to 2-D modulation is difficult with solid-metal attenuation-based modulators. This work proposes to use liquids and gas to attenuate the x-ray beam which can be arrayed allowing for 2-D fluence modulation. The thickness of liquid and the pressure for a given path length of gas were determined that provided the same attenuation as 30 cm of soft tissue at 80, 100, 120, and 140 kV. Gaseous Xenon and liquid Iodine, Zinc Chloride, and Cerium Chloride were studied. Additionally, we performed some proof-of-concept experiments in which (1) a single cell of liquid was connected to a reservoir which allowed the liquid thickness to be modulated and (2) a 96 cell array was constructed in which the liquid thickness in each cell was adjusted manually. Liquid thickness varied as a function of kV and chemical composition, with Zinc Chloride allowing for the smallest thickness; 1.8, 2.25, 3, and 3.6 cm compensated for 30 cm of soft tissue at 80, 100, 120, and 140 kV respectively. The 96 cell Iodine attenuator allowed for a reduction in both dynamic range to the detector and scatter to primary ratio. Successful modulation of a single cell was performed at 0, 90, and 130 degrees using a simple piston/actuator. The thickness of liquids and the Xenon gas pressure seem logistically implementable within the constraints of CBCT and diagnostic CT systems.

  16. Downhole pressure attenuation apparatus

    SciTech Connect

    Ricles, T.D.; Barton, J.A.

    1992-02-18

    This patent describes a process for preventing damage to tool strings and other downhole equipment in a well caused by pressures produced during detonation of one or more downhole explosive devices. It comprises adding to a tool string at least one pressure attenuating apparatus for attenuating the peak pressure wave and quasi-static pressure pulse produced by the explosive devices, the pressure attenuating apparatus including an initially closed relief vent including tubing means supporting a plurality of charge port assemblies each including an explosive filled shaped charge and a prestressed disc, the shaped charges interconnected by a detonating cord, the amount of explosive in each shaped charge being sufficient to rupture its associated disc without damaging surrounding tubular bodies in the well, and a vent chamber defined by the tubing means and providing a liquid free volume, and opening the relief vent substantially contemporaneously with downhole explosive device detonation by detonating the shaped charges to rupture the discs of the charge port assemblies.

  17. Flexible graphene based microwave attenuators.

    PubMed

    Byun, Kisik; Ju Park, Yong; Ahn, Jong-Hyun; Min, Byung-Wook

    2015-02-01

    We demonstrate flexible 3 dB and 6 dB microwave attenuators using multilayer graphene grown by the chemical vapor deposition method. On the basis of the characterized results of multilayer graphene and graphene-Au ohmic contacts, the graphene attenuators are designed and measured. The flexible graphene-based attenuators have 3 dB and 6 dB attenuation with a return loss of less than -15 dB at higher than 5 GHz. The devices have shown durability in a bending cycling test of 100 times. The circuit model of the attenuator based on the characterized results matches the experimental results well. PMID:25590144

  18. Control algorithms for dynamic attenuators

    SciTech Connect

    Hsieh, Scott S.; Pelc, Norbert J.

    2014-06-15

    Purpose: The authors describe algorithms to control dynamic attenuators in CT and compare their performance using simulated scans. Dynamic attenuators are prepatient beam shaping filters that modulate the distribution of x-ray fluence incident on the patient on a view-by-view basis. These attenuators can reduce dose while improving key image quality metrics such as peak or mean variance. In each view, the attenuator presents several degrees of freedom which may be individually adjusted. The total number of degrees of freedom across all views is very large, making many optimization techniques impractical. The authors develop a theory for optimally controlling these attenuators. Special attention is paid to a theoretically perfect attenuator which controls the fluence for each ray individually, but the authors also investigate and compare three other, practical attenuator designs which have been previously proposed: the piecewise-linear attenuator, the translating attenuator, and the double wedge attenuator. Methods: The authors pose and solve the optimization problems of minimizing the mean and peak variance subject to a fixed dose limit. For a perfect attenuator and mean variance minimization, this problem can be solved in simple, closed form. For other attenuator designs, the problem can be decomposed into separate problems for each view to greatly reduce the computational complexity. Peak variance minimization can be approximately solved using iterated, weighted mean variance (WMV) minimization. Also, the authors develop heuristics for the perfect and piecewise-linear attenuators which do not requirea priori knowledge of the patient anatomy. The authors compare these control algorithms on different types of dynamic attenuators using simulated raw data from forward projected DICOM files of a thorax and an abdomen. Results: The translating and double wedge attenuators reduce dose by an average of 30% relative to current techniques (bowtie filter with tube current

  19. Ultrasonic attenuation in pearlitic steel.

    PubMed

    Du, Hualong; Turner, Joseph A

    2014-03-01

    Expressions for the attenuation coefficients of longitudinal and transverse ultrasonic waves are developed for steel with pearlitic microstructure. This type of lamellar duplex microstructure influences attenuation because of the lamellar spacing. In addition, longitudinal attenuation measurements were conducted using an unfocused transducer with 10 MHz central frequency on the cross section of a quenched railroad wheel sample. The dependence of longitudinal attenuation on the pearlite microstructure is observed from the changes of longitudinal attenuation from the quenched tread surface to deeper locations. The results show that the attenuation value is lowest and relatively constant within the quench depth, then increases linearly. The experimental results demonstrate a reasonable agreement with results from the theoretical model. Ultrasonic attenuation provides an important non-destructive method to evaluate duplex microstructure within grains which can be implemented for quality control in conjunction with other manufacturing processes. PMID:24268679

  20. Study on plate silencer with general boundary conditions

    NASA Astrophysics Data System (ADS)

    Liu, Gongmin; Zhao, Xiaochen; Zhang, Wenping; Li, Shuaijun

    2014-09-01

    A plate silencer consists of an expansion chamber with two side-branch rigid cavities covered by plates. Previous studies showed that, in a duct, the introduction of simply supported or clamped plates into an air conveying system could achieve broadband quieting from low to medium frequencies. In this study, analytical formulation is extended to the plate silencer with general boundary conditions. A set of static beam functions, which are a combination of sine series and third-order polynomial, is employed as the trial functions of the plate vibration velocity. Greens function and Kirchhoff-Helmholtz integral are used to solve the sound radiation in the duct and the cavity, and then the vibration velocity of the plate is obtained. Having obtained the vibration velocity, the pressure perturbations induced by the plate oscillation and the transmission loss are found. Optimization is carried out in order to obtain the widest stopband. The transmission loss calculated by the analytical method agrees closely with the result of the finite element method simulation. Further studies with regard to the plate under several different classical boundary conditions based on the validated model show that a clamped-free plate silencer has the worst stopband. Attempts to release the boundary restriction of the plate are also made to study its effect on sound reflection. Results show that a softer end for a clamped-clamped plate silencer helps increase the optimal bandwidth, while the same treatment for simply supported plate silencer will result in performance degradation.

  1. Foxtail Mosaic Virus-Induced Gene Silencing in Monocot Plants.

    PubMed

    Liu, Na; Xie, Ke; Jia, Qi; Zhao, Jinping; Chen, Tianyuan; Li, Huangai; Wei, Xiang; Diao, Xianmin; Hong, Yiguo; Liu, Yule

    2016-07-01

    Virus-induced gene silencing (VIGS) is a powerful technique to study gene function in plants. However, very few VIGS vectors are available for monocot plants. Here we report that Foxtail mosaic virus (FoMV) can be engineered as an effective VIGS system to induce efficient silencing of endogenous genes in monocot plants including barley (Hordeum vulgare L.), wheat (Triticum aestivum) and foxtail millet (Setaria italica). This is evidenced by FoMV-based silencing of phytoene desaturase (PDS) and magnesium chelatase in barley, of PDS and Cloroplastos alterados1 in foxtail millet and wheat, and of an additional gene IspH in foxtail millet. Silencing of these genes resulted in photobleached or chlorosis phenotypes in barley, wheat, and foxtail millet. Furthermore, our FoMV-based gene silencing is the first VIGS system reported for foxtail millet, an important C4 model plant. It may provide an efficient toolbox for high-throughput functional genomics in economically important monocot crops. PMID:27225900

  2. Reconstitution of Heterochromatin-Dependent Transcriptional Gene Silencing

    PubMed Central

    Johnson, Aaron; Li, Geng; Sikorski, Timothy W.; Buratowski, Stephen; Woodcock, Christopher L.; Moazed, Danesh

    2009-01-01

    Summary Heterochromatin assembly in budding yeast requires the SIR complex, which contains the NAD-dependent deacetylase Sir2 and the Sir3 and Sir4 proteins. Sir3 binds to nucleosomes containing deacetylated histone H4 lysine 16 (H4K16) and, with Sir4, promotes spreading of Sir2 and deacetylation along the chromatin fiber. Combined action of histone modifying and binding activities is a conserved hallmark of heterochromatin, but the relative contribution of each activity to silencing has remained unclear. Here we reconstitute SIR-chromatin complexes using purified components and show that the SIR complex efficiently deacetylates chromatin templates and promotes the assembly of altered structures that silence Gal4-VP16-activated transcription. Silencing requires all three Sir proteins, even with fully deacetylated chromatin, and involves the specific association of Sir3 with deacetylated H4K16. These results define a minimal set of components that mediate heterochromatic gene silencing and demonstrate distinct contributions for histone deacetylation and nucleosome binding in the silencing mechanism. PMID:19782027

  3. Is the Efficiency of RNA Silencing Evolutionarily Regulated?

    PubMed Central

    Ui-Tei, Kumiko

    2016-01-01

    Small interfering RNAs (siRNAs) and microRNAs (miRNAs) regulate gene expression in a sequence-specific manner. Genes with partial complementarity to siRNA/miRNA sequences in their 3′-untranslated regions (UTRs) are suppressed by a mechanism referred to as the siRNA off-target effect or miRNA-mediated RNA silencing. However, the determinants of such RNA silencing efficiency are poorly understood. Previously, I and co-workers reported that the efficiency of RNA silencing is strongly correlated with the thermodynamic stability of base pairing in the duplex formed within an siRNA/miRNA and between the seed region and its target mRNA. In this review, I first summarize our previous studies that identified the thermodynamic parameter to estimate the silencing efficiency using the calculated base pairing stability: siRNAs downregulate the expression of off-target genes depending on the stability of binding between the siRNA seed region (nucleotides 2–8) and off-target mRNAs, and miRNAs downregulate target mRNA expression depending on the stability of the duplex formed between the 5′ terminus of the miRNA and its target mRNA. I further discuss the possibility that such thermodynamic features of silencing efficiency may have arisen during evolution with increasing body temperature in various organisms. PMID:27187367

  4. Is the Efficiency of RNA Silencing Evolutionarily Regulated?

    PubMed

    Ui-Tei, Kumiko

    2016-01-01

    Small interfering RNAs (siRNAs) and microRNAs (miRNAs) regulate gene expression in a sequence-specific manner. Genes with partial complementarity to siRNA/miRNA sequences in their 3'-untranslated regions (UTRs) are suppressed by a mechanism referred to as the siRNA off-target effect or miRNA-mediated RNA silencing. However, the determinants of such RNA silencing efficiency are poorly understood. Previously, I and co-workers reported that the efficiency of RNA silencing is strongly correlated with the thermodynamic stability of base pairing in the duplex formed within an siRNA/miRNA and between the seed region and its target mRNA. In this review, I first summarize our previous studies that identified the thermodynamic parameter to estimate the silencing efficiency using the calculated base pairing stability: siRNAs downregulate the expression of off-target genes depending on the stability of binding between the siRNA seed region (nucleotides 2-8) and off-target mRNAs, and miRNAs downregulate target mRNA expression depending on the stability of the duplex formed between the 5' terminus of the miRNA and its target mRNA. I further discuss the possibility that such thermodynamic features of silencing efficiency may have arisen during evolution with increasing body temperature in various organisms. PMID:27187367

  5. On the robustness of SAC silencing in closed mitosis

    NASA Astrophysics Data System (ADS)

    Ruth, Donovan; Liu, Jian

    Mitosis equally partitions sister chromatids to two daughter cells. This is achieved by properly attaching these chromatids via their kinetochores to microtubules that emanate from the spindle poles. Once the last kinetochore is properly attached, the spindle microtubules pull the sister chromatids apart. Due to the dynamic nature of microtubules, however, kinetochore-microtubule attachment often goes wrong. When this erroneous attachment occurs, it locally activates an ensemble of proteins, called the spindle assembly checkpoint proteins (SAC), which halts the mitotic progression until all the kinetochores are properly attached by spindle microtubules. The timing of SAC silencing thus determines the fidelity of chromosome segregation. We previously established a spatiotemporal model that addresses the robustness of SAC silencing in open mitosis for the first time. Here, we focus on closed mitosis by examining yeast mitosis as a model system. Though much experimental work has been done to study the SAC in cells undergoing closed mitosis, the processes responsible are not well understood. We leverage and extend our previous model to study SAC silencing mechanism in closed mitosis. We show that a robust signal of the SAC protein accumulation at the spindle pole body can be achieved. This signal is a nonlinear increasing function of number of kinetochore-microtubule attachments, and can thus serve as a robust trigger to time the SAC silencing. Together, our mechanism provides a unified framework across species that ensures robust SAC silencing and fidelity of chromosome segregation in mitosis. Intramural research program in NHLBI at NIH.

  6. Breaking the Silence: Protein Stabilization Uncovers Silenced Biosynthetic Gene Clusters in the Fungus Aspergillus nidulans

    PubMed Central

    Gerke, Jennifer; Bayram, Özgür; Feussner, Kirstin; Landesfeind, Manuel; Shelest, Ekaterina; Feussner, Ivo

    2012-01-01

    The genomes of filamentous fungi comprise numerous putative gene clusters coding for the biosynthesis of chemically and structurally diverse secondary metabolites (SMs), which are rarely expressed under laboratory conditions. Previous approaches to activate these genes were based primarily on artificially targeting the cellular protein synthesis apparatus. Here, we applied an alternative approach of genetically impairing the protein degradation apparatus of the model fungus Aspergillus nidulans by deleting the conserved eukaryotic csnE/CSN5 deneddylase subunit of the COP9 signalosome. This defect in protein degradation results in the activation of a previously silenced gene cluster comprising a polyketide synthase gene producing the antibiotic 2,4-dihydroxy-3-methyl-6-(2-oxopropyl)benzaldehyde (DHMBA). The csnE/CSN5 gene is highly conserved in fungi, and therefore, the deletion is a feasible approach for the identification of new SMs. PMID:23001671

  7. Digitally Controlled Beam Attenuator

    NASA Astrophysics Data System (ADS)

    Peppler, W. W.; Kudva, B.; Dobbins, J. T.; Lee, C. S.; Van Lysel, M. S.; Hasegawa, B. H.; Mistretta, C. A.

    1982-12-01

    In digital fluorographic techniques the video camera must accommodate a wide dynamic range due to the large variation in the subject thickness within the field of view. Typically exposure factors and the optical aperture are selected such that the maximum video signal is obtained in the most transmissive region of the subject. Consequently, it has been shown that the signal-to-noise ratio is severely reduced in the dark regions. We have developed a prototype digital beam attenuator (DBA) which will alleviate this and some related problems in digital fluorography. The prototype DBA consists of a 6x6 array of pistons which are individually controlled. A membrane containing an attenuating solu-tion of (CeC13) in water and the piston matrix are placed between the x-ray tube and the subject. Under digital control the pistons are moved into the attenuating material in order to adjust the beam intensity over each of the 36 cells. The DBA control unit which digitizes the image during patient positioning will direct the pistons under hydraulic control to produce a uniform x-ray field exiting the subject. The pistons were designed to produce very little structural background in the image. In subtraction studies any structure would be cancelled. For non-subtraction studies such as cine-cardiology we are considering higher cell densities (eg. 64x64). Due to the narrow range of transmission provided by the DBA, in such studies ultra-high contrast films could be used to produce a high resolution quasi-subtraction display. Additional benefits of the DBA are: 1) reduced dose to the bright image areas when the dark areas are properly exposed. 2) improved scatter and glare to primary ratios, leading to improved contrast in the dark areas.

  8. RNA interference for CFTR attenuates lung fluid absorption at birth in rats

    PubMed Central

    Li, Tianbo; Koshy, Shyny; Folkesson, Hans G

    2008-01-01

    Background Small interfering RNA (siRNA) against αENaC (α-subunit of the epithelial Na channel) and CFTR (cystic fibrosis transmembrane conductance regulator) was used to explore ENaC and CTFR function in newborn rat lungs. Methods Twenty-four hours after trans-thoracic intrapulmonary (ttip) injection of siRNA-generating plasmid DNA (pSi-0, pSi-4, or pSi-C2), we measured CFTR and ENaC expression, extravascular lung water, and mortality. Results αENaC and CFTR mRNA and protein decreased by ~80% and ~85%, respectively, following αENaC and CFTR silencing. Extravascular lung water and mortality increased after αENaC and CFTR-silencing. In pSi-C2-transfected isolated DLE cells there were attenuated CFTR mRNA and protein. In pSi-4-transfected DLE cells αENaC mRNA and protein were both reduced. Interestingly, CFTR-silencing also reduced αENaC mRNA and protein. αENaC silencing, on the other hand, only slightly reduced CFTR mRNA and protein. Conclusion Thus, ENaC and CFTR are both involved in the fluid secretion to absorption conversion around at birth. PMID:18652671

  9. Radiation Imaging and Attenuation

    NASA Astrophysics Data System (ADS)

    Davison, Candace; Yocum, Douglas

    2008-03-01

    X-ray and neutron images are used to demonstrate materials' different radiation attenuation properties. This leads to discussion of applications in medicine, industry and research. The Penn State Radiation Science and Engineering Center (RSEC) uses neutron radioscopy to image the inside of a working hydrogen fuel cell. This is one of the many educational activities that are conducted when students visit the RSEC. To encourage pre-college students to apply these principles and learn more about nuclear technology, we are sponsoring a design competition. For more information visit www.rsec.psu.edu

  10. Structure and Gene-Silencing Mechanisms of Small Noncoding RNAs

    NASA Astrophysics Data System (ADS)

    Chu, Chia-Ying; Rana, Tariq M.

    Small (19-31-nucleotides) noncoding RNAs were identified in the past 10 years for their distinct function in gene silencing. The best known gene-silencing phenomenon, RNA interference (RNAi), is triggered in a sequence-specific manner by endogenously produced or exogenously introduced small doubled-stranded RNAs. As knowledge of the structure and function of the RNAi machinery has expanded, this phenomenon has become a powerful tool for biochemical research; it has enormous potential for therapeutics. This chapter summarizes significant aspects of three major classes of small noncoding, regulatory RNAs: small interfering RNAs (siRNAs), microRNAs (miRNAs), and Piwi-interacting RNAs (piRNAs). Here, we focus on the biogenesis of these small RNAs, their structural features and coupled effectors as well as the mechanisms of each small regulatory RNA pathway which reveal fascinating ways by which gene silencing is controlled and fine-tuned at an epigenetic level.

  11. Delivery of gene silencing agents for breast cancer therapy

    PubMed Central

    2013-01-01

    The discovery of RNA interference has opened the door for the development of a new class of cancer therapeutics. Small inhibitory RNA oligos are being designed to specifically suppress expression of proteins that are traditionally considered nondruggable, and microRNAs are being evaluated to exert broad control of gene expression for inhibition of tumor growth. Since most naked molecules are not optimized for in vivo applications, the gene silencing agents need to be packaged into delivery vehicles in order to reach the target tissues as their destinations. Thus, the selection of the right delivery vehicles serves as a crucial step in the development of cancer therapeutics. The current review summarizes the status of gene silencing agents in breast cancer and recent development of candidate cancer drugs in clinical trials. Nanotechnology-based delivery vectors for the formulation and packaging of gene silencing agents are also described. PMID:23659575

  12. Gene Silencing in Crustaceans: From Basic Research to Biotechnologies

    PubMed Central

    Sagi, Amir; Manor, Rivka; Ventura, Tomer

    2013-01-01

    Gene silencing through RNA interference (RNAi) is gaining momentum for crustaceans, both in basic research and for commercial development. RNAi has proven instrumental in a growing number of crustacean species, revealing the functionality of novel crustacean genes essential among others to development, growth, metabolism and reproduction. Extensive studies have also been done on silencing of viral transcripts in crustaceans, contributing to the understanding of the defense mechanisms of crustaceans and strategies employed by viruses to overcome these. The first practical use of gene silencing in aquaculture industry has been recently achieved, through manipulation of a crustacean insulin-like androgenic gland hormone. This review summarizes the advancements in the use of RNAi in crustaceans, and assesses the advantages of this method, as well as the current hurdles that hinder its large-scale practice. PMID:24705266

  13. Temporal control of gene silencing by in ovo electroporation.

    PubMed

    Baeriswyl, Thomas; Mauti, Olivier; Stoeckli, Esther T

    2008-01-01

    The analysis of gene function during embryonic development asks for tight temporal control of gene expression. Classic genetic tools do not allow for this, as the absence of a gene during the early stages of development will preclude its functional analysis during later stages. In contrast, RNAi technology allows one to achieve temporal control of gene silencing especially when used with oviparous animal models. In contrast to mammals, reptiles and birds are easily accessible during embryonic development. We have developed approaches to use RNAi for the analysis of gene function during nervous system development in the chicken embryo. Although the protocol given here describes a method for gene silencing in the developing spinal cord, it can easily be adapted to other parts of the developing nervous system. The combination of the easy accessibility of the chicken embryo and RNAi provides a unique opportunity for temporal and spatial control of gene silencing during development. PMID:18369789

  14. Biomimetic RNA Silencing Nanocomplexes Overcome Multidrug Resistance in Cancer Cells**

    PubMed Central

    Wang, Zhongliang; Wang, Zhe; Liu, Dingbin; Yan, Xuefeng; Wang, Fu; Niu, Gang

    2015-01-01

    RNA interference (RNAi) is an RNA-dependent gene silencing approach controlled by RNA-induced silencing complex (RISC). Here we represent a synthetic RISC-mimic nanocomplex, which can actively cleave its target RNA in a sequence-specific manner. With high enzymatic stability and efficient self-delivery to target cells, the designed nanocomplex can selectively and potently induce gene silencing without cytokine activation. The nanocomplexes targeting to multidrug resistance are able to not only bypass P-glycoprotein (Pgp) transporter due to their nano-size effect, but also effectively suppress the Pgp expression, thus resulting in successful restoration of drug sensitivity of OVCAR8/ADR cells to Pgp-transportable cytotoxic agents. This nanocomplex approach has the potential for both functional genomics and cancer therapy. PMID:24446433

  15. Ribosomal Stalk Protein Silencing Partially Corrects the ΔF508-CFTR Functional Expression Defect.

    PubMed

    Veit, Guido; Oliver, Kathryn; Apaja, Pirjo M; Perdomo, Doranda; Bidaud-Meynard, Aurélien; Lin, Sheng-Ting; Guo, Jingyu; Icyuz, Mert; Sorscher, Eric J; Hartman Iv, John L; Lukacs, Gergely L

    2016-05-01

    The most common cystic fibrosis (CF) causing mutation, deletion of phenylalanine 508 (ΔF508 or Phe508del), results in functional expression defect of the CF transmembrane conductance regulator (CFTR) at the apical plasma membrane (PM) of secretory epithelia, which is attributed to the degradation of the misfolded channel at the endoplasmic reticulum (ER). Deletion of phenylalanine 670 (ΔF670) in the yeast oligomycin resistance 1 gene (YOR1, an ABC transporter) of Saccharomyces cerevisiae phenocopies the ΔF508-CFTR folding and trafficking defects. Genome-wide phenotypic (phenomic) analysis of the Yor1-ΔF670 biogenesis identified several modifier genes of mRNA processing and translation, which conferred oligomycin resistance to yeast. Silencing of orthologues of these candidate genes enhanced the ΔF508-CFTR functional expression at the apical PM in human CF bronchial epithelia. Although knockdown of RPL12, a component of the ribosomal stalk, attenuated the translational elongation rate, it increased the folding efficiency as well as the conformational stability of the ΔF508-CFTR, manifesting in 3-fold augmented PM density and function of the mutant. Combination of RPL12 knockdown with the corrector drug, VX-809 (lumacaftor) restored the mutant function to ~50% of the wild-type channel in primary CFTRΔF508/ΔF508 human bronchial epithelia. These results and the observation that silencing of other ribosomal stalk proteins partially rescue the loss-of-function phenotype of ΔF508-CFTR suggest that the ribosomal stalk modulates the folding efficiency of the mutant and is a potential therapeutic target for correction of the ΔF508-CFTR folding defect. PMID:27168400

  16. Silencing miR-146a influences B cells and ameliorates experimental autoimmune myasthenia gravis

    PubMed Central

    Zhang, JunMei; Jia, Ge; Liu, Qun; Hu, Jue; Yan, Mei; Yang, BaiFeng; Yang, Huan; Zhou, WenBin; Li, Jing

    2015-01-01

    MicroRNAs have been shown to be important regulators of immune homeostasis as patients with aberrant microRNA expression appeared to be more susceptible to autoimmune diseases. We recently found that miR-146a was up-regulated in activated B cells in response to rat acetylcholine receptor (AChR) α-subunit 97-116 peptide, and this up-regulation was significantly attenuated by AntagomiR-146a. Our data also demonstrated that silencing miR-146a with its inhibitor AntagomiR-146a effectively ameliorated clinical myasthenic symptoms in mice with ongoing experimental autoimmune myasthenia gravis. Furthermore, multiple defects were observed after miR-146a was knocked down in B cells, including decreased anti-R97-116 antibody production and class switching, reduced numbers of plasma cells, memory B cells and B-1 cells, and weakened activation of B cells. Previously, miR-146a has been identified as a nuclear factor-κB-dependent gene and predicted to base pair with the tumour necrosis factor receptor-associated factor 6 (TRAF6) and interleukin-1 receptor-associated kinase 1 (IRAK1) genes to regulate the immune response. However, our study proved that miR-146a inhibition had no effect on the expression of TRAF6 and IRAK1 in B cells. This result suggests that the function of miR-146a in B cells does not involve these two target molecules. We conclude that silencing miR-146a exerts its therapeutic effects by influencing the B-cell functions that contribute to the autoimmune pathogenesis of myasthenia gravis. PMID:24962817

  17. Ribosomal Stalk Protein Silencing Partially Corrects the ΔF508-CFTR Functional Expression Defect

    PubMed Central

    Veit, Guido; Oliver, Kathryn; Apaja, Pirjo M.; Perdomo, Doranda; Bidaud-Meynard, Aurélien; Guo, Jingyu; Icyuz, Mert; Sorscher, Eric J.; Hartman IV, John L.; Lukacs, Gergely L.

    2016-01-01

    The most common cystic fibrosis (CF) causing mutation, deletion of phenylalanine 508 (ΔF508 or Phe508del), results in functional expression defect of the CF transmembrane conductance regulator (CFTR) at the apical plasma membrane (PM) of secretory epithelia, which is attributed to the degradation of the misfolded channel at the endoplasmic reticulum (ER). Deletion of phenylalanine 670 (ΔF670) in the yeast oligomycin resistance 1 gene (YOR1, an ABC transporter) of Saccharomyces cerevisiae phenocopies the ΔF508-CFTR folding and trafficking defects. Genome-wide phenotypic (phenomic) analysis of the Yor1-ΔF670 biogenesis identified several modifier genes of mRNA processing and translation, which conferred oligomycin resistance to yeast. Silencing of orthologues of these candidate genes enhanced the ΔF508-CFTR functional expression at the apical PM in human CF bronchial epithelia. Although knockdown of RPL12, a component of the ribosomal stalk, attenuated the translational elongation rate, it increased the folding efficiency as well as the conformational stability of the ΔF508-CFTR, manifesting in 3-fold augmented PM density and function of the mutant. Combination of RPL12 knockdown with the corrector drug, VX-809 (lumacaftor) restored the mutant function to ~50% of the wild-type channel in primary CFTRΔF508/ΔF508 human bronchial epithelia. These results and the observation that silencing of other ribosomal stalk proteins partially rescue the loss-of-function phenotype of ΔF508-CFTR suggest that the ribosomal stalk modulates the folding efficiency of the mutant and is a potential therapeutic target for correction of the ΔF508-CFTR folding defect. PMID:27168400

  18. Chopping-Wheel Optical Attenuator

    NASA Technical Reports Server (NTRS)

    Leviton, Douglas B.

    1988-01-01

    Star-shaped rotating chopping wheel provides adjustable time-averaged attenuation of narrow beam of light without changing length of optical path or spectral distribution of light. Duty cycle or attenuation factor of chopped beam controlled by adjusting radius at which beam intersects wheel. Attenuation factor independent of wavelength. Useful in systems in which chopping frequency above frequency-response limits of photodetectors receiving chopped light. Used in systems using synchronous detection with lock-in amplifiers.

  19. Ultrasonic Attenuation in Zircaloy-4

    SciTech Connect

    Gomez, M.P.; Banchik, A.D.; Lopez Pumarega, M.I.; Ruzzante, J.E.

    2005-04-09

    In this work the relationship between Zircaloy-4 grain size and ultrasonic attenuation behavior was studied for longitudinal waves in the frequency range of 10-90 MHz. The attenuation was analyzed as a function of frequency for samples with different mechanical and heat treatments having recrystallized and Widmanstatten structures with different grain size. The attenuation behavior was analyzed by different scattering models, depending on grain size, wavelength and frequency.

  20. LINE-ABOVE-GROUND ATTENUATOR

    DOEpatents

    Wilds, R.B.; Ames, J.R.

    1957-09-24

    The line-above-ground attenuator provides a continuously variable microwave attenuator for a coaxial line that is capable of high attenuation and low insertion loss. The device consists of a short section of the line-above- ground plane type transmission lime, a pair of identical rectangular slabs of lossy material like polytron, whose longitudinal axes are parallel to and indentically spaced away from either side of the line, and a geared mechanism to adjust amd maintain this spaced relationship. This device permits optimum fineness and accuracy of attenuator control which heretofore has been difficult to achieve.

  1. Small RNA binding is a common strategy to suppress RNA silencing by several viral suppressors

    PubMed Central

    Lakatos, Lóránt; Csorba, Tibor; Pantaleo, Vitantonio; Chapman, Elisabeth J; Carrington, James C; Liu, Yu-Ping; Dolja, Valerian V; Calvino, Lourdes Fernández; López-Moya, Juan José; Burgyán, József

    2006-01-01

    RNA silencing is an evolutionarily conserved system that functions as an antiviral mechanism in higher plants and insects. To counteract RNA silencing, viruses express silencing suppressors that interfere with both siRNA- and microRNA-guided silencing pathways. We used comparative in vitro and in vivo approaches to analyse the molecular mechanism of suppression by three well-studied silencing suppressors. We found that silencing suppressors p19, p21 and HC-Pro each inhibit the intermediate step of RNA silencing via binding to siRNAs, although the molecular features required for duplex siRNA binding differ among the three proteins. None of the suppressors affected the activity of preassembled RISC complexes. In contrast, each suppressor uniformly inhibited the siRNA-initiated RISC assembly pathway by preventing RNA silencing initiator complex formation. PMID:16724105

  2. Small RNA binding is a common strategy to suppress RNA silencing by several viral suppressors.

    PubMed

    Lakatos, Lóránt; Csorba, Tibor; Pantaleo, Vitantonio; Chapman, Elisabeth J; Carrington, James C; Liu, Yu-Ping; Dolja, Valerian V; Calvino, Lourdes Fernández; López-Moya, Juan José; Burgyán, József

    2006-06-21

    RNA silencing is an evolutionarily conserved system that functions as an antiviral mechanism in higher plants and insects. To counteract RNA silencing, viruses express silencing suppressors that interfere with both siRNA- and microRNA-guided silencing pathways. We used comparative in vitro and in vivo approaches to analyse the molecular mechanism of suppression by three well-studied silencing suppressors. We found that silencing suppressors p19, p21 and HC-Pro each inhibit the intermediate step of RNA silencing via binding to siRNAs, although the molecular features required for duplex siRNA binding differ among the three proteins. None of the suppressors affected the activity of preassembled RISC complexes. In contrast, each suppressor uniformly inhibited the siRNA-initiated RISC assembly pathway by preventing RNA silencing initiator complex formation. PMID:16724105

  3. Fiber optic attenuator

    NASA Technical Reports Server (NTRS)

    Buzzetti, Mike F. (Inventor)

    1994-01-01

    A fiber optic attenuator of the invention is a mandrel structure through which a bundle of optical fibers is wrapped around in a complete circle. The mandrel structure includes a flexible cylindrical sheath through which the bundle passes. A set screw on the mandrel structure impacts one side of the sheath against two posts on the opposite side of the sheath. By rotating the screw, the sheath is deformed to extend partially between the two posts, bending the fiber optic bundle to a small radius controlled by rotating the set screw. Bending the fiber optic bundle to a small radius causes light in each optical fiber to be lost in the cladding, the amount depending upon the radius about which the bundle is bent.

  4. An attenuated philosophical gentleman.

    PubMed

    Christie, John R R

    2014-06-20

    Dr. Joseph Black had at one time, a house near us to the west. He was a striking and beautiful person; tall, very thin, and cadaverously pale; his hair carefully powdered, though there was little of it except what was collected in a long thin queue; his eyes dark, clear and large, like deep pools of pure water. He wore black speckless clothes, silk stockings, silver buckles, and either a slim green umbrella, or a genteel brown cane. The general frame and air were feeble and slender. The wildest boy respected Black. No lad could be irreverent toward a man so pale, so gentle, so elegant and so illustrious. So he glided, like a spirit, through our rather mischievous sportiveness, unharmed. He died seated, with a bowl of milk upon his knee, of which his ceasing to be did not spill a drop; a departure which it seemed, after the event, might have been foretold of this attenuated philosophical gentleman. PMID:24921110

  5. Genetic diversity and silencing suppression effects of Rice yellow mottle virus and the P1 protein

    PubMed Central

    Siré, Christelle; Bangratz-Reyser, Martine; Fargette, Denis; Brugidou, Christophe

    2008-01-01

    Background PTGS (post-transcriptional gene silencing) is used to counter pathogenic invasions, particularly viruses. In return, many plant viruses produce proteins which suppress silencing directed against their RNA. The diversity of silencing suppression at the species level in natural hosts is unknown. Results We investigated the functional diversity of silencing suppression among isolates of the African RYMV (Rice yellow mottle virus) in rice. The RYMV-P1 protein is responsible for cell-to-cell movement and is a silencing suppressor. Transgenic gus-silencing rice lines were used to investigate intra-specific and serogroup silencing suppression diversity at two different levels: that of the virion and the P1 silencing suppressor protein. Our data provide evidence that silencing suppression is a universal phenomenon for RYMV species. However, we found considerable diversity in their ability to suppress silencing which was not linked to RYMV phylogeny, or pathogenicity. At the level of the silencing suppressor P1 alone, we found similar results to those previously found at the virion level. In addition, we showed that cell-to-cell movement of P1 was crucial for the efficiency of silencing suppression. Mutagenesis of P1 demonstrated a strong link between some amino acids and silencing suppression features with, one on the hand, the conserved amino acids C95 and C64 involved in cell-to-cell movement and the strength of suppression, respectively, and on the other hand, the non conserved F88 was involved in the strength of silencing suppression. Conclusion We demonstrated that intra-species diversity of silencing suppression is highly variable and by mutagenesis of P1 we established the first link between silencing suppression and genetic diversity. These results are potentially important for understanding virus-host interactions. PMID:18447922

  6. Virus-Induced Gene Silencing in Hexaploid Wheat

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Functional genomics analysis in hexaploid wheat is greatly impeded by the genetic redundancy of polyploidy and the difficulties in generating large numbers of transgenic plants required in insertional mutagenesis strategies. Virus-induced gene silencing (VIGS), however, is a strategy for creating g...

  7. A reciprocal identity method for large silencer analysis

    NASA Astrophysics Data System (ADS)

    Zhou, L.; Wu, T. W.; Ruan, K.; Herrin, D. W.

    2016-03-01

    Conventional techniques used in the boundary element method for evaluating muffler transmission loss have been limited by the cutoff frequency of the inlet and outlet ducts. Even though the boundary element method itself is a truly three-dimensional analysis tool, it has not been effectively used on large silencers due to the large inlet and outlet cross sections. In this paper, a numerical technique based on the reciprocal identity and the boundary element impedance matrix is proposed as a post-processing filter to extract the transmission loss of large silencers at all frequencies. Each reciprocal identity couples two different sound fields on the same silencer geometry. The first sound field has the analytical modal expansion in the inlet and outlet ducts, while the second sound field is the boundary element solution associated with a random boundary condition set. Depending on how many modes exist in the inlet and outlet ducts at a certain frequency, a minimum number of random boundary condition sets must be applied to the boundary element model. The boundary element impedance matrix provides more than enough such solution sets for the reciprocal identity coupling. The overdetermined system is then solved by a least-squares procedure. The proposed method is verified by comparing to the analytical solutions of a simple expansion chamber and a round bar silencer.

  8. Strain Specific Factors Control Effector Gene Silencing in Phytophthora sojae

    PubMed Central

    Shrestha, Sirjana Devi; Chapman, Patrick; Zhang, Yun; Gijzen, Mark

    2016-01-01

    The Phytophthora sojae avirulence gene Avr3a encodes an effector that is capable of triggering immunity on soybean plants carrying the resistance gene Rps3a. P. sojae strains that express Avr3a are avirulent to Rps3a plants, while strains that do not are virulent. To study the inheritance of Avr3a expression and virulence towards Rps3a, genetic crosses and self-fertilizations were performed. A cross between P. sojae strains ACR10 X P7076 causes transgenerational gene silencing of Avr3a allele, and this effect is meiotically stable up to the F5 generation. However, test-crosses of F1 progeny (ACR10 X P7076) with strain P6497 result in the release of silencing of Avr3a. Expression of Avr3a in the progeny is variable and correlates with the phenotypic penetrance of the avirulence trait. The F1 progeny from a direct cross of P6497 X ACR10 segregate for inheritance for Avr3a expression, a result that could not be explained by parental imprinting or heterozygosity. Analysis of small RNA arising from the Avr3a gene sequence in the parental strains and hybrid progeny suggests that the presence of small RNA is necessary but not sufficient for gene silencing. Overall, we conclude that inheritance of the Avr3a gene silenced phenotype relies on factors that are variable among P. sojae strains. PMID:26930612

  9. Bureaucratic Constructions of Sexual Diversity: "Sensitive", "Controversial" and Silencing

    ERIC Educational Resources Information Center

    Ullman, Jacqueline; Ferfolja, Tania

    2015-01-01

    National research illustrates the high degree of discrimination that prevails against lesbian, gay, bisexual, transgender and queer (LGBTQ) students resulting in diminished educational outcomes, both academic and social. This phenomenon is influenced by the prevalence of whole-school silences around LGBTQ topics in many Australian schools. This…

  10. Love Styles and Self-Silencing in Romantic Relationships

    ERIC Educational Resources Information Center

    Collins, Kerry A.; Cramer, Kenneth M.; Singleton-Jackson, Jill A.

    2005-01-01

    Six love styles have been theorized to be related to several personality constructs (e.g., self-esteem) (Lee, 1973). Despite the interpersonal nature of love, investigations have yet to evaluate related variables and their association to love styles in romantic relationships. As a stable cognitive schema, silencing the self is proposed to account…

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

    PubMed Central

    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 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. PMID:27455310

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

    PubMed

    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 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. PMID:27455310

  13. Multiple roles for Piwi in silencing Drosophila transposons

    PubMed Central

    Rozhkov, Nikolay V.; Hammell, Molly; Hannon, Gregory J.

    2013-01-01

    Silencing of transposons in the Drosophila ovary relies on three Piwi family proteins—Piwi, Aubergine (Aub), and Ago3—acting in concert with their small RNA guides, the Piwi-interacting RNAs (piRNAs). Aub and Ago3 are found in the germ cell cytoplasm, where they function in the ping-pong cycle to consume transposon mRNAs. The nuclear Piwi protein is required for transposon silencing in both germ and somatic follicle cells, yet the precise mechanisms by which Piwi acts remain largely unclear. We investigated the role of Piwi by combining cell type-specific knockdowns with measurements of steady-state transposon mRNA levels, nascent RNA synthesis, chromatin state, and small RNA abundance. In somatic cells, Piwi loss led to concerted effects on nascent transcripts and transposon mRNAs, indicating that Piwi acts through transcriptional gene silencing (TGS). In germ cells, Piwi loss showed disproportionate impacts on steady-state RNA levels, indicating that it also exerts an effect on post-transcriptional gene silencing (PTGS). Piwi knockdown affected levels of germ cell piRNAs presumably bound to Aub and Ago3, perhaps explaining its post-transcriptional impacts. Overall, our results indicate that Piwi plays multiple roles in the piRNA pathway, in part enforcing transposon repression through effects on local chromatin states and transcription but also participating in germ cell piRNA biogenesis. PMID:23392609

  14. Breaking Classroom Silences: A View from Linguistic Ethnography

    ERIC Educational Resources Information Center

    Rampton, Ben; Charalambous, Constadina

    2016-01-01

    This paper addresses potentially problematic classroom episodes in which someone foregrounds a social division that is normally taken for granted. It illustrates the way in which linguistic ethnography can unpack the layered processes that collide in the breaking of silence, showing how linguistic form and practice, individual positioning, local…

  15. Organizational Justice As a Predictor of Organizational Silence

    ERIC Educational Resources Information Center

    Tan, Çetin

    2014-01-01

    In this study, relation between teachers' perception for organizational justice and their organizational silence was examined. Sample of this study consists of 300 teachers who work at elementary schools in Siirt. Relational Scanning model was utilized in performance of this study. In this study, Organizational Justice Scale and…

  16. Strain Specific Factors Control Effector Gene Silencing in Phytophthora sojae.

    PubMed

    Shrestha, Sirjana Devi; Chapman, Patrick; Zhang, Yun; Gijzen, Mark

    2016-01-01

    The Phytophthora sojae avirulence gene Avr3a encodes an effector that is capable of triggering immunity on soybean plants carrying the resistance gene Rps3a. P. sojae strains that express Avr3a are avirulent to Rps3a plants, while strains that do not are virulent. To study the inheritance of Avr3a expression and virulence towards Rps3a, genetic crosses and self-fertilizations were performed. A cross between P. sojae strains ACR10 X P7076 causes transgenerational gene silencing of Avr3a allele, and this effect is meiotically stable up to the F5 generation. However, test-crosses of F1 progeny (ACR10 X P7076) with strain P6497 result in the release of silencing of Avr3a. Expression of Avr3a in the progeny is variable and correlates with the phenotypic penetrance of the avirulence trait. The F1 progeny from a direct cross of P6497 X ACR10 segregate for inheritance for Avr3a expression, a result that could not be explained by parental imprinting or heterozygosity. Analysis of small RNA arising from the Avr3a gene sequence in the parental strains and hybrid progeny suggests that the presence of small RNA is necessary but not sufficient for gene silencing. Overall, we conclude that inheritance of the Avr3a gene silenced phenotype relies on factors that are variable among P. sojae strains. PMID:26930612

  17. Tissue homogeneity requires inhibition of unequal gene silencing during development.

    PubMed

    Le, Hai H; Looney, Monika; Strauss, Benjamin; Bloodgood, Michael; Jose, Antony M

    2016-08-01

    Multicellular organisms can generate and maintain homogenous populations of cells that make up individual tissues. However, cellular processes that can disrupt homogeneity and how organisms overcome such disruption are unknown. We found that ∼100-fold differences in expression from a repetitive DNA transgene can occur between intestinal cells in Caenorhabditis elegans These differences are caused by gene silencing in some cells and are actively suppressed by parental and zygotic factors such as the conserved exonuclease ERI-1. If unsuppressed, silencing can spread between some cells in embryos but can be repeat specific and independent of other homologous loci within each cell. Silencing can persist through DNA replication and nuclear divisions, disrupting uniform gene expression in developed animals. Analysis at single-cell resolution suggests that differences between cells arise during early cell divisions upon unequal segregation of an initiator of silencing. Our results suggest that organisms with high repetitive DNA content, which include humans, could use similar developmental mechanisms to achieve and maintain tissue homogeneity. PMID:27458132

  18. Design curves for circular and annular duct silencers

    NASA Technical Reports Server (NTRS)

    Watson, Willie R.; Ramakrishnan, R.

    1989-01-01

    Conventional models of sound propagation between porous walls (Scott, 1946) are adapted in order to calculate design curves for the lined circular and annular-duct silencers used in HVAC systems. The derivation of the governing equations is outlined, and results for two typical cases are presented graphically. Good agreement with published experimental data is demonstrated.

  19. Organizational Silence and Hidden Threats to Patient Safety

    PubMed Central

    Henriksen, Kerm; Dayton, Elizabeth

    2006-01-01

    Organizational silence refers to a collective-level phenomenon of saying or doing very little in response to significant problems that face an organization. The paper focuses on some of the less obvious factors contributing to organizational silence that can serve as threats to patient safety. Converging areas of research from the cognitive, social, and organizational sciences and the study of sociotechnical systems help to identify some of the underlying factors that serve to shape and sustain organizational silence. These factors have been organized under three levels of analysis: (1) individual factors, including the availability heuristic, self-serving bias, and the status quo trap; (2) social factors, including conformity, diffusion of responsibility, and microclimates of distrust; and (3) organizational factors, including unchallenged beliefs, the good provider fallacy, and neglect of the interdependencies. Finally, a new role for health care leaders and managers is envisioned. It is one that places high value on understanding system complexity and does not take comfort in organizational silence. PMID:16898978

  20. Virus-induced gene silencing (VIGS) in barley seedling leaves

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Virus-induced gene silencing (VIGS) is one of the most potent reverse genetics technologies for gene functional characterization. This method exploits a dsRNA-mediated antiviral defense mechanism in plants. Using this method allows researchers to generate rapid phenotypic data in a relatively rapid ...

  1. Breaking the Silence: Helping Battered Moms and Their Children.

    ERIC Educational Resources Information Center

    Texas Child Care, 2000

    2000-01-01

    Discusses issues of domestic violence to help child caregivers care for children more effectively. Considers the causes of domestic violence and its impact on children and families, and offers resources for more information as well as suggestions for helping an abused person get help. Includes lists of "Do's and don'ts for breaking the silence"…

  2. Silencing of an arabidopsis Myb regulatory transgene in tobacco

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We have been exploring the use of genetically engineered anthocyanin over-production as a visual indicator of gene activation and silencing in plants. Previous work demonstrated that constitutive over-expression of genes encoding specific Arabidopsis transcription factors from the myb family can pr...

  3. The Silenced Dialogue and Pre-Service Teachers

    ERIC Educational Resources Information Center

    Curry, Kristal

    2013-01-01

    In this article, the author reflects on the 1988 article "The Silenced Dialogue," by Lisa Delpit, which described the lack of communication dividing Black and White educators when it comes to the issue of race, specifically due to the disparity between reliance on theory (White) and reliance on cultural understanding (Black). Nearly a…

  4. When to Shut Students Up: Civility, Silencing, and Free Speech

    ERIC Educational Resources Information Center

    Callan, Eamonn

    2011-01-01

    Teachers sometimes shut students up for the sake of civility. My question is whether silencing for the sake of civility can be morally justified when a student derogates fellow students as members of some widely stigmatized group, and the offending speech is not for any further reason to be deplored, for example, as a personally targeted insult.…

  5. Breaking the Silence Surrounding Mental Health on Campus

    ERIC Educational Resources Information Center

    Roper, Larry D.

    2013-01-01

    Mentally ill students are able to participate in higher education at unprecedented rates. While colleges and universities have been responsive to the therapeutic needs, we have failed to successfully create supportive campus climates. Campus leaders are challenged to demonstrate ethical leadership that breaks the silence and confronts the stigma…

  6. Breaking the Silence: Facing Undocumented Issues in Teacher Practice

    ERIC Educational Resources Information Center

    Jefferies, Julián; Dabach, Dafney Blanca

    2014-01-01

    This conceptual article addresses the need for educators to interrupt status-quo silences surrounding the role of immigration status in schools--an issue that disproportionately impacts Latina/os. In this article we: (a) articulate the need for teacher education to address the impact of undocumented status in school settings; (b) present…

  7. Adjustable Optical-Fiber Attenuator

    NASA Technical Reports Server (NTRS)

    Buzzetti, Mike F.

    1994-01-01

    Adjustable fiber-optic attenuator utilizes bending loss to reduce strength of light transmitted along it. Attenuator functions without introducing measurable back-reflection or insertion loss. Relatively insensitive to vibration and changes in temperature. Potential applications include cable television, telephone networks, other signal-distribution networks, and laboratory instrumentation.

  8. Range Restriction and Attenuation Corrections.

    ERIC Educational Resources Information Center

    Mumford, Michael D.; Mendoza, Jorge L.

    The present paper reviews the techniques commonly used to correct an observed correlation coefficient for the simultaneous influence of attenuation and range restriction effects. It is noted that the procedure which is currently in use may be somewhat biased because it treats range restriction and attenuation as independent restrictive influences.…

  9. Suicide Risk: Amplifiers and Attenuators.

    ERIC Educational Resources Information Center

    Plutchik, Robert; Van Praag, Herman M.

    1994-01-01

    Attempts to integrate findings on correlates of suicide and violent risk in terms of a theory called a two-stage model of countervailing forces, which assumes that the strength of aggressive impulses is modified by amplifiers and attenuators. The vectorial interaction of amplifiers and attenuators creates an unstable equilibrium making prediction…

  10. Different patterns of gene silencing in position-effect variegation.

    PubMed

    Lloyd, Vett K; Dyment, David; Sinclair, Donald A R; Grigliatti, Thomas A

    2003-12-01

    Position-effect variegation (PEV) results when a fully functional gene is moved from its normal position to a position near to a broken heterochromatic-euchromatic boundary. In this new position, the gene, while remaining unaltered at the DNA level, is transcriptionally silenced in some cells but active in others, producing a diagnostic mosaic phenotype. Many variegating stocks show phenotypic instability, in that the level of variegation is dramatically different in different isolates or when out crossed. To test if this phenotypic instability was due to segregation of spontaneously accumulated mutations that suppress variegation, four different and well-characterized strains showing PEV for the white+ gene (wm4, wmMc, wm51b, and wmJ) and representing both large and small spot variegators were repeatedly out crossed to a strain free of modifiers, and the phenotypes of these variegators were monitored for 30 generations. Once free of modifiers, these variegating strains were then allowed to reaccumulate modifiers. The spontaneous suppressors of variegation were found to include both dominant and recessive, autosomal and X-linked alleles selected to reduce the detrimental effects of silencing white+ and adjacent genes. The time of peak sensitivity to temperature during development was also determined for these four variegators. Although large and small spot variegators have previously been attributed to early and late silencing events, respectively, the variegators we examined all shared a common early period of peak sensitivity to temperature. Once free of their variegation suppressors, the different variegating strains showed considerable differences in the frequency of inactivation at a cellular level (the number of cells showing silencing of a given gene) and the extent of variegation within the cell (the number of silenced genes). These results suggest that large and small spot variegation may be a superficial consequence of spontaneous variegation suppressors