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Sample records for reticulum segregate cell

  1. Endoplasmic reticulum involvement in yeast cell death

    International Nuclear Information System (INIS)

    Nicanor Austriaco, O.

    2012-01-01

    Yeast cells undergo programed cell death (PCD) with characteristic markers associated with apoptosis in mammalian cells including chromatin breakage, nuclear fragmentation, reactive oxygen species generation, and metacaspase activation. Though significant research has focused on mitochondrial involvement in this phenomenon, more recent work with both Saccharomyces cerevisiae and Schizosaccharomyces pombe has also implicated the endoplasmic reticulum (ER) in yeast PCD. This minireview provides an overview of ER stress-associated cell death (ER-SAD) in yeast. It begins with a description of ER structure and function in yeast before moving to a discussion of ER-SAD in both mammalian and yeast cells. Three examples of yeast cell death associated with the ER will be highlighted here including inositol starvation, lipid toxicity, and the inhibition of N-glycosylation. It closes by suggesting ways to further examine the involvement of the ER in yeast cell death.

  2. Polarity and reorganization of the endoplasmic reticulum during fertilization and ooplasmic segregation in the ascidian egg

    OpenAIRE

    1993-01-01

    During the first cell cycle of the ascidian egg, two phases of ooplasmic segregation create distinct cytoplasmic domains that are crucial for later development. We recently defined a domain enriched in ER in the vegetal region of Phallusia mammillata eggs. To explore the possible physiological and developmental function of this ER domain, we here investigate its organization and fate by labeling the ER network in vivo with DiIC16(3), and observing its distribution before and after fertilizati...

  3. Blood Cell Interactions and Segregation in Flow

    OpenAIRE

    Munn, Lance L.; Dupin, Michael M.

    2008-01-01

    For more than a century, pioneering researchers have been using novel experimental and computational approaches to probe the mysteries of blood flow. Thanks to their efforts, we know that blood cells generally prefer to migrate to the axis of flow, that red and white cells segregate in flow, and that cell deformability and their tendency to reversibly aggregate contribute to the non-Newtonian nature of this unique fluid. All of these properties have beneficial physiological consequences, allo...

  4. Analysis of endoplasmic reticulum of tobacco cells using confocal microscopy

    Czech Academy of Sciences Publication Activity Database

    Radochová, Barbora; Janáček, Jiří; Schwarzerová, K.; Demjénová, E.; Tomori, Z.; Karen, Petr; Kubínová, Lucie

    2005-01-01

    Roč. 24, č. 11 (2005), s. 181-185 ISSN 1580-3139 R&D Projects: GA AV ČR(CZ) KJB6011309 Institutional research plan: CEZ:AV0Z50110509 Keywords : confocal microscopy * endoplasmic reticulum * image analysis Subject RIV: EA - Cell Biology

  5. Blood cell interactions and segregation in flow.

    Science.gov (United States)

    Munn, Lance L; Dupin, Michael M

    2008-04-01

    For more than a century, pioneering researchers have been using novel experimental and computational approaches to probe the mysteries of blood flow. Thanks to their efforts, we know that blood cells generally prefer to migrate to the axis of flow, that red and white cells segregate in flow, and that cell deformability and their tendency to reversibly aggregate contribute to the non-Newtonian nature of this unique fluid. All of these properties have beneficial physiological consequences, allowing blood to perform a variety of critical functions. Our current understanding of these unusual flow properties of blood have been made possible by the ingenuity and diligence of a number of researchers, including Harry Goldsmith, who developed novel technologies to visualize and quantify the flow of blood at the level of individual cells. Here we summarize efforts in our lab to continue this tradition and to further our understanding of how blood cells interact with each other and with the blood vessel wall.

  6. Chromosome and cell wall segregation in Streptococcus faecium ATCC 9790

    International Nuclear Information System (INIS)

    Higgins, M.L.; Glaser, D.; Dicker, D.T.; Zito, E.T.

    1989-01-01

    Segregation was studied by measuring the positions of autoradiographic grain clusters in chains formed from single cells containing on average less than one radiolabeled chromosome strand. The degree to which chromosomal and cell wall material cosegregated was quantified by using the methods of S. Cooper and M. Weinberger, dividing the number of chains labeled at the middle. This analysis indicated that in contrast to chromosomal segregation in Escherichia coli and, in some studies, to that in gram-positive rods, chromosomal segregation in Streptococcus faecium was slightly nonrandom and did not vary with growth rate. Results were not significantly affected by strand exchange. In contrast, labeled cell wall segregated predominantly nonrandomly

  7. Observation of endoplasmic reticulum tubules via TOF-SIMS tandem mass spectrometry imaging of transfected cells.

    Science.gov (United States)

    Chini, Corryn E; Fisher, Gregory L; Johnson, Ben; Tamkun, Michael M; Kraft, Mary L

    2018-02-26

    Advances in three-dimensional secondary ion mass spectrometry (SIMS) imaging have enabled visualizing the subcellular distributions of various lipid species within individual cells. However, the difficulty of locating organelles using SIMS limits efforts to study their lipid compositions. Here, the authors have assessed whether endoplasmic reticulum (ER)-Tracker Blue White DPX ® , which is a commercially available stain for visualizing the endoplasmic reticulum using fluorescence microscopy, produces distinctive ions that can be used to locate the endoplasmic reticulum using SIMS. Time-of-flight-SIMS tandem mass spectrometry (MS 2 ) imaging was used to identify positively and negatively charged ions produced by the ER-Tracker stain. Then, these ions were used to localize the stain and thus the endoplasmic reticulum, within individual human embryonic kidney cells that contained higher numbers of endoplasmic reticulum-plasma membrane junctions on their surfaces. By performing MS 2 imaging of selected ions in parallel with the precursor ion (MS 1 ) imaging, the authors detected a chemical interference native to the cell at the same nominal mass as the pentafluorophenyl fragment from the ER-Tracker stain. Nonetheless, the fluorine secondary ions produced by the ER-Tracker stain provided a distinctive signal that enabled locating the endoplasmic reticulum using SIMS. This simple strategy for visualizing the endoplasmic reticulum in individual cells using SIMS could be combined with existing SIMS methodologies for imaging intracellular lipid distribution and to study the lipid composition within the endoplasmic reticulum.

  8. Endoplasmic Reticulum Stress, Unfolded Protein Response, and Cancer Cell Fate

    Directory of Open Access Journals (Sweden)

    Marco Corazzari

    2017-04-01

    Full Text Available Perturbation of endoplasmic reticulum (ER homeostasis results in a stress condition termed “ER stress” determining the activation of a finely regulated program defined as unfolded protein response (UPR and whose primary aim is to restore this organelle’s physiological activity. Several physiological and pathological stimuli deregulate normal ER activity causing UPR activation, such as hypoxia, glucose shortage, genome instability, and cytotoxic compounds administration. Some of these stimuli are frequently observed during uncontrolled proliferation of transformed cells, resulting in tumor core formation and stage progression. Therefore, it is not surprising that ER stress is usually induced during solid tumor development and stage progression, becoming an hallmark of such malignancies. Several UPR components are in fact deregulated in different tumor types, and accumulating data indicate their active involvement in tumor development/progression. However, although the UPR program is primarily a pro-survival process, sustained and/or prolonged stress may result in cell death induction. Therefore, understanding the mechanism(s regulating the cell survival/death decision under ER stress condition may be crucial in order to specifically target tumor cells and possibly circumvent or overcome tumor resistance to therapies. In this review, we discuss the role played by the UPR program in tumor initiation, progression and resistance to therapy, highlighting the recent advances that have improved our understanding of the molecular mechanisms that regulate the survival/death switch.

  9. The endoplasmic reticulum in plant immunity and cell death.

    Science.gov (United States)

    Eichmann, Ruth; Schäfer, Patrick

    2012-01-01

    The endoplasmic reticulum (ER) is a highly dynamic organelle in eukaryotic cells and a major production site of proteins destined for vacuoles, the plasma membrane, or apoplast in plants. At the ER, these secreted proteins undergo multiple processing steps, which are supervised and conducted by the ER quality control system. Notably, processing of secreted proteins can considerably elevate under stress conditions and exceed ER folding capacities. The resulting accumulation of unfolded proteins is defined as ER stress. The efficiency of cells to re-establish proper ER function is crucial for stress adaptation. Besides delivering proteins directly antagonizing and resolving stress conditions, the ER monitors synthesis of immune receptors. This indicates the significance of the ER for the establishment and function of the plant immune system. Recent studies point out the fragility of the entire system and highlight the ER as initiator of programed cell death (PCD) in plants as was reported for vertebrates. This review summarizes current knowledge on the impact of the ER on immune and PCD signaling. Understanding the integration of stress signals by the ER bears a considerable potential to optimize development and to enhance stress resistance of plants.

  10. The endoplasmic reticulum in plant immunity and cell death

    Directory of Open Access Journals (Sweden)

    Patrick eSchäfer

    2012-08-01

    Full Text Available The endoplasmic reticulum (ER is a highly dynamic organelle in eukaryotic cells and a major production site of proteins destined for vacuoles, the plasma membrane or apoplast in plants. At the ER, these secreted proteins undergo multiple processing steps, which are supervised and conducted by the ER quality control system. Notably, processing of secreted proteins can considerably elevate under stress conditions and exceed ER folding capacities. The resulting accumulation of unfolded proteins is defined as ER stress. The efficiency of cells to re-establish proper ER function is crucial for stress adaptation. Besides delivering proteins directly antagonizing and resolving stress conditions, the ER monitors synthesis of immune receptors. This indicates the significance of the ER for the establishment and function of the plant immune system. Recent studies point out the fragility of the entire system and highlight the ER as initiator of programmed cell death (PCD in plants as was reported for vertebrates. This review summarizes current knowledge on the impact of the ER on immune and PCD signalling. Understanding the integration of stress signals by the ER bears a considerable potential to optimize development and to enhance stress resistance of plants.

  11. Non-Random Chromosome Segregation in Stem Cells

    OpenAIRE

    Conboy, Michael J; Karasov, Ariela O; Rando, Thomas A

    2007-01-01

    Decades ago, the "immortal strand hypothesis" was proposed as a means by which stem cells might limit acquiring mutations that could give rise to cancer, while continuing to proliferate for the life of an organism. Originally based on observations in embryonic cells, and later studied in terms of stem cell self-renewal, this hypothesis has remained largely unaccepted because of few additional reports, the rarity of the cells displaying template strand segregation, and alternative interpretati...

  12. Preconditioning with endoplasmic reticulum stress ameliorates endothelial cell inflammation.

    Science.gov (United States)

    Leonard, Antony; Paton, Adrienne W; El-Quadi, Monaliza; Paton, James C; Fazal, Fabeha

    2014-01-01

    Endoplasmic Reticulum (ER) stress, caused by disturbance in ER homeostasis, has been implicated in several pathological conditions such as ischemic injury, neurodegenerative disorders, metabolic diseases and more recently in inflammatory conditions. Our present study aims at understanding the role of ER stress in endothelial cell (EC) inflammation, a critical event in the pathogenesis of acute lung injury (ALI). We found that preconditioning human pulmonary artery endothelial cells (HPAEC) to ER stress either by depleting ER chaperone and signaling regulator BiP using siRNA, or specifically cleaving (inactivating) BiP using subtilase cytotoxin (SubAB), alleviates EC inflammation. The two approaches adopted to abrogate BiP function induced ATF4 protein expression and the phosphorylation of eIF2α, both markers of ER stress, which in turn resulted in blunting the activation of NF-κB, and restoring endothelial barrier integrity. Pretreatment of HPAEC with BiP siRNA inhibited thrombin-induced IκBα degradation and its resulting downstream signaling pathway involving NF-κB nuclear translocation, DNA binding, phosphorylation at serine536, transcriptional activation and subsequent expression of adhesion molecules. However, TNFα-mediated NF-κB signaling was unaffected upon BiP knockdown. In an alternative approach, SubAB-mediated inactivation of NF-κB was independent of IκBα degradation. Mechanistic analysis revealed that pretreatment of EC with SubAB interfered with the binding of the liberated NF-κB to the DNA, thereby resulting in reduced expression of adhesion molecules, cytokines and chemokines. In addition, both knockdown and inactivation of BiP stimulated actin cytoskeletal reorganization resulting in restoration of endothelial permeability. Together our studies indicate that BiP plays a central role in EC inflammation and injury via its action on NF-κB activation and regulation of vascular permeability.

  13. Evolutionary dynamics of adult stem cells: Comparison of random and immortal strand segregation mechanisms

    OpenAIRE

    Tannenbaum, Emmanuel; Sherley, James L.; Shakhnovich, Eugene I.

    2004-01-01

    This paper develops a point-mutation model describing the evolutionary dynamics of a population of adult stem cells. Such a model may prove useful for quantitative studies of tissue aging and the emergence of cancer. We consider two modes of chromosome segregation: (1) Random segregation, where the daughter chromosomes of a given parent chromosome segregate randomly into the stem cell and its differentiating sister cell. (2) ``Immortal DNA strand'' co-segregation, for which the stem cell reta...

  14. Endoplasmic reticulum redox state is not perturbed by pharmacological or pathological endoplasmic reticulum stress in live pancreatic β-cells.

    Directory of Open Access Journals (Sweden)

    Irmgard Schuiki

    Full Text Available Accumulation of unfolded, misfolded and aggregated proteins in the endoplasmic reticulum (ER causes ER stress. ER stress can result from physiological situations such as acute increases in secretory protein biosynthesis or pathological conditions that perturb ER homeostasis such as alterations in the ER redox state. Here we monitored ER redox together with transcriptional output of the Unfolded Protein Response (UPR in INS-1 insulinoma cells stably expressing eroGFP (ER-redox-sensor and mCherry protein driven by a GRP78 promoter (UPR-sensor. Live cell imaging, flow cytometry and biochemical characterization were used to examine these parameters in response to various conditions known to induce ER stress. As expected, treatment of the cells with the reducing agent dithiothreitol caused a decrease in the oxidation state of the ER accompanied by an increase in XBP-1 splicing. Unexpectedly however, other treatments including tunicamycin, thapsigargin, DL-homocysteine, elevated free fatty acids or high glucose had essentially no influence on the ER redox state, despite inducing ER stress. Comparable results were obtained with dispersed rat islet cells expressing eroGFP. Thus, unlike in yeast cells, ER stress in pancreatic β-cells is not associated with a more reducing ER environment.

  15. Immune responsiveness and incidence of reticulum cell sarcoma in long-term syngeneic radiation chimeras

    International Nuclear Information System (INIS)

    Adorini, L.; Gorini, G.; Covelli, V.; Ballardin, E.; di Michele, A.; Bassani, B.; Metalli, P.; Doria, G.

    1976-01-01

    Long-term syngeneic radiation chimeras displayed a very low incidence of reticulum cell sarcoma as compared with control mice. Immune reactivity of these animals was studied in vivo by anti-dinitrophenyl antibody titer and affinity and in vitro by mitotic responsiveness to phytohemagglutinin, concanavalin A and lipopolysaccharide. Antibody titer and affinity as well as the response to T lectins were found to be increased in chimeras. These results were attributed to increased function of mature T2 cells, which could explain the reduced incidence of reticulum cell sarcoma in chimeras

  16. Stem cell identity and template DNA strand segregation.

    Science.gov (United States)

    Tajbakhsh, Shahragim

    2008-12-01

    The quest for stem cell properties to distinguish their identity from that of committed daughters has led to a re-investigation of the notion that DNA strands are not equivalent, and 'immortal' DNA strands are retained in stem cells whereas newly replicated DNA strands segregate to the differentiating daughter cell during mitosis. Whether this process occurs only in stem cells, and also in all tissues, remains unclear. That individual chromosomes can be also partitioned non-randomly raises the question if this phenomenon is related to the immortal DNA hypothesis, and it underscores the need for high-resolution techniques to observe these events empirically. Although initially postulated as a mechanism to avoid DNA replication errors, alternative views including epigenetic regulation and sister chromatid silencing may provide insights into this process.

  17. Organization of the cytoplasmic reticulum in the central vacuole of parenchyma cells in Allium cepa L.

    Directory of Open Access Journals (Sweden)

    Tomasz J. Wodzicki

    2015-01-01

    Full Text Available An elaborate and complex cytoplasmic reticulum composed of fine filaments and lamellae ranging from 0.1 to 4 microns in size is revealed by viewing the central vacuole of onion bulb parenchyma cells with the scanning election microscope. The larger cytoplasmic strands, visible with the light microscope, are composed of numerous smaller filaments (some tubular which might explain the observed bidirectional movement of particles in these larger strands. The finely divided cytoplasmic network of filaments is continuous with the parietal cytoplasm inclosing the vacuolar sap. In these highly vacuolated cells the mass of the protoplast is in the form of an intravacuolar reticulum immersed in the cell sap. The probable significance of the vacuolar sap in relation to physiological processes of the cell is discussed.

  18. Titanium Dioxide Nanoparticles Induce Endoplasmic Reticulum Stress-Mediated Autophagic Cell Death via Mitochondria-Associated Endoplasmic Reticulum Membrane Disruption in Normal Lung Cells

    Science.gov (United States)

    Yu, Kyeong-Nam; Chang, Seung-Hee; Park, Soo Jin; Lim, Joohyun; Lee, Jinkyu; Yoon, Tae-Jong; Kim, Jun-Sung; Cho, Myung-Haing

    2015-01-01

    Nanomaterials are used in diverse fields including food, cosmetic, and medical industries. Titanium dioxide nanoparticles (TiO2-NP) are widely used, but their effects on biological systems and mechanism of toxicity have not been elucidated fully. Here, we report the toxicological mechanism of TiO2-NP in cell organelles. Human bronchial epithelial cells (16HBE14o-) were exposed to 50 and 100 μg/mL TiO2-NP for 24 and 48 h. Our results showed that TiO2-NP induced endoplasmic reticulum (ER) stress in the cells and disrupted the mitochondria-associated endoplasmic reticulum membranes (MAMs) and calcium ion balance, thereby increasing autophagy. In contrast, an inhibitor of ER stress, tauroursodeoxycholic acid (TUDCA), mitigated the cellular toxic response, suggesting that TiO2-NP promoted toxicity via ER stress. This novel mechanism of TiO2-NP toxicity in human bronchial epithelial cells suggests that further exhaustive research on the harmful effects of these nanoparticles in relevant organisms is needed for their safe application. PMID:26121477

  19. Sodium Butyrate Induces Endoplasmic Reticulum Stress and Autophagy in Colorectal Cells: Implications for Apoptosis.

    Directory of Open Access Journals (Sweden)

    Jintao Zhang

    Full Text Available Butyrate, a short-chain fatty acid derived from dietary fiber, inhibits proliferation and induces cell death in colorectal cancer cells. However, clinical trials have shown mixed results regarding the anti-tumor activities of butyrate. We have previously shown that sodium butyrate increases endoplasmic reticulum stress by altering intracellular calcium levels, a well-known autophagy trigger. Here, we investigated whether sodium butyrate-induced endoplasmic reticulum stress mediated autophagy, and whether there was crosstalk between autophagy and the sodium butyrate-induced apoptotic response in human colorectal cancer cells.Human colorectal cancer cell lines (HCT-116 and HT-29 were treated with sodium butyrate at concentrations ranging from 0.5-5mM. Cell proliferation was assessed using MTT tetrazolium salt formation. Autophagy induction was confirmed through a combination of Western blotting for associated proteins, acridine orange staining for acidic vesicles, detection of autolysosomes (MDC staining, and electron microscopy. Apoptosis was quantified by flow cytometry using standard annexinV/propidium iodide staining and by assessing PARP-1 cleavage by Western blot.Sodium butyrate suppressed colorectal cancer cell proliferation, induced autophagy, and resulted in apoptotic cell death. The induction of autophagy was supported by the accumulation of acidic vesicular organelles and autolysosomes, and the expression of autophagy-associated proteins, including microtubule-associated protein II light chain 3 (LC3-II, beclin-1, and autophagocytosis-associated protein (Atg3. The autophagy inhibitors 3-methyladenine (3-MA and chloroquine inhibited sodium butyrate induced autophagy. Furthermore, sodium butyrate treatment markedly enhanced the expression of endoplasmic reticulum stress-associated proteins, including BIP, CHOP, PDI, and IRE-1a. When endoplasmic reticulum stress was inhibited by pharmacological (cycloheximide and mithramycin and genetic

  20. Lipolysis Response to Endoplasmic Reticulum Stress in Adipose Cells*

    Science.gov (United States)

    Deng, Jingna; Liu, Shangxin; Zou, Liangqiang; Xu, Chong; Geng, Bin; Xu, Guoheng

    2012-01-01

    In obesity and diabetes, adipocytes show significant endoplasmic reticulum (ER) stress, which triggers a series of responses. This study aimed to investigate the lipolysis response to ER stress in rat adipocytes. Thapsigargin, tunicamycin, and brefeldin A, which induce ER stress through different pathways, efficiently activated a time-dependent lipolytic reaction. The lipolytic effect of ER stress occurred with elevated cAMP production and protein kinase A (PKA) activity. Inhibition of PKA reduced PKA phosphosubstrates and attenuated the lipolysis. Although both ERK1/2 and JNK are activated during ER stress, lipolysis is partially suppressed by inhibiting ERK1/2 but not JNK and p38 MAPK and PKC. Thus, ER stress induces lipolysis by activating cAMP/PKA and ERK1/2. In the downstream lipolytic cascade, phosphorylation of lipid droplet-associated protein perilipin was significantly promoted during ER stress but attenuated on PKA inhibition. Furthermore, ER stress stimuli did not alter the levels of hormone-sensitive lipase and adipose triglyceride lipase but caused Ser-563 and Ser-660 phosphorylation of hormone-sensitive lipase and moderately elevated its translocation from the cytosol to lipid droplets. Accompanying these changes, total activity of cellular lipases was promoted to confer the lipolysis. These findings suggest a novel pathway of the lipolysis response to ER stress in adipocytes. This lipolytic activation may be an adaptive response that regulates energy homeostasis but with sustained ER stress challenge could contribute to lipotoxicity, dyslipidemia, and insulin resistance because of persistently accelerated free fatty acid efflux from adipocytes to the bloodstream and other tissues. PMID:22223650

  1. A turn-on fluorescent probe for endogenous formaldehyde in the endoplasmic reticulum of living cells

    Science.gov (United States)

    Tang, Yonghe; Ma, Yanyan; Xu, An; Xu, Gaoping; Lin, Weiying

    2017-06-01

    As the simplest aldehyde compounds, formaldehyde (FA) is implicated in nervous system diseases and cancer. Endoplasmic reticulum is an organelle that plays important functions in living cells. Accordingly, the development of efficient methods for FA detection in the endoplasmic reticulum (ER) is of great biomedical importance. In this work, we developed the first ER-targeted fluorescent FA probe Na-FA-ER. The detection is based on the condensation reaction of the hydrazine group and FA to suppress the photo-induced electron transfer (PET) pathway, resulting in a fluorescence increase. The novel Na-FA-ER showed high sensitivity to FA. In addition, the Na-FA-ER enabled the bio-imaging of exogenous and endogenous FA in living HeLa cells. Most significantly, the new Na-FA-ER was employed to visualize the endogenous FA in the ER in living cells for the first time.

  2. The endoplasmic reticulum exerts control over organelle streaming during cell expansion.

    Science.gov (United States)

    Stefano, Giovanni; Renna, Luciana; Brandizzi, Federica

    2014-03-01

    Cytoplasmic streaming is crucial for cell homeostasis and expansion but the precise driving forces are largely unknown. In plants, partial loss of cytoplasmic streaming due to chemical and genetic ablation of myosins supports the existence of yet-unknown motors for organelle movement. Here we tested a role of the endoplasmic reticulum (ER) as propelling force for cytoplasmic streaming during cell expansion. Through quantitative live-cell analyses in wild-type Arabidopsis thaliana cells and mutants with compromised ER structure and streaming, we demonstrate that cytoplasmic streaming undergoes profound changes during cell expansion and that it depends on motor forces co-exerted by the ER and the cytoskeleton.

  3. Prodigiosin activates endoplasmic reticulum stress cell death pathway in human breast carcinoma cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Pan, Mu-Yun [Institute of Biomedical Sciences, National Chung Hsing University, Taichung, Taiwan (China); Shen, Yuh-Chiang [Institute of Biomedical Sciences, National Chung Hsing University, Taichung, Taiwan (China); National Research Institute of Chinese Medicine, Taipei, Taiwan (China); Lu, Chien-Hsing [Department of Obstetrics and Gynecology, Taichung Veterans General Hospital, Taichung, Taiwan (China); Department of Obstetrics and Gynecology, National Yang-Ming University School of Medicine, Taipei, Taiwan (China); Yang, Shu-Yi [Institute of Biomedical Sciences, National Chung Hsing University, Taichung, Taiwan (China); Ho, Tsing-Fen [Department of Medical Laboratory Science and Biotechnology, Central Taiwan University of Science and Technology, Taichung, Taiwan (China); Peng, Yu-Ta [Institute of Biomedical Sciences, National Chung Hsing University, Taichung, Taiwan (China); Chang, Chia-Che, E-mail: chia_che@dragon.nchu.edu.tw [Institute of Biomedical Sciences, National Chung Hsing University, Taichung, Taiwan (China); Agricultural Biotechnology Center, National Chung Hsing University, Taichung, Taiwan (China); Graduate Institute of Basic Medical Science, China Medical University, Taichung, Taiwan (China)

    2012-12-15

    Prodigiosin is a bacterial tripyrrole pigment with potent cytotoxicity against diverse human cancer cell lines. Endoplasmic reticulum (ER) stress is initiated by accumulation of unfolded or misfolded proteins in the ER lumen and may induce cell death when irremediable. In this study, the role of ER stress in prodigiosin-induced cytotoxicity was elucidated for the first time. Comparable to the ER stress inducer thapsigargin, prodigiosin up-regulated signature ER stress markers GRP78 and CHOP in addition to activating the IRE1, PERK and ATF6 branches of the unfolded protein response (UPR) in multiple human breast carcinoma cell lines, confirming prodigiosin as an ER stress inducer. Prodigiosin transcriptionally up-regulated CHOP, as evidenced by its promoting effect on the CHOP promoter activity. Of note, knockdown of CHOP effectively lowered prodigiosin's capacity to evoke PARP cleavage, reduce cell viability and suppress colony formation, highlighting an essential role of CHOP in prodigiosin-induced cytotoxic ER stress response. In addition, prodigiosin down-regulated BCL2 in a CHOP-dependent manner. Importantly, restoration of BCL2 expression blocked prodigiosin-induced PARP cleavage and greatly enhanced the survival of prodigiosin-treated cells, suggesting that CHOP-dependent BCL2 suppression mediates prodigiosin-elicited cell death. Moreover, pharmacological inhibition of JNK by SP600125 or dominant-negative blockade of PERK-mediated eIF2α phosphorylation impaired prodigiosin-induced CHOP up-regulation and PARP cleavage. Collectively, these results identified ER stress-mediated cell death as a mode-of-action of prodigiosin's tumoricidal effect. Mechanistically, prodigiosin engages the IRE1–JNK and PERK–eIF2α branches of the UPR signaling to up-regulate CHOP, which in turn mediates BCL2 suppression to induce cell death. Highlights: ► Prodigiosin is a bacterial tripyrrole pigment with potent anticancer effect. ► Prodigiosin is herein identified

  4. Prodigiosin activates endoplasmic reticulum stress cell death pathway in human breast carcinoma cell lines

    International Nuclear Information System (INIS)

    Pan, Mu-Yun; Shen, Yuh-Chiang; Lu, Chien-Hsing; Yang, Shu-Yi; Ho, Tsing-Fen; Peng, Yu-Ta; Chang, Chia-Che

    2012-01-01

    Prodigiosin is a bacterial tripyrrole pigment with potent cytotoxicity against diverse human cancer cell lines. Endoplasmic reticulum (ER) stress is initiated by accumulation of unfolded or misfolded proteins in the ER lumen and may induce cell death when irremediable. In this study, the role of ER stress in prodigiosin-induced cytotoxicity was elucidated for the first time. Comparable to the ER stress inducer thapsigargin, prodigiosin up-regulated signature ER stress markers GRP78 and CHOP in addition to activating the IRE1, PERK and ATF6 branches of the unfolded protein response (UPR) in multiple human breast carcinoma cell lines, confirming prodigiosin as an ER stress inducer. Prodigiosin transcriptionally up-regulated CHOP, as evidenced by its promoting effect on the CHOP promoter activity. Of note, knockdown of CHOP effectively lowered prodigiosin's capacity to evoke PARP cleavage, reduce cell viability and suppress colony formation, highlighting an essential role of CHOP in prodigiosin-induced cytotoxic ER stress response. In addition, prodigiosin down-regulated BCL2 in a CHOP-dependent manner. Importantly, restoration of BCL2 expression blocked prodigiosin-induced PARP cleavage and greatly enhanced the survival of prodigiosin-treated cells, suggesting that CHOP-dependent BCL2 suppression mediates prodigiosin-elicited cell death. Moreover, pharmacological inhibition of JNK by SP600125 or dominant-negative blockade of PERK-mediated eIF2α phosphorylation impaired prodigiosin-induced CHOP up-regulation and PARP cleavage. Collectively, these results identified ER stress-mediated cell death as a mode-of-action of prodigiosin's tumoricidal effect. Mechanistically, prodigiosin engages the IRE1–JNK and PERK–eIF2α branches of the UPR signaling to up-regulate CHOP, which in turn mediates BCL2 suppression to induce cell death. Highlights: ► Prodigiosin is a bacterial tripyrrole pigment with potent anticancer effect. ► Prodigiosin is herein identified as an

  5. Trichodermin induces cell apoptosis through mitochondrial dysfunction and endoplasmic reticulum stress in human chondrosarcoma cells

    International Nuclear Information System (INIS)

    Su, Chen-Ming; Wang, Shih-Wei; Lee, Tzong-Huei; Tzeng, Wen-Pei; Hsiao, Che-Jen; Liu, Shih-Chia; Tang, Chih-Hsin

    2013-01-01

    Chondrosarcoma is the second most common primary bone tumor, and it responds poorly to both chemotherapy and radiation treatment. Nalanthamala psidii was described originally as Myxosporium in 1926. This is the first study to investigate the anti-tumor activity of trichodermin (trichothec-9-en-4-ol, 12,13-epoxy-, acetate), an endophytic fungal metabolite from N. psidii against human chondrosarcoma cells. We demonstrated that trichodermin induced cell apoptosis in human chondrosarcoma cell lines (JJ012 and SW1353 cells) instead of primary chondrocytes. In addition, trichodermin triggered endoplasmic reticulum (ER) stress protein levels of IRE1, p-PERK, GRP78, and GRP94, which were characterized by changes in cytosolic calcium levels. Furthermore, trichodermin induced the upregulation of Bax and Bid, the downregulation of Bcl-2, and the dysfunction of mitochondria, which released cytochrome c and activated caspase-3 in human chondrosarcoma. In addition, animal experiments illustrated reduced tumor volume, which led to an increased number of terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL)-positive cells and an increased level of cleaved PARP protein following trichodermin treatment. Together, this study demonstrates that trichodermin is a novel anti-tumor agent against human chondrosarcoma cells both in vitro and in vivo via mitochondrial dysfunction and ER stress. - Highlights: • Trichodermin induces chondrosarcoma apoptosis. • ER stress is involved in trichodermin-induced cell death. • Trichodermin induces chondrosarcoma death in vivo.

  6. Trichodermin induces cell apoptosis through mitochondrial dysfunction and endoplasmic reticulum stress in human chondrosarcoma cells

    Energy Technology Data Exchange (ETDEWEB)

    Su, Chen-Ming [Graduate Institute of Basic Medical Science, China Medical University, Taichung, Taiwan (China); Wang, Shih-Wei [Department of Medicine, Mackay Medical College, New Taipei City, Taiwan (China); Lee, Tzong-Huei [Graduate Institute of Pharmacognosy, Taipei Medical University, Taipei, Taiwan (China); Tzeng, Wen-Pei [Graduate Institute of Sports and Health, National Changhua University of Education, Changhua, Taiwan (China); Hsiao, Che-Jen [School of Respiratory Therapy, College of Medicine, Taipei Medical University, Taipei, Taiwan (China); Liu, Shih-Chia [Department of Orthopaedics, Mackay Memorial Hospital, Taipei, Taiwan (China); Tang, Chih-Hsin, E-mail: chtang@mail.cmu.edu.tw [Graduate Institute of Basic Medical Science, China Medical University, Taichung, Taiwan (China); Department of Pharmacology, School of Medicine, China Medical University, Taichung, Taiwan (China); Department of Biotechnology, College of Health Science, Asia University, Taichung, Taiwan (China)

    2013-10-15

    Chondrosarcoma is the second most common primary bone tumor, and it responds poorly to both chemotherapy and radiation treatment. Nalanthamala psidii was described originally as Myxosporium in 1926. This is the first study to investigate the anti-tumor activity of trichodermin (trichothec-9-en-4-ol, 12,13-epoxy-, acetate), an endophytic fungal metabolite from N. psidii against human chondrosarcoma cells. We demonstrated that trichodermin induced cell apoptosis in human chondrosarcoma cell lines (JJ012 and SW1353 cells) instead of primary chondrocytes. In addition, trichodermin triggered endoplasmic reticulum (ER) stress protein levels of IRE1, p-PERK, GRP78, and GRP94, which were characterized by changes in cytosolic calcium levels. Furthermore, trichodermin induced the upregulation of Bax and Bid, the downregulation of Bcl-2, and the dysfunction of mitochondria, which released cytochrome c and activated caspase-3 in human chondrosarcoma. In addition, animal experiments illustrated reduced tumor volume, which led to an increased number of terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL)-positive cells and an increased level of cleaved PARP protein following trichodermin treatment. Together, this study demonstrates that trichodermin is a novel anti-tumor agent against human chondrosarcoma cells both in vitro and in vivo via mitochondrial dysfunction and ER stress. - Highlights: • Trichodermin induces chondrosarcoma apoptosis. • ER stress is involved in trichodermin-induced cell death. • Trichodermin induces chondrosarcoma death in vivo.

  7. THE RESPONSE OF DISSEMINATED RETICULUM CELL SARCOMA TO THE INTRAVENOUS INJECTION OF COLLOIDAL RADIOACTIVE GOLD

    Energy Technology Data Exchange (ETDEWEB)

    Rubin, Philip; Levitt, Seymour H.

    1963-06-15

    Case histories of two patients treated with colloidal radiogold for diffuse reticulum cell sarcoma are presented. Further analysis of the method is suggested by the unusually long survival time of one of the patients. It was concluded that, although external radiotherapy remains the treatment of choice in localized reticulum cell sarcoma, intravenous colloidal radiogold may be a useful agent in lymphosarcomas with diffuse minute neoplastic liver and spleen involvements. Intravenous colloidal radiogold can produce bone marrow depression and thrombocytopenia which can lead to death. This factor tends to argue against therapeutic use of the agent. It is suggested that no more than 50 mC Au/sup 198/ intravenously should be used for treatment of this disease. (R.M.G.)

  8. Acyl-CoA-binding protein (ACBP) localizes to the endoplasmic reticulum and Golgi in a ligand-dependent manner in mammalian cells

    DEFF Research Database (Denmark)

    Hansen, Jesper S; Færgeman, Nils J; Kragelund, Birthe B

    2008-01-01

    showed that ACBP targeted to the ER (endoplasmic reticulum) and Golgi in a ligand-binding-dependent manner. A variant Y28F/K32A-FACI-50, which is unable to bind acyl-CoA, did no longer show association with the ER and became segregated from the Golgi, as analysed by intensity correlation calculations....... Depletion of fatty acids from cells by addition of FAFBSA (fatty-acid-free BSA) significantly decreased FACI-50 association with the Golgi, whereas fatty acid overloading increased Golgi association, strongly supporting that ACBP associates with the Golgi in a ligand-dependent manner. FRAP (fluorescence...... recovery after photobleaching) showed that the fatty-acid-induced targeting of FACI-50 to the Golgi resulted in a 5-fold reduction in FACI-50 mobility. We suggest that ACBP is targeted to the ER and Golgi in a ligand-binding-dependent manner in living cells and propose that ACBP may be involved...

  9. Evolutionary dynamics of adult stem cells: comparison of random and immortal-strand segregation mechanisms.

    Science.gov (United States)

    Tannenbaum, Emmanuel; Sherley, James L; Shakhnovich, Eugene I

    2005-04-01

    This paper develops a point-mutation model describing the evolutionary dynamics of a population of adult stem cells. Such a model may prove useful for quantitative studies of tissue aging and the emergence of cancer. We consider two modes of chromosome segregation: (1) random segregation, where the daughter chromosomes of a given parent chromosome segregate randomly into the stem cell and its differentiating sister cell and (2) "immortal DNA strand" co-segregation, for which the stem cell retains the daughter chromosomes with the oldest parent strands. Immortal strand co-segregation is a mechanism, originally proposed by [Cairns Nature (London) 255, 197 (1975)], by which stem cells preserve the integrity of their genomes. For random segregation, we develop an ordered strand pair formulation of the dynamics, analogous to the ordered strand pair formalism developed for quasispecies dynamics involving semiconservative replication with imperfect lesion repair (in this context, lesion repair is taken to mean repair of postreplication base-pair mismatches). Interestingly, a similar formulation is possible with immortal strand co-segregation, despite the fact that this segregation mechanism is age dependent. From our model we are able to mathematically show that, when lesion repair is imperfect, then immortal strand co-segregation leads to better preservation of the stem cell lineage than random chromosome segregation. Furthermore, our model allows us to estimate the optimal lesion repair efficiency for preserving an adult stem cell population for a given period of time. For human stem cells, we obtain that mispaired bases still present after replication and cell division should be left untouched, to avoid potentially fixing a mutation in both DNA strands.

  10. Pekinenin E Inhibits the Growth of Hepatocellular Carcinoma by Promoting Endoplasmic Reticulum Stress Mediated Cell Death

    Directory of Open Access Journals (Sweden)

    Lu Fan

    2017-06-01

    Full Text Available Hepatocellular carcinoma (HCC is a malignant primary liver cancer with poor prognosis. In the present study, we report that pekinenin E (PE, a casbane diterpenoid derived from the roots of Euphorbia pekinensis, has a strong antitumor activity against human HCC cells both in vitro and in vivo. PE suppressed the growth of human HCC cells Hep G2 and SMMC-7721. In addition, PE-mediated endoplasmic reticulum (ER stress caused increasing expressions of C/EBP homologous protein (CHOP, leading to apoptosis in HCC cells both in vitro and in vivo. Inhibition of ER stress with CHOP small interfering RNA or 4-phenyl-butyric acid partially reversed PE-induced cell death. Furthermore, PE induced S cell cycle arrest, which could also be partially reversed by CHOP knockdown. In all, these findings suggest that PE causes ER stress-associated cell death and cell cycle arrest, and it may serve as a potent agent for curing human HCC.

  11. Endoplasmic reticulum calcium transport ATPase expression during differentiation of colon cancer and leukaemia cells

    International Nuclear Information System (INIS)

    Papp, Bela; Brouland, Jean-Philippe; Gelebart, Pascal; Kovacs, Tuende; Chomienne, Christine

    2004-01-01

    The calcium homeostasis of the endoplasmic reticulum (ER) is connected to a multitude of cell functions involved in intracellular signal transduction, control of proliferation, programmed cell death, or the synthesis of mature proteins. Calcium is accumulated in the ER by various biochemically distinct sarco/endoplasmic reticulum calcium transport ATPase isoenzymes (SERCA isoforms). Experimental data indicate that the SERCA composition of some carcinoma and leukaemia cell types undergoes significant changes during differentiation, and that this is accompanied by modifications of SERCA-dependent calcium accumulation in the ER. Because ER calcium homeostasis can also influence cell differentiation, we propose that the modulation of the expression of various SERCA isoforms, and in particular, the induction of the expression of SERCA3-type proteins, is an integral part of the differentiation program of some cancer and leukaemia cell types. The SERCA content of the ER may constitute a new parameter by which the calcium homeostatic characteristics of the organelle are adjusted. The cross-talk between ER calcium homeostasis and cell differentiation may have some implications for the better understanding of the signalling defects involved in the acquisition and maintenance of the malignant phenotype

  12. Involvement of Endoplasmic Reticulum Stress in Capsaicin-Induced Apoptosis of Human Pancreatic Cancer Cells

    Directory of Open Access Journals (Sweden)

    Shengzhang Lin

    2013-01-01

    Full Text Available Capsaicin, main pungent ingredient of hot chilli peppers, has been shown to have anticarcinogenic effect on various cancer cells through multiple mechanisms. In this study, we investigated the apoptotic effect of capsaicin on human pancreatic cancer cells in both in vitro and in vivo systems, as well as the possible mechanisms involved. In vitro, treatment of both the pancreatic cancer cells (PANC-1 and SW1990 with capsaicin resulted in cells growth inhibition, G0/G1 phase arrest, and apoptosis in a dose-dependent manner. Knockdown of growth arrest- and DNA damage-inducible gene 153 (GADD153, a marker of the endoplasmic-reticulum-stress- (ERS- mediated apoptosis pathway, by specific siRNA attenuated capsaicin-induced apoptosis both in PANC-1 and SW1990 cells. Moreover, in vivo studies capsaicin effectively inhibited the growth and metabolism of pancreatic cancer and prolonged the survival time of pancreatic cancer xenograft tumor-induced mice. Furthermore, capsaicin increased the expression of some key ERS markers, including glucose-regulated protein 78 (GRP78, phosphoprotein kinase-like endoplasmic reticulum kinase (phosphoPERK, and phosphoeukaryotic initiation factor-2α (phospho-eIF2α, activating transcription factor 4 (ATF4 and GADD153 in tumor tissues. In conclusion, we for the first time provide important evidence to support the involvement of ERS in the induction of apoptosis in pancreatic cancer cells by capsaicin.

  13. Hesperidin inhibits HeLa cell proliferation through apoptosis mediated by endoplasmic reticulum stress pathways and cell cycle arrest

    International Nuclear Information System (INIS)

    Wang, Yaoxian; Yu, Hui; Zhang, Jin; Gao, Jing; Ge, Xin; Lou, Ge

    2015-01-01

    Hesperidin (30, 5, 9-dihydroxy-40-methoxy-7-orutinosyl flavanone) is a flavanone that is found mainly in citrus fruits and has been shown to have some anti-neoplastic effects. The aim of the present study was to investigate the effect of hesperidin on apoptosis in human cervical cancer HeLa cells and to identify the mechanism involved. Cells were treated with hesperidin (0, 20, 40, 60, 80, and 100 μM) for 24, 48, or 72 h and relative cell viability was assessed using the 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. Hesperidin inhibited the proliferation of HeLa cells in a concentration- and time-dependent manner. Hesperidin-induced apoptosis in HeLa cells was characterized by increased nuclear condensation and DNA fragmentation. Furthermore, increased levels of GADD153/CHOP and GRP78 indicated hesperidin-induced apoptosis in HeLa cells involved a caspase-dependent pathway, presumably downstream of the endoplasmic reticulum stress pathway. Both of these proteins are hallmarks of endoplasmic reticulum stress. Hesperidin also promoted the formation of reactive oxygen species, mobilization of intracellular Ca 2+ , loss of mitochondrial membrane potential (ΔΨm), increased release of cytochrome c and apoptosis-inducing factor from mitochondria, and promoted capase-3 activation. It also arrested HeLa cells in the G0/G1 phase in the cell cycle by downregulating the expression of cyclinD1, cyclinE1, and cyclin-dependent kinase 2 at the protein level. The effect of hesperidin was also verified on the human colon cancer cell HT-29 cells. We concluded that hesperidin inhibited HeLa cell proliferation through apoptosis involving endoplasmic reticulum stress pathways and cell cycle arrest

  14. High Glucose Inhibits Neural Stem Cell Differentiation Through Oxidative Stress and Endoplasmic Reticulum Stress.

    Science.gov (United States)

    Chen, Xi; Shen, Wei-Bin; Yang, Penghua; Dong, Daoyin; Sun, Winny; Yang, Peixin

    2018-06-01

    Maternal diabetes induces neural tube defects by suppressing neurogenesis in the developing neuroepithelium. Our recent study further revealed that high glucose inhibited embryonic stem cell differentiation into neural lineage cells. However, the mechanism whereby high glucose suppresses neural differentiation is unclear. To investigate whether high glucose-induced oxidative stress and endoplasmic reticulum (ER) stress lead to the inhibition of neural differentiation, the effect of high glucose on neural stem cell (the C17.2 cell line) differentiation was examined. Neural stem cells were cultured in normal glucose (5 mM) or high glucose (25 mM) differentiation medium for 3, 5, and 7 days. High glucose suppressed neural stem cell differentiation by significantly decreasing the expression of the neuron marker Tuj1 and the glial cell marker GFAP and the numbers of Tuj1 + and GFAP + cells. The antioxidant enzyme superoxide dismutase mimetic Tempol reversed high glucose-decreased Tuj1 and GFAP expression and restored the numbers of neurons and glial cells differentiated from neural stem cells. Hydrogen peroxide treatment imitated the inhibitory effect of high glucose on neural stem cell differentiation. Both high glucose and hydrogen peroxide triggered ER stress, whereas Tempol blocked high glucose-induced ER stress. The ER stress inhibitor, 4-phenylbutyrate, abolished the inhibition of high glucose or hydrogen peroxide on neural stem cell differentiation. Thus, oxidative stress and its resultant ER stress mediate the inhibitory effect of high glucose on neural stem cell differentiation.

  15. Germ Cell-less Promotes Centrosome Segregation to Induce Germ Cell Formation

    Directory of Open Access Journals (Sweden)

    Dorothy A. Lerit

    2017-01-01

    Full Text Available The primordial germ cells (PGCs specified during embryogenesis serve as progenitors to the adult germline stem cells. In Drosophila, the proper specification and formation of PGCs require both centrosomes and germ plasm, which contains the germline determinants. Centrosomes are microtubule (MT-organizing centers that ensure the faithful segregation of germ plasm into PGCs. To date, mechanisms that modulate centrosome behavior to engineer PGC development have remained elusive. Only one germ plasm component, Germ cell-less (Gcl, is known to play a role in PGC formation. Here, we show that Gcl engineers PGC formation by regulating centrosome dynamics. Loss of gcl leads to aberrant centrosome separation and elaboration of the astral MT network, resulting in inefficient germ plasm segregation and aborted PGC cellularization. Importantly, compromising centrosome separation alone is sufficient to mimic the gcl loss-of-function phenotypes. We conclude Gcl functions as a key regulator of centrosome separation required for proper PGC development.

  16. Cytoskeletal-assisted dynamics of the mitochondrial reticulum in living cells.

    Science.gov (United States)

    Knowles, Michelle K; Guenza, Marina G; Capaldi, Roderick A; Marcus, Andrew H

    2002-11-12

    Subcellular organelle dynamics are strongly influenced by interactions with cytoskeletal filaments and their associated motor proteins, and lead to complex multiexponential relaxations that occur over a wide range of spatial and temporal scales. Here we report spatio-temporal measurements of the fluctuations of the mitochondrial reticulum in osteosarcoma cells by using Fourier imaging correlation spectroscopy, over time and distance scales of 10(-2) to 10(3) s and 0.5-2.5 microm. We show that the method allows a more complete description of mitochondrial dynamics, through the time- and length-scale-dependent collective diffusion coefficient D(k,tau), than available by other means. Addition of either nocodazole to disrupt microtubules or cytochalasin D to disassemble microfilaments simplifies the intermediate scattering function. When both drugs are used, the reticulum morphology of mitochondria is retained even though the cytoskeletal elements have been de-polymerized. The dynamics of the organelle are then primarily diffusive and can be modeled as a collection of friction points interconnected by elastic springs. This study quantitatively characterizes organelle dynamics in terms of collective cytoskeletal interactions in living cells.

  17. The glutathione mimic ebselen inhibits oxidative stress but not endoplasmic reticulum stress in endothelial cells.

    Science.gov (United States)

    Ahwach, Salma Makhoul; Thomas, Melanie; Onstead-Haas, Luisa; Mooradian, Arshag D; Haas, Michael J

    2015-08-01

    Reactive oxygen species are associated with cardiovascular disease, diabetes, and atherosclerosis, yet the use of antioxidants in clinical trials has been ineffective at improving outcomes. In endothelial cells, high-dextrose-induced oxidative stress and endoplasmic reticulum stress promote endothelial dysfunction leading to the recruitment and activation of peripheral blood lymphocytes and the breakdown of barrier function. Ebselen, a glutathione peroxidase 1 (GPX1) mimic, has been shown to improve β-cell function in diabetes and prevent atherosclerosis. To determine if ebselen inhibits both oxidative stress and endoplasmic reticulum (ER) stress in endothelial cells, we examined its effects in human umbilical vein endothelial cells (HUVEC) and human coronary artery endothelial cells (HCAEC) with and without high-dextrose. Oxidative stress and ER stress were measured by 2-methyl-6-(4-methoxyphenyl)-3,7-dihydroimidazo[1,2-A]pyrazin-3-one hydrochloride chemiluminescence and ER stress alkaline phosphatase assays, respectively. GPX1 over-expression and knockdown were performed by transfecting cells with a GPX1 expression construct or a GPX1-specific siRNA, respectively. Ebselen inhibited dextrose-induced oxidative stress but not ER stress in both HUVEC and HCAEC. Ebselen also had no effect on tunicamycin-induced ER stress in HCAEC. Furthermore, augmentation of GPX1 activity directly by sodium selenite supplementation or transfection of a GPX1 expression plasmid decreased dextrose-induced oxidative stress but not ER stress, while GPX1 knockout enhanced oxidative stress but had no effect on ER stress. These results suggest that ebselen targets only oxidative stress but not ER stress. Copyright © 2015. Published by Elsevier Inc.

  18. The orphan nuclear receptor NR4A1 (Nur77) regulates oxidative and endoplasmic reticulum stress in pancreatic cancer cells.

    Science.gov (United States)

    Lee, Syng-Ook; Jin, Un-Ho; Kang, Jeong Han; Kim, Sang Bae; Guthrie, Aaron S; Sreevalsan, Sandeep; Lee, Ju-Seog; Safe, Stephen

    2014-04-01

    NR4A1 (Nur77, TR3) is an orphan nuclear receptor that is overexpressed in pancreatic cancer and exhibits pro-oncogenic activity. RNA interference of NR4A1 expression in Panc-1 cells induced apoptosis and subsequent proteomic analysis revealed the induction of several markers of endoplasmic reticulum stress, including glucose-related protein 78 (GRP78), CCAAT/enhancer-binding protein-homologous protein (CHOP), and activating transcription factor-4 (ATF-4). Treatment of pancreatic cancer cells with the NR4A1 antagonist 1,1-bis(3'-indolyl)-1-(p-hydroxyphenyl)methane (DIM-C-pPhOH) gave similar results. Moreover, both NR4A1 knockdown and DIM-C-pPhOH induced reactive oxygen species (ROS), and induction of ROS and endoplasmic reticulum stress by these agents was attenuated after cotreatment with antioxidants. Manipulation of NR4A1 expression coupled with gene expression profiling identified a number of ROS metabolism transcripts regulated by NR4A1. Knockdown of one of these transcripts, thioredoxin domain containing 5 (TXNDC5), recapitulated the elevated ROS and endoplasmic reticulum stress; thus, demonstrating that NR4A1 regulates levels of endoplasmic reticulum stress and ROS in pancreatic cancer cells to facilitate cell proliferation and survival. Finally, inactivation of NR4A1 by knockdown or DIM-C-pPhOH decreased TXNDC5, resulting in activation of the ROS/endoplasmic reticulum stress and proapoptotic pathways. The NR4A1 receptor is pro-oncogenic, regulates the ROS/endoplasmic reticulum stress pathways, and inactivation of the receptor represents a novel pathway for inducing cell death in pancreatic cancer. Mol Cancer Res; 12(4); 527-38. ©2014 AACR.

  19. Template DNA-strand co-segregation and asymmetric cell division in skeletal muscle stem cells.

    Science.gov (United States)

    Shinin, Vasily; Gayraud-Morel, Barbara; Tajbakhsh, Shahragim

    2009-01-01

    Stem cells are present in all tissues and organs, and are crucial for normal regulated growth. How the pool size of stem cells and their progeny is regulated to establish the tissue prenatally, then maintain it throughout life, is a key question in biology and medicine. The ability to precisely locate stem and progenitors requires defining lineage progression from stem to differentiated cells, assessing the mode of cell expansion and self-renewal and identifying markers to assess the different cell states within the lineage. We have shown that during lineage progression from a quiescent adult muscle satellite cell to a differentiated myofibre, both symmetric and asymmetric divisions take place. Furthermore, we provide evidence that a sub-population of label retaining satellite cells co-segregate template DNA strands to one daughter cell. These findings provide a means of identifying presumed stem and progenitor cells within the lineage. In addition, asymmetric segregation of template DNA and the cytoplasmic protein Numb provides a landmark to define cell behaviour as self-renewal and differentiation decisions are being executed.

  20. 3-Bromopyruvate induces endoplasmic reticulum stress, overcomes autophagy and causes apoptosis in human HCC cell lines.

    Science.gov (United States)

    Ganapathy-Kanniappan, Shanmugasundaram; Geschwind, Jean-Francois H; Kunjithapatham, Rani; Buijs, Manon; Syed, Labiq H; Rao, Pramod P; Ota, Shinichi; Kwak, Byung Kook; Loffroy, Romaric; Vali, Mustafa

    2010-03-01

    Autophagy, a cellular response to stress, plays a role in resistance to chemotherapy in cancer cells. Resistance renders systemic chemotherapy generally ineffective against human hepatocellular carcinoma (HCC). Recently, we reported that the pyruvate analog 3-bromopyruvate (3-BrPA) promoted tumor cell death by targeting GAPDH. In continuance, we investigated the intracellular response of two human HCC cell lines (Hep3B and SK-Hep1) that differ in their status of key apoptotic regulators, p53 and Fas. 3-BrPA treatment induced endoplasmic reticulum (ER) stress, translation inhibition and apoptosis based on Western blot and qPCR, pulse labeling, Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay and active caspase-3 in both the cell lines. However, electron microscopy revealed that 3-BrPA treated SK-Hep1 cells underwent classical apoptotic cell death while Hep3B cells initially responded with the protective autophagy that failed to prevent eventual apoptosis. 3-BrPA treatment promotes apoptosis in human HCC cell lines, irrespective of the intracellular response.

  1. Naphthoquinone Derivative PPE8 Induces Endoplasmic Reticulum Stress in p53 Null H1299 Cells

    Directory of Open Access Journals (Sweden)

    Jin-Cherng Lien

    2015-01-01

    Full Text Available Endoplasmic reticulum (ER plays a key role in synthesizing secretory proteins and sensing signal function in eukaryotic cells. Responding to calcium disturbance, oxidation state change, or pharmacological agents, ER transmembrane protein, inositol-regulating enzyme 1 (IRE1, senses the stress and triggers downstream signals. Glucose-regulated protein 78 (GRP78 dissociates from IRE1 to assist protein folding and guard against cell death. In prolonged ER stress, IRE1 recruits and activates apoptosis signal-regulating kinase 1 (ASK1 as well as downstream JNK for cell death. Naphthoquinones are widespread natural phenolic compounds. Vitamin K3, a derivative of naphthoquinone, inhibits variant tumor cell growth via oxygen uptake and oxygen stress. We synthesized a novel naphthoquinone derivative PPE8 and evaluated capacity to induce ER stress in p53 null H1299 and p53 wild-type A549 cells. In H1299 cells, PPE8 induced ER enlargement, GRP78 expression, and transient IER1 activation. Activated IRE1 recruited ASK1 for downstream JNK phosphorylation. IRE1 knockdown by siRNA attenuated PPE8-induced JNK phosphorylation and cytotoxicity. Prolonged JNK phosphorylation may be involved in PPE8-induced cytotoxicity. Such results did not arise in A549 cells, but p53 knockdown by siRNA restored PPE8-induced GRP78 expression and JNK phosphorylation. We offer a novel compound to induce ER stress and cytotoxicity in p53-deficient cancer cells, presenting an opportunity for treatment.

  2. Interdependence of bacterial cell division and genome segregation and its potential in drug development.

    Science.gov (United States)

    Misra, Hari S; Maurya, Ganesh K; Chaudhary, Reema; Misra, Chitra S

    2018-03-01

    Cell division and genome segregation are mutually interdependent processes, which are tightly linked with bacterial multiplication. Mechanisms underlying cell division and the cellular machinery involved are largely conserved across bacteria. Segregation of genome elements on the other hand, follows different pathways depending upon its type and the functional components encoded on these elements. Small molecules, that are known to inhibit cell division and/or resolution of intertwined circular chromosome and maintenace of DNA topology have earlier been tested as antibacterial agents. The utility of such drugs in controlling bacterial infections has witnessed only partial success, possibly due to functional redundancy associated with targeted components. However, in due course, literature has grown with newer information. This review has brought forth some recent findings on bacterial cell division with special emphasis on crosstalk between cell division and genome segregation that could be explored as novel targets in drug development. Copyright © 2018 Elsevier GmbH. All rights reserved.

  3. Retention of CXCR4 in the endoplasmic reticulum blocks dissemination of a T cell hybridoma.

    Science.gov (United States)

    Zeelenberg, I S; Ruuls-Van Stalle, L; Roos, E

    2001-07-01

    The dissemination of T cell hybridomas to multiple nonhematopoietic tissues is blocked by pertussis toxin, suggesting the involvement of a chemokine. To study whether this chemokine is SDF-1, we employed a strategy proposed previously for gene therapy of AIDS, whereby the SDF-1 receptor CXCR4 (also a coreceptor for HIV) is retained in the endoplasmic reticulum (ER) and fails to reach the cell surface. We transfected SDF-1, carrying an ER retention sequence, into a T cell hybridoma. This altered chemokine is retained in the ER, where it binds CXCR4 and prevents the latter protein from reaching the surface. These cells failed to migrate toward SDF-1 or to invade fibroblast monolayers, although they could still migrate toward thymus and activation-regulated chemokine (TARC) and invade TARC-treated monolayers. Furthermore, the ability of the transfected cells to disseminate to multiple organs upon intravenous injection into mice was abolished. This dissemination reflects the in vivo migration patterns of activated and memory T cells into nonhematopoietic tissues, which is thus likely to depend on CXCR4. Attempts to block CXCR4 function as a therapy for AIDS may affect this migration with consequences for T cell function. Our results also suggest a decisive role for CXCR4 in the dissemination of hematopoietic malignancies expressing this receptor.

  4. Protective effect of catechin in type I Gaucher disease cells by reducing endoplasmic reticulum stress

    International Nuclear Information System (INIS)

    Lee, Yea-Jin; Kim, Sung-Jo; Heo, Tae-Hwe

    2011-01-01

    Highlights: → Catechin reduces the expression level of ER stress marker protein in type I Gaucher disease cells. → Catechin induces the proliferation rate of GD cells similar levels to normal cells. → Catechin improves wound healing activity. → Catechin-mediated reductions in ER stress may be associated with enhanced cell survival. → We identified catechin as a protective agent against ER stress in GD cells. -- Abstract: Gaucher disease (GD) is the most common lysosomal storage disorder (LSD) and is divided into three phenotypes, I, II, and III. Type I is the most prevalent form and has its onset in adulthood. The degree of endoplasmic reticulum (ER) stress is one of the factors that determine GD severity. It has recently been reported that antioxidants reduce ER stress and apoptosis by scavenging the oxidants that cause oxidative stress. For this report, we investigated the possibility that catechin can act on type I GD patient cells to alleviate the pathogenic conditions of GD. We treated GD cells with catechin and examined the expression level of GRP78/BiP (an ER stress marker) by western blots and fluorescence microscopy, the proliferation rate of GD cells, and scratch-induced wound healing activity. Our results show that catechin reduces the expression level of GRP78/BiP, leads to cell proliferation rates of GD cells similar levels to normal cells, and improves wound healing activity. We conclude that catechin protects against ER stress in GD cells and catechin-mediated reductions in ER stress may be associated with enhanced cell survival.

  5. Protective effect of catechin in type I Gaucher disease cells by reducing endoplasmic reticulum stress

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Yea-Jin [Department of Biotechnology, Hoseo University, Baebang, Asan, Chungnam, 336-795 (Korea, Republic of); Kim, Sung-Jo, E-mail: sungjo@hoseo.edu [Department of Biotechnology, Hoseo University, Baebang, Asan, Chungnam, 336-795 (Korea, Republic of); Heo, Tae-Hwe, E-mail: thhur92@catholic.ac.kr [College of Pharmacy, The Catholic University of Korea, Bucheon 420-743 (Korea, Republic of)

    2011-09-23

    Highlights: {yields} Catechin reduces the expression level of ER stress marker protein in type I Gaucher disease cells. {yields} Catechin induces the proliferation rate of GD cells similar levels to normal cells. {yields} Catechin improves wound healing activity. {yields} Catechin-mediated reductions in ER stress may be associated with enhanced cell survival. {yields} We identified catechin as a protective agent against ER stress in GD cells. -- Abstract: Gaucher disease (GD) is the most common lysosomal storage disorder (LSD) and is divided into three phenotypes, I, II, and III. Type I is the most prevalent form and has its onset in adulthood. The degree of endoplasmic reticulum (ER) stress is one of the factors that determine GD severity. It has recently been reported that antioxidants reduce ER stress and apoptosis by scavenging the oxidants that cause oxidative stress. For this report, we investigated the possibility that catechin can act on type I GD patient cells to alleviate the pathogenic conditions of GD. We treated GD cells with catechin and examined the expression level of GRP78/BiP (an ER stress marker) by western blots and fluorescence microscopy, the proliferation rate of GD cells, and scratch-induced wound healing activity. Our results show that catechin reduces the expression level of GRP78/BiP, leads to cell proliferation rates of GD cells similar levels to normal cells, and improves wound healing activity. We conclude that catechin protects against ER stress in GD cells and catechin-mediated reductions in ER stress may be associated with enhanced cell survival.

  6. Effect of endoplasmic reticulum stress on the response of HeLa cells to carbon ion radiation

    International Nuclear Information System (INIS)

    Xia Jiefang; Wang Zhuanzi; Wei Wei; Dang Bingrong; Li Wenjian

    2015-01-01

    To investigate the effect of endoplasmic reticulum stress on HeLa cells to "1"2C"6"+ ion irradiation, HeLa cells were pretreated with 2.5 mmol/L dithiothreitol and irradiated by "1"2C"6"+ ions with different doses. The results showed that, compared with IR alone, dithiothreitol combined with carbon ion irradiation caused HeLa cell survival decreased, and the apoptosis increased. Moreover, dithiothreitol and carbon ion radiation combination treatment led to a significant increase of G_2/M phase, and autophagy was activated obviously in combination treatment group. The results imply that continuous endoplasmic reticulum stress can change the response of HeLa cells to "1"2C"6"+ irradiation, and dithiothreitol may affect HeLa cells through the autophagy cell death pathway. (authors)

  7. Intestinal Epithelial Cell Endoplasmic Reticulum Stress and Inflammatory Bowel Disease Pathogenesis: An Update Review

    Directory of Open Access Journals (Sweden)

    Xiaoshi Ma

    2017-10-01

    Full Text Available The intestinal epithelial cells serve essential roles in maintaining intestinal homeostasis, which relies on appropriate endoplasmic reticulum (ER function for proper protein folding, modification, and secretion. Exogenous or endogenous risk factors with an ability to disturb the ER function can impair the intestinal barrier function and activate inflammatory responses in the host. The last decade has witnessed considerable progress in the understanding of the functional role of ER stress and unfolded protein response (UPR in the gut homeostasis and its significant contribution to the pathogenesis of inflammatory bowel disease (IBD. Herein, we review recent evidence supporting the viewpoint that deregulation of ER stress and UPR signaling in the intestinal epithelium, including the absorptive cells, Paneth cells, goblet cells, and enteroendocrine cells, mediates the action of genetic or environmental factors driving colitis in experimental animals and IBD patients. In addition, we highlight pharmacologic application of chaperones or small molecules that enhance protein folding and modification capacity or improve the function of the ER. These molecules represent potential therapeutic strategies in the prevention or treatment of IBD through restoring ER homeostasis in intestinal epithelial cells.

  8. Melatonin Modulates Neuronal Cell Death Induced by Endoplasmic Reticulum Stress under Insulin Resistance Condition.

    Science.gov (United States)

    Song, Juhyun; Kim, Oh Yoen

    2017-06-10

    Insulin resistance (IR) is an important stress factor in the central nervous system, thereby aggravating neuropathogenesis and triggering cognitive decline. Melatonin, which is an antioxidant phytochemical and synthesized by the pineal gland, has multiple functions in cellular responses such as apoptosis and survival against stress. This study investigated whether melatonin modulates the signaling of neuronal cell death induced by endoplasmic reticulum (ER) stress under IR condition using SH-SY5Y neuroblastoma cells. Apoptosis cell death signaling markers (cleaved Poly [ADP-ribose] polymerase 1 (PARP), p53, and Bax) and ER stress markers (phosphorylated eIF2α (p-eIF2α), ATF4, CHOP, p-IRE1 , and spliced XBP1 (sXBP1)) were measured using reverse transcription-PCR, quantitative PCR, and western blottings. Immunofluorescence staining was also performed for p-ASK1 and p-IRE1 . The mRNA or protein expressions of cell death signaling markers and ER stress markers were increased under IR condition, but significantly attenuated by melatonin treatment. Insulin-induced activation of ASK1 ( p-ASK1 ) was also dose dependently attenuated by melatonin treatment. The regulatory effect of melatonin on neuronal cells under IR condition was associated with ASK1 signaling. In conclusion, the result suggested that melatonin may alleviate ER stress under IR condition, thereby regulating neuronal cell death signaling.

  9. Baicalein Induces Apoptosis and Autophagy via Endoplasmic Reticulum Stress in Hepatocellular Carcinoma Cells

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    Zhongxia Wang

    2014-01-01

    Full Text Available Background. Hepatocellular carcinoma (HCC remains a disastrous disease and the treatment for HCC is rather limited. Separation and identification of active compounds from traditionally used herbs in HCC treatment may shed light on novel therapeutic drugs for HCC. Methods. Cell viability and colony forming assay were conducted to determine anti-HCC activity. Morphology of cells and activity of caspases were analyzed. Antiapoptotic Bcl-2 family proteins and JNK were also examined. Levels of unfolded protein response (UPR markers were determined and intracellular calcium was assayed. Small interfering RNAs (siRNAs were used to investigate the role of UPR and autophagy in baicalein-induced cell death. Results. Among four studied flavonoids, only baicalein exhibited satisfactory inhibition of viability and colony formation of HCC cells within water-soluble concentration. Baicalein induced apoptosis via endoplasmic reticulum (ER stress, possibly by downregulating prosurvival Bcl-2 family, increasing intracellular calcium, and activating JNK. CHOP was the executor of cell death during baicalein-induced ER stress while eIF2α and IRE1α played protective roles. Protective autophagy was also triggered by baicalein in HCC cells. Conclusion. Baicalein exhibits prominent anti-HCC activity. This flavonoid induces apoptosis and protective autophagy via ER stress. Combination of baicalein and autophagy inhibitors may represent a promising therapy against HCC.

  10. Arsenic induces cell apoptosis in cultured osteoblasts through endoplasmic reticulum stress

    International Nuclear Information System (INIS)

    Tang, C.-H.; Chiu, Y.-C.; Huang, C.-F.; Chen, Y.-W.; Chen, P.-C.

    2009-01-01

    Osteoporosis is characterized by low bone mass resulting from an imbalance between bone resorption by osteoclasts and bone formation by osteoblasts. Therefore, decreased bone formation by osteoblasts may lead to the development of osteoporosis, and rate of apoptosis is responsible for the regulation of bone formation. Arsenic (As) exists ubiquitously in our environment and increases the risk of neurotoxicity, liver injury, peripheral vascular disease and cancer. However, the effect of As on apoptosis of osteoblasts is mostly unknown. Here, we found that As induced cell apoptosis in osteoblastic cell lines (including hFOB, MC3T3-E1 and MG-63) and mouse bone marrow stromal cells (M2-10B4). As also induced upregulation of Bax and Bak, downregulation of Bcl-2 and dysfunction of mitochondria in osteoblasts. As also triggered endoplasmic reticulum (ER) stress, as indicated by changes in cytosolic-calcium levels. We found that As increased the expression and activities of glucose-regulated protein 78 (GRP78) and calpain. Transfection of cells with GRP78 or calpain siRNA reduced As-mediated cell apoptosis in osteoblasts. Therefore, our results suggest that As increased cell apoptosis in cultured osteoblasts and increased the risk of osteoporosis.

  11. Selective modulation of endoplasmic reticulum stress markers in prostate cancer cells by a standardized mangosteen fruit extract.

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    Gongbo Li

    Full Text Available The increased proliferation of cancer cells is directly dependent on the increased activity of the endoplasmic reticulum (ER machinery which is responsible for protein folding, assembly, and transport. In fact, it is so critical that perturbations in the endoplasmic reticulum can lead to apoptosis. This carefully regulated organelle represents a unique target of cancer cells while sparing healthy cells. In this study, a standardized mangosteen fruit extract (MFE was evaluated for modulating ER stress proteins in prostate cancer. Two human prostate cancer cell lines, 22Rv1 and LNCaP, and prostate epithelial cells (PrECs procured from two patients undergoing radical prostatectomy were treated with MFE. Flow cytometry, MTT, BrdU and Western blot were used to evaluate cell apoptosis, viability, proliferation and ER stress. Next, we evaluated MFE for microsomal stability and anti-cancer activity in nude mice. MFE induced apoptosis, decreased viability and proliferation in prostate cancer cells. MFE increased the expression of ER stress proteins. Interestingly, MFE selectively promotes ER stress in prostate cancer cells while sparing PrECs. MFE suppressed tumor growth in a xenograft tumor model without obvious toxicity. Mangosteen fruit extract selectively promotes endoplasmic reticulum stress in cancer cells while sparing non-tumorigenic prostate epithelial cells. Furthermore, in an in vivo setting mangosteen fruit extract significantly reduces xenograft tumor formation.

  12. Selective modulation of endoplasmic reticulum stress markers in prostate cancer cells by a standardized mangosteen fruit extract.

    Science.gov (United States)

    Li, Gongbo; Petiwala, Sakina M; Pierce, Dana R; Nonn, Larisa; Johnson, Jeremy J

    2013-01-01

    The increased proliferation of cancer cells is directly dependent on the increased activity of the endoplasmic reticulum (ER) machinery which is responsible for protein folding, assembly, and transport. In fact, it is so critical that perturbations in the endoplasmic reticulum can lead to apoptosis. This carefully regulated organelle represents a unique target of cancer cells while sparing healthy cells. In this study, a standardized mangosteen fruit extract (MFE) was evaluated for modulating ER stress proteins in prostate cancer. Two human prostate cancer cell lines, 22Rv1 and LNCaP, and prostate epithelial cells (PrECs) procured from two patients undergoing radical prostatectomy were treated with MFE. Flow cytometry, MTT, BrdU and Western blot were used to evaluate cell apoptosis, viability, proliferation and ER stress. Next, we evaluated MFE for microsomal stability and anti-cancer activity in nude mice. MFE induced apoptosis, decreased viability and proliferation in prostate cancer cells. MFE increased the expression of ER stress proteins. Interestingly, MFE selectively promotes ER stress in prostate cancer cells while sparing PrECs. MFE suppressed tumor growth in a xenograft tumor model without obvious toxicity. Mangosteen fruit extract selectively promotes endoplasmic reticulum stress in cancer cells while sparing non-tumorigenic prostate epithelial cells. Furthermore, in an in vivo setting mangosteen fruit extract significantly reduces xenograft tumor formation.

  13. Collective motion of cells mediates segregation and pattern formation in co-cultures.

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    Elod Méhes

    Full Text Available Pattern formation by segregation of cell types is an important process during embryonic development. We show that an experimentally yet unexplored mechanism based on collective motility of segregating cells enhances the effects of known pattern formation mechanisms such as differential adhesion, mechanochemical interactions or cell migration directed by morphogens. To study in vitro cell segregation we use time-lapse videomicroscopy and quantitative analysis of the main features of the motion of individual cells or groups. Our observations have been extensive, typically involving the investigation of the development of patterns containing up to 200,000 cells. By either comparing keratocyte types with different collective motility characteristics or increasing cells' directional persistence by the inhibition of Rac1 GTP-ase we demonstrate that enhanced collective cell motility results in faster cell segregation leading to the formation of more extensive patterns. The growth of the characteristic scale of patterns generally follows an algebraic scaling law with exponent values up to 0.74 in the presence of collective motion, compared to significantly smaller exponents in case of diffusive motion.

  14. Valsartan protects HK-2 cells from contrast media-induced apoptosis by inhibiting endoplasmic reticulum stress.

    Science.gov (United States)

    Peng, Ping-An; Wang, Le; Ma, Qian; Xin, Yi; Zhang, Ou; Han, Hong-Ya; Liu, Xiao-Li; Ji, Qing-Wei; Zhou, Yu-Jie; Zhao, Ying-Xin

    2015-12-01

    Contrast-induced acute kidney injury (CI-AKI) is associated with increasing in-hospital and long-term adverse clinical outcomes in high-risk patients undergoing percutaneous coronary intervention (PCI). Contrast media (CM)-induced renal tubular cell apoptosis is reported to participate in this process by activating endoplasmic reticulum (ER) stress. An angiotensin II type 1 receptor (AT1R) antagonist can alleviate ER stress-induced renal apoptosis in streptozotocin (STZ)-induced diabetic mice and can reduce CM-induced renal apoptosis by reducing oxidative stress and reversing the enhancement of bax mRNA and the reduction of bcl-2 mRNA, but the effect of the AT1R blocker on ER stress in the pathogenesis of CI-AKI is still unknown. In this study, we explored the effect of valsartan on meglumine diatrizoate-induced human renal tubular cell apoptosis by measuring changes in ER stress-related biomarkers. The results showed that meglumine diatrizoate caused significant cell apoptosis by up-regulating the expression of ER stress markers, including glucose-regulated protein 78 (GRP78), activating transcription factor 4 (ATF4), CCAAT/enhancer-binding protein-homologous protein (CHOP) and caspase 12, in a time- and dose-dependent manner, which could be alleviated by preincubation with valsartan. In conclusion, valsartan had a potential nephroprotective effect on meglumine diatrizoate-induced renal cell apoptosis by inhibiting ER stress. © 2015 International Federation for Cell Biology.

  15. Fluorescence Dynamics in the Endoplasmic Reticulum of a Live Cell: Time-Resolved Confocal Microscopy.

    Science.gov (United States)

    Ghosh, Shirsendu; Nandi, Somen; Ghosh, Catherine; Bhattacharyya, Kankan

    2016-09-19

    Fluorescence dynamics in the endoplasmic reticulum (ER) of a live non-cancer lung cell (WI38) and a lung cancer cell (A549) are studied by using time-resolved confocal microscopy. To selectively study the organelle, ER, we have used an ER-Tracker dye. From the emission maximum (λmaxem) of the ER-Tracker dye, polarity (i.e. dielectric constant, ϵ) in the ER region of the cells (≈500 nm in WI38 and ≈510 nm in A549) is estimated to be similar to that of chloroform (λmaxem =506 nm, ϵ≈5). The red shift by 10 nm in λmaxem in the cancer cell (A549) suggests a slightly higher polarity compared to the non-cancer cell (WI38). The fluorescence intensity of the ER-Tracker dye exhibits prolonged intermittent oscillations on a timescale of 2-6 seconds for the cancer cell (A549). For the non-cancer cell (WI38), such fluorescence oscillations are much less prominent. The marked fluorescence intensity oscillations in the cancer cell are attributed to enhanced calcium oscillations. The average solvent relaxation time () of the ER region in the lung cancer cell (A549, 250±50 ps) is about four times faster than that in the non-cancer cell (WI38, 1000±50 ps). © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  16. Colonic stem cell data are consistent with the immortal model of stem cell division under non-random strand segregation.

    Science.gov (United States)

    Walters, K

    2009-06-01

    Colonic stem cells are thought to reside towards the base of crypts of the colon, but their numbers and proliferation mechanisms are not well characterized. A defining property of stem cells is that they are able to divide asymmetrically, but it is not known whether they always divide asymmetrically (immortal model) or whether there are occasional symmetrical divisions (stochastic model). By measuring diversity of methylation patterns in colon crypt samples, a recent study found evidence in favour of the stochastic model, assuming random segregation of stem cell DNA strands during cell division. Here, the effect of preferential segregation of the template strand is considered to be consistent with the 'immortal strand hypothesis', and explore the effect on conclusions of previously published results. For a sample of crypts, it is shown how, under the immortal model, to calculate mean and variance of the number of unique methylation patterns allowing for non-random strand segregation and compare them with those observed. The calculated mean and variance are consistent with an immortal model that incorporates non-random strand segregation for a range of stem cell numbers and levels of preferential strand segregation. Allowing for preferential strand segregation considerably alters previously published conclusions relating to stem cell numbers and turnover mechanisms. Evidence in favour of the stochastic model may not be as strong as previously thought.

  17. Endoplasmic Reticulum Stress Cooperates in Zearalenone-Induced Cell Death of RAW 264.7 Macrophages

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    Fenglei Chen

    2015-08-01

    Full Text Available Zearalenone (ZEA is a fungal mycotoxin that causes cell apoptosis and necrosis. However, little is known about the molecular mechanisms of ZEA toxicity. The objective of this study was to explore the effects of ZEA on the proliferation and apoptosis of RAW 264.7 macrophages and to uncover the signaling pathway underlying the cytotoxicity of ZEA in RAW 264.7 macrophages. This study demonstrates that the endoplasmic reticulum (ER stress pathway cooperated in ZEA-induced cell death of the RAW 264.7 macrophages. Our results show that ZEA treatment reduced the viability of RAW 264.7 macrophages in a dose- and time-dependent manner as shown by the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide assay (MTT and flow cytometry assay. Western blots analysis revealed that ZEA increased the expression of glucose-regulated protein 78 (GRP78 and CCAAT/enhancer binding protein homologous protein (CHOP, two ER stress-related marker genes. Furthermore, treating the cells with the ER stress inhibitors 4-phenylbutyrate (4-PBA or knocking down CHOP, using lentivirus encoded short hairpin interfering RNAs (shRNAs, significantly diminished the ZEA-induced increases in GRP78 and CHOP, and cell death. In summary, our results suggest that ZEA induces the apoptosis and necrosis of RAW 264.7 macrophages in a dose- and time-dependent manner via the ER stress pathway in which the activation of CHOP plays a critical role.

  18. Protective effect of mild endoplasmic reticulum stress on radiation-induced bystander effects in hepatocyte cells

    Science.gov (United States)

    Xie, Yuexia; Ye, Shuang; Zhang, Jianghong; He, Mingyuan; Dong, Chen; Tu, Wenzhi; Liu, Peifeng; Shao, Chunlin

    2016-01-01

    Radiation-induced bystander effect (RIBE) has important implications for secondary cancer risk assessment during cancer radiotherapy, but the defense and self-protective mechanisms of bystander normal cells are still largely unclear. The present study found that micronuclei (MN) formation could be induced in the non-irradiated HL-7702 hepatocyte cells after being treated with the conditioned medium from irradiated hepatoma HepG2 cells under either normoxia or hypoxia, where the ratio of the yield of bystander MN induction to the yield of radiation-induced MN formation under hypoxia was much higher than that of normoxia. Nonetheless, thapsigargin induced endoplasmic reticulum (ER) stress and dramatically suppressed this bystander response manifested as the decrease of MN and apoptosis inductions. Meanwhile, the interference of BiP gene, a major ER chaperone, amplified the detrimental RIBE. More precisely, thapsigargin provoked ER sensor of PERK to initiate an instantaneous and moderate ER stress thus defensed the hazard form RIBE, while BiP depletion lead to persistently destroyed homeostasis of ER and exacerbated cell injury. These findings provide new insights that the mild ER stress through BiP-PERK-p-eIF2α signaling pathway has a profound role in protecting cellular damage from RIBE and hence may decrease the potential secondary cancer risk after cancer radiotherapy. PMID:27958308

  19. Glucosamine-induced endoplasmic reticulum stress affects GLUT4 expression via activating transcription factor 6 in rat and human skeletal muscle cells

    DEFF Research Database (Denmark)

    Raciti, G A; Iadicicco, C; Ulianich, L

    2010-01-01

    Glucosamine, generated during hyperglycaemia, causes insulin resistance in different cells. Here we sought to evaluate the possible role of endoplasmic reticulum (ER) stress in the induction of insulin resistance by glucosamine in skeletal muscle cells.......Glucosamine, generated during hyperglycaemia, causes insulin resistance in different cells. Here we sought to evaluate the possible role of endoplasmic reticulum (ER) stress in the induction of insulin resistance by glucosamine in skeletal muscle cells....

  20. Protein-accumulating cells and dilated cisternae of the endoplasmic reticulum in three glucosinolate-containing genera: Armoracia, Capparis, Drypetes.

    Science.gov (United States)

    Jørgensen, L B; Behnke, H D; Mabry, T J

    1977-01-01

    Three glucosinolate-containing species, Armoracia rusticana Gaertner, Meyer et Scherbius (Brassicaceae), Capparis cynophallophora L. (Capparaceae) and Drypetes roxburghii (Wall.) Hurusawa (Euphorbiaceae), are shown by both light and electron microscopy to contain protein-accumulating cells (PAC). The PAC of Armoracia and Copparis (former "myrosin cells") occur as idioblasts. The PAC of Drypetes are usual members among axial phloem parenchyma cells rather than idioblasts. In Drypetes the vacuoles of the PAC are shown ultrastructurally to contain finely fibrillar material and to originate from local dilatations of the endoplasmic reticulum. The vacuoles in PAC of Armoracia and Capparis seem to originate in the same way; but ultrastructurally, their content is finely granular. In addition, Armoracia and Capparis are shown by both light and electron microscopy to contain dilated cisternae (DC) of the endoplasmic reticulum in normal parenchyma cells, in accord with previous findings for several species within Brassicaceae. The relationship of PAC and DC to glucosinolates and the enzyme myrosinase is discussed.

  1. Protein disulfide isomerases in the endoplasmic reticulum promote anchorage-independent growth of breast cancer cells.

    Science.gov (United States)

    Wise, Randi; Duhachek-Muggy, Sara; Qi, Yue; Zolkiewski, Michal; Zolkiewska, Anna

    2016-06-01

    Metastatic breast cancer cells are exposed to stress of detachment from the extracellular matrix (ECM). Cultured breast cancer cells that survive this stress and are capable of anchorage-independent proliferation form mammospheres. The purpose of this study was to explore a link between mammosphere growth, ECM gene expression, and the protein quality control system in the endoplasmic reticulum (ER). We compared the mRNA and protein levels of ER folding factors in SUM159PT and MCF10DCIS.com breast cancer cells grown as mammospheres versus adherent conditions. Publicly available gene expression data for mammospheres formed by primary breast cancer cells and for circulating tumor cells (CTCs) were analyzed to assess the status of ECM/ER folding factor genes in clinically relevant samples. Knock-down of selected protein disulfide isomerase (PDI) family members was performed to examine their roles in SUM159PT mammosphere growth. We found that cells grown as mammospheres had elevated expression of ECM genes and ER folding quality control genes. CTC gene expression data for an index patient indicated that upregulation of ECM and ER folding factor genes occurred at the time of acquired therapy resistance and disease progression. Knock-down of PDI, ERp44, or ERp57, three members of the PDI family with elevated protein levels in mammospheres, in SUM159PT cells partially inhibited the mammosphere growth. Thus, breast cancer cell survival and growth under detachment conditions require enhanced assistance of the ER protein folding machinery. Targeting ER folding factors, in particular members of the PDI family, may improve the therapeutic outcomes in metastatic breast cancer.

  2. HMGB1 induces an inflammatory response in endothelial cells via the RAGE-dependent endoplasmic reticulum stress pathway

    International Nuclear Information System (INIS)

    Luo, Ying; Li, Shu-Jun; Yang, Jian; Qiu, Yuan-Zhen; Chen, Fang-Ping

    2013-01-01

    Highlights: •Mechanisms of inflammatory response induced by HMGB1 are incompletely understood. •We found that endoplasmic reticulum stress mediate the inflammatory response induced by HMGB1. •RAGE-mediated ERS pathways are involved in those processes. •We reported a new mechanism for HMGB1 induced inflammatory response. -- Abstract: The high mobility group 1B protein (HMGB1) mediates chronic inflammatory responses in endothelial cells, which play a critical role in atherosclerosis. However, the underlying mechanism is unknown. The goal of our study was to identify the effects of HMGB1 on the RAGE-induced inflammatory response in endothelial cells and test the possible involvement of the endoplasmic reticulum stress pathway. Our results showed that incubation of endothelial cells with HMGB1 (0.01–1 μg/ml) for 24 h induced a dose-dependent activation of endoplasmic reticulum stress transducers, as assessed by PERK and IRE1 protein expression. Moreover, HMGB1 also promoted nuclear translocation of ATF6. HMGB1-mediated ICAM-1 and P-selectin production was dramatically suppressed by PERK siRNA or IRE1 siRNA. However, non-targeting siRNA had no such effects. HMGB1-induced increases in ICAM-1 and P-selectin expression were also inhibited by a specific eIF2α inhibitor (salubrinal) and a specific JNK inhibitor (SP600125). Importantly, a blocking antibody specifically targeted against RAGE (anti-RAGE antibody) decreased ICAM-1, P-selectin and endoplasmic reticulum stress molecule (PERK, eIF2α, IRE1 and JNK) protein expression levels. Collectively, these novel findings suggest that HMGB1 promotes an inflammatory response by inducing the expression of ICAM-1 and P-selectin via RAGE-mediated stimulation of the endoplasmic reticulum stress pathway

  3. Kaempferol induces hepatocellular carcinoma cell death via endoplasmic reticulum stress-CHOP-autophagy signaling pathway.

    Science.gov (United States)

    Guo, Haiqing; Lin, Wei; Zhang, Xiangying; Zhang, Xiaohui; Hu, Zhongjie; Li, Liying; Duan, Zhongping; Zhang, Jing; Ren, Feng

    2017-10-10

    Kaempferol is a flavonoid compound that has gained widespread attention due to its antitumor functions. However, the underlying mechanisms are still not clear. The present study investigated the effect of kaempferol on hepatocellular carcinoma and its underlying mechanisms. Kaempferol induced autophagy in a concentration- and time-dependent manner in HepG2 or Huh7 cells, which was evidenced by the significant increase of autophagy-related genes. Inhibition of autophagy pathway, through 3-methyladenine or Atg7 siRNA, strongly diminished kaempferol-induced apoptosis. We further hypothesized that kaempferol can induce autophagy via endoplasmic reticulum (ER) stress pathway. Indeed, blocking ER stress by 4-phenyl butyric acid (4-PBA) or knockdown of CCAAT/enhancer-binding protein homologous protein (CHOP) with siRNA alleviated kaempferol-induced HepG2 or Huh7 cells autophagy; while transfection with plasmid overexpressing CHOP reversed the effect of 4-PBA on kaempferol-induced autophagy. Our results demonstrated that kaempferol induced hepatocarcinoma cell death via ER stress and CHOP-autophagy signaling pathway; kaempferol may be used as a potential chemopreventive agent for patients with hepatocellular carcinoma.

  4. 'Candidatus Liberibacter asiaticus' Accumulates inside Endoplasmic Reticulum Associated Vacuoles in the Gut Cells of Diaphorina citri.

    Science.gov (United States)

    Ghanim, Murad; Achor, Diann; Ghosh, Saptarshi; Kontsedalov, Svetlana; Lebedev, Galina; Levy, Amit

    2017-12-05

    Citrus greening disease known also as Huanglongbing (HLB) caused by the phloem-limited bacterium 'Candidatus Liberibacter asiaticus' (CLas) has resulted in tremendous losses and the death of millions of trees worldwide. CLas is transmitted by the Asian citrus psyllid Diaphorina citri. The closely-related bacteria 'Candidatus Liberibacter solanacearum' (CLso), associated with vegetative disorders in carrots, is transmitted by the carrot psyllid Bactericera trigonica. A promising approach to prevent the transmission of these pathogens is to interfere with the vector-pathogen interactions, but our understanding of these processes is limited. It was recently reported that CLas induced changes in the nuclear architecture, and activated programmed cell death, in D. citri midgut cells. Here, we used electron and fluorescent microscopy and show that CLas induces the formation of endoplasmic reticulum (ER)-associated bodies. The bacterium recruits those ER structures into Liberibacter containing vacuoles (LCVs), in which bacterial cells seem to propagate. ER- associated LCV formation was unique to CLas, as we could not detect these bodies in B. trigonica infected with CLso. ER recruitment is hypothesized to generate a safe replicative body to escape cellular immune responses in the insect gut. Understanding the molecular interactions that undelay these responses will open new opportunities for controlling CLas.

  5. Pterostilbene Inhibits the Growth of Human Esophageal Cancer Cells by Regulating Endoplasmic Reticulum Stress

    Directory of Open Access Journals (Sweden)

    Yingtong Feng

    2016-03-01

    Full Text Available Background/Aims: Pterostilbene (PTE, a natural dimethylated resveratrol analog from blueberries, is known to have diverse pharmacological activities, including anticancer properties. In this study, we investigated the anticancer activity of PTE against human esophageal cancer cells both in vitro and in vivo and explored the role of endoplasmic reticulum (ER stress (ERS signaling in this process. Methods: Cell viability, the apoptotic index, Caspase 3 activity, adhesion, migration, reactive oxygen species (ROS levels, and glutathione (GSH levels were detected to explore the effect of PTE on human EC109 esophageal cancer cells. Furthermore, siRNA transfection and a chemical inhibitor were employed to confirm the role of ERS. Results: PTE treatment dose- and time-dependently decreased the viability of human esophageal cancer EC109 cells. PTE also decreased tumor cell adhesion, migration and intracellular GSH levels while increasing the apoptotic index, Caspase 3 activity and ROS levels, which suggest the strong anticancer activity of PTE. Furthermore, PTE treatment increased the expression of ERS-related molecules (GRP78, ATF6, p-PERK, p-eIF2α and CHOP, upregulated the pro-apoptosis-related protein PUMA and downregulated the anti-apoptosis-related protein Bcl-2 while promoting the translocation of cytochrome c from mitochondria to cytosol and the activation of Caspase 9 and Caspase 12. The downregulation of ERS signaling by CHOP siRNA desensitized esophageal cancer cells to PTE treatment, whereas upregulation of ERS signaling by thapsigargin (THA had the opposite effect. N-Acetylcysteine (NAC, a ROS scavenger, also desensitized esophageal cancer cells to PTE treatment. Conclusions: Overall, the results indicate that PTE is a potent anti-cancer pharmaceutical against human esophageal cancer, and the possible mechanism involves the activation of ERS signaling pathways.

  6. Protective effects of 4-phenylbutyrate derivatives on the neuronal cell death and endoplasmic reticulum stress.

    Science.gov (United States)

    Mimori, Seisuke; Okuma, Yasunobu; Kaneko, Masayuki; Kawada, Koichi; Hosoi, Toru; Ozawa, Koichiro; Nomura, Yasuyuki; Hamana, Hiroshi

    2012-01-01

    Endoplasmic reticulum (ER) stress responses play an important role in neurodegenerative diseases. Sodium 4-phenylbutyrate (4-PBA) is a terminal aromatic substituted fatty acid that has been used for the treatment of urea cycle disorders. 4-PBA possesses in vitro chemical chaperone activity and reduces the accumulation of Parkin-associated endothelin receptor-like receptor (Pael-R), which is involved in autosomal recessive juvenile parkinsonism (AR-JP). In this study, we show that terminal aromatic substituted fatty acids, including 3-phenylpropionate (3-PPA), 4-PBA, 5-phenylvaleric acid, and 6-phenylhexanoic acid, prevented the aggregation of lactalbumin and bovine serum albumin. Aggregation inhibition increased relative to the number of carbons in the fatty acids. Moreover, these compounds protected cells against ER stress-induced neuronal cell death. The cytoprotective effect correlated with the in vitro chemical chaperone activity. Similarly, cell viability decreased on treatment with tunicamycin, an ER stress inducer, and was dependent on the number of carbons in the fatty acids. Moreover, the expression of glucose-regulated proteins 94 and 78 (GRP94, 78) decreased according to the number of carbons in the fatty acids. Furthermore, we investigated the effects of these compounds on the accumulation of Pael-R in neuroblastoma cells. 3-PPA and 4-PBA significantly suppressed neuronal cell death caused by ER stress induced by the overexpression of Pael-R. Overexpressed Pael-R accumulated in the ER of cells. With 3-PPA and 4-PBA treatment, the localization of the overexpressed Pael-R shifted away from the ER to the cytoplasmic membrane. These results suggest that terminal aromatic substituted fatty acids are potential candidates for the treatment of neurodegenerative diseases.

  7. γ-Oryzanol protects pancreatic β-cells against endoplasmic reticulum stress in male mice.

    Science.gov (United States)

    Kozuka, Chisayo; Sunagawa, Sumito; Ueda, Rei; Higa, Moritake; Tanaka, Hideaki; Shimizu-Okabe, Chigusa; Ishiuchi, Shogo; Takayama, Chitoshi; Matsushita, Masayuki; Tsutsui, Masato; Miyazaki, Jun-ichi; Oyadomari, Seiichi; Shimabukuro, Michio; Masuzaki, Hiroaki

    2015-04-01

    Endoplasmic reticulum (ER) stress is profoundly involved in dysfunction of β-cells under high-fat diet and hyperglycemia. Our recent study in mice showed that γ-oryzanol, a unique component of brown rice, acts as a chemical chaperone in the hypothalamus and improves feeding behavior and diet-induced dysmetabolism. However, the entire mechanism whereby γ-oryzanol improves glucose metabolism throughout the body still remains unclear. In this context, we tested whether γ-oryzanol reduces ER stress and improves function and survival of pancreatic β-cells using murine β-cell line MIN6. In MIN6 cells with augmented ER stress by tunicamycin, γ-oryzanol decreased exaggerated expression of ER stress-related genes and phosphorylation of eukaryotic initiation factor-2α, resulting in restoration of glucose-stimulated insulin secretion and prevention of apoptosis. In islets from high-fat diet-fed diabetic mice, oral administration of γ-oryzanol improved glucose-stimulated insulin secretion on following reduction of exaggerated ER stress and apoptosis. Furthermore, we examined the impact of γ-oryzanol on low-dose streptozotocin-induced diabetic mice, where exaggerated ER stress and resultant apoptosis in β-cells were observed. Also in this model, γ-oryzanol attenuated mRNA level of genes involved in ER stress and apoptotic signaling in islets, leading to amelioration of glucose dysmetabolism. Taken together, our findings demonstrate that γ-oryzanol directly ameliorates ER stress-induced β-cell dysfunction and subsequent apoptosis, highlighting usefulness of γ-oryzanol for the treatment of diabetes mellitus.

  8. Role of radiation therapy in the management of ocular reticulum cell sarcoma

    International Nuclear Information System (INIS)

    Margolis, L.; Fraser, R.; Lichter, A.; Char, D.H.

    1980-01-01

    Nine patients with ocular lymphomas were seen in the Department of Ophthalmology and the Division of Radiation Oncology at UCSF and Ralph K. Davies Medical Center, San Francisco, from 1968 through 1974. Six of the 9 patients had visual symptoms as the first manifestation of their disease. Eight of the 9 patients developed intracranial lymphoma at some time during the course of the disease. Despite lymphoma work-up including bone marrow biopsies and lymphangiogram, only 1 patient was found to have documented systemic involvement. The diagnosis of ocular lymphoma was based on pathologic material from the eye in 5 cases or from central nervous system biopsy in 4 patients in association with tumor cell infiltrates in the retina and vitreous clouding. Radiation therapy to the eyes improved vision in 10 of 13 eyes treated in 8 patients. The usual dose was in the range of 3500 to 4500 rads given over 4 to 5 weeks. In addition, 7 patients received central nervous system irradiation. Review of the literature reinforced the findings of this series showing the frequent association of ocular lymphoma with intracranial lymphoma and the rare systemic dissemination. This disease process has previously been referred to as ocular reticulum cell sarcoma

  9. Endoplasmic reticulum (ER Chaperones and Oxidoreductases: Critical Regulators of Tumor Cell Survival and Immunorecognition

    Directory of Open Access Journals (Sweden)

    Thomas eSimmen

    2014-10-01

    Full Text Available Endoplasmic reticulum (ER chaperones and oxidoreductases are abundant enzymes that mediate the production of fully folded secretory and transmembrane proteins. Resisting the Golgi and plasma membrane-directed bulk flow, ER chaperones and oxidoreductases enter retrograde trafficking whenever they are pulled outside of the ER. However, solid tumors are characterized by the increased production of reactive oxygen species (ROS, combined with reduced blood flow that leads to low oxygen supply and ER stress. Under these conditions, hypoxia and the unfolded protein response (UPR upregulate ER chaperones and oxidoreductases. When this occurs, ER oxidoreductases and chaperones become important regulators of tumor growth. However, under these conditions, these proteins not only promote the production of proteins, but also alter the properties of the plasma membrane and hence modulate tumor immune recognition. For instance, high levels of calreticulin serve as an eat-me signal on the surface of tumor cells. Conversely, both intracellular and surface BiP/GRP78 promotes tumor growth. Other ER folding assistants able to modulate the properties of tumor tissue include protein disulfide isomerase (PDI, Ero1α and GRP94. Understanding the roles and mechanisms of ER chaperones in regulating tumor cell functions and immunorecognition will lead to important insight for the development of novel cancer therapies.

  10. GABAB receptor cell surface export is controlled by an endoplasmic reticulum gatekeeper

    Science.gov (United States)

    Doly, Stéphane; Shirvani, Hamasseh; Gäta, Gabriel; Meye, Frank; Emerit, Michel-Boris; Enslen, Hervé; Achour, Lamia; Pardo-Lopez, Liliana; Kwon, Yang Seung; Armand, Vincent; Gardette, Robert; Giros, Bruno; Gassmann, Martin; Bettler, Bernhard; Mameli, Manuel; Darmon, Michèle; Marullo, Stefano

    2016-01-01

    Summary Endoplasmic reticulum (ER) release and cell surface export of many G protein-coupled receptors (GPCRs), are tightly regulated. For GABAB receptors of GABA, the major mammalian inhibitory neurotransmitter, the ligand-binding GB1 subunit is maintained in the ER by unknown mechanisms in the absence of hetero-dimerization with the GB2 subunit. We report that GB1 retention is regulated by a specific gatekeeper, PRAF2. This ER resident transmembrane protein binds to GB1, preventing its progression in the biosynthetic pathway. GB1 release occurs upon competitive displacement from PRAF2 by GB2. PRAF2 concentration, relative to that of GB1 and GB2, tightly controls cell surface receptor density and controls GABAB function in neurons. Experimental perturbation of PRAF2 levels in vivo caused marked hyperactivity disorders in mice. These data reveal an unanticipated major impact of specific ER gate-keepers on GPCR function and identify PRAF2 as a new molecular target with therapeutic potential for psychiatric and neurological diseases involving GABAB function. PMID:26033241

  11. ATAD3 proteins: brokers of a mitochondria-endoplasmic reticulum connection in mammalian cells.

    Science.gov (United States)

    Baudier, Jacques

    2018-05-01

    In yeast, a sequence of physical and genetic interactions termed the endoplasmic reticulum (ER)-mitochondria organizing network (ERMIONE) controls mitochondria-ER interactions and mitochondrial biogenesis. Several functions that characterize ERMIONE complexes are conserved in mammalian cells, suggesting that a similar tethering complex must exist in metazoans. Recent studies have identified a new family of nuclear-encoded ATPases associated with diverse cellular activities (AAA+-ATPase) mitochondrial membrane proteins specific to multicellular eukaryotes, called the ATPase family AAA domain-containing protein 3 (ATAD3) proteins (ATAD3A and ATAD3B). These proteins are crucial for normal mitochondrial-ER interactions and lie at the heart of processes underlying mitochondrial biogenesis. ATAD3A orthologues have been studied in flies, worms, and mammals, highlighting the widespread importance of this gene during embryonic development and in adulthood. ATAD3A is a downstream effector of target of rapamycin (TOR) signalling in Drosophila and exhibits typical features of proteins from the ERMIONE-like complex in metazoans. In humans, mutations in the ATAD3A gene represent a new link between altered mitochondrial-ER interaction and recognizable neurological syndromes. The primate-specific ATAD3B protein is a biomarker of pluripotent embryonic stem cells. Through negative regulation of ATAD3A function, ATAD3B supports mitochondrial stemness properties. © 2017 Cambridge Philosophical Society.

  12. ERLIN2 promotes breast cancer cell survival by modulating endoplasmic reticulum stress pathways

    International Nuclear Information System (INIS)

    Wang, Guohui; Yang, Zeng-Quan; Liu, Gang; Wang, Xiaogang; Sethi, Seema; Ali-Fehmi, Rouba; Abrams, Judith; Zheng, Ze; Zhang, Kezhong; Ethier, Stephen

    2012-01-01

    Amplification of the 8p11-12 region has been found in approximately 15% of human breast cancer and is associated with poor prognosis. Previous genomic analysis has led us to identify the endoplasmic reticulum (ER) lipid raft-associated 2 (ERLIN2) gene as one of the candidate oncogenes within the 8p11-12 amplicon in human breast cancer, particularly in the luminal subtype. ERLIN2, an ER membrane protein, has recently been identified as a novel mediator of ER-associated degradation. Yet, the biological roles of ERLIN2 and molecular mechanisms by which ERLIN2 coordinates ER pathways in breast carcinogenesis remain unclear. We established the MCF10A-ERLIN2 cell line, which stably over expresses ERLIN2 in human nontransformed mammary epithelial cells (MCF10A) using the pLenti6/V5-ERLIN2 construct. ERLIN2 over expressing cells and their respective parental cell lines were assayed for in vitro transforming phenotypes. Next, we knocked down the ERLIN2 as well as the ER stress sensor IRE1α activity in the breast cancer cell lines to characterize the biological roles and molecular basis of the ERLIN2 in carcinogenesis. Finally, immunohistochemical staining was performed to detect ERLIN2 expression in normal and cancerous human breast tissues We found that amplification of the ERLIN2 gene and over expression of the ERLIN2 protein occurs in both luminal and Her2 subtypes of breast cancer. Gain- and loss-of-function approaches demonstrated that ERLIN2 is a novel oncogenic factor associated with the ER stress response pathway. The IRE1α/XBP1 axis in the ER stress pathway modulated expression of ERLIN2 protein levels in breast cancer cells. We also showed that over expression of ERLIN2 facilitated the adaptation of breast epithelial cells to ER stress by supporting cell growth and protecting the cells from ER stress-induced cell death. ERLIN2 may confer a selective growth advantage for breast cancer cells by facilitating a cytoprotective response to various cellular stresses

  13. ERLIN2 promotes breast cancer cell survival by modulating endoplasmic reticulum stress pathways

    Directory of Open Access Journals (Sweden)

    Wang Guohui

    2012-06-01

    Full Text Available Abstract Background Amplification of the 8p11-12 region has been found in approximately 15% of human breast cancer and is associated with poor prognosis. Previous genomic analysis has led us to identify the endoplasmic reticulum (ER lipid raft-associated 2 (ERLIN2 gene as one of the candidate oncogenes within the 8p11-12 amplicon in human breast cancer, particularly in the luminal subtype. ERLIN2, an ER membrane protein, has recently been identified as a novel mediator of ER-associated degradation. Yet, the biological roles of ERLIN2 and molecular mechanisms by which ERLIN2 coordinates ER pathways in breast carcinogenesis remain unclear. Methods We established the MCF10A-ERLIN2 cell line, which stably over expresses ERLIN2 in human nontransformed mammary epithelial cells (MCF10A using the pLenti6/V5-ERLIN2 construct. ERLIN2 over expressing cells and their respective parental cell lines were assayed for in vitro transforming phenotypes. Next, we knocked down the ERLIN2 as well as the ER stress sensor IRE1α activity in the breast cancer cell lines to characterize the biological roles and molecular basis of the ERLIN2 in carcinogenesis. Finally, immunohistochemical staining was performed to detect ERLIN2 expression in normal and cancerous human breast tissues Results We found that amplification of the ERLIN2 gene and over expression of the ERLIN2 protein occurs in both luminal and Her2 subtypes of breast cancer. Gain- and loss-of-function approaches demonstrated that ERLIN2 is a novel oncogenic factor associated with the ER stress response pathway. The IRE1α/XBP1 axis in the ER stress pathway modulated expression of ERLIN2 protein levels in breast cancer cells. We also showed that over expression of ERLIN2 facilitated the adaptation of breast epithelial cells to ER stress by supporting cell growth and protecting the cells from ER stress-induced cell death. Conclusions ERLIN2 may confer a selective growth advantage for breast cancer cells by

  14. Quantification of plasmodesmatal endoplasmic reticulum coupling between sieve elements and companion cells using fluorescence redistribution after photobleaching

    DEFF Research Database (Denmark)

    Martens, Helle; Roberts, Alison G.; Oparka, Karl J.

    2006-01-01

    retrieval along the pathway is an integral component of phloem function. GFP fluorescence was limited to CCs where it was visualized as a well-developed ER network in close proximity to the plasma membrane. ER coupling between CC and SEs was tested in wild-type tobacco using an ER-specific fluorochrome......Transgenic tobacco (Nicotiana tabacum) was studied to localize the activity of phloem loading during development and to establish whether the endoplasmic reticulum (ER) of the companion cell (CC) and the sieve element (SE) reticulum is continuous by using a SUC2 promoter-green fluorescent protein...... and fluorescence redistribution after photobleaching (FRAP), and showed that the ER is continuous via pore-plasmodesma units. ER coupling between CC and SE was quantified by determining the mobile fraction and half-life of fluorescence redistribution and compared with that of other cell types. In all tissues...

  15. Genes and Gene Networks Involved in Sodium Fluoride-Elicited Cell Death Accompanying Endoplasmic Reticulum Stress in Oral Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Yoshiaki Tabuchi

    2014-05-01

    Full Text Available Here, to understand the molecular mechanisms underlying cell death induced by sodium fluoride (NaF, we analyzed gene expression patterns in rat oral epithelial ROE2 cells exposed to NaF using global-scale microarrays and bioinformatics tools. A relatively high concentration of NaF (2 mM induced cell death concomitant with decreases in mitochondrial membrane potential, chromatin condensation and caspase-3 activation. Using 980 probe sets, we identified 432 up-regulated and 548 down-regulated genes, that were differentially expressed by >2.5-fold in the cells treated with 2 mM of NaF and categorized them into 4 groups by K-means clustering. Ingenuity® pathway analysis revealed several gene networks from gene clusters. The gene networks Up-I and Up-II included many up-regulated genes that were mainly associated with the biological function of induction or prevention of cell death, respectively, such as Atf3, Ddit3 and Fos (for Up-I and Atf4 and Hspa5 (for Up-II. Interestingly, knockdown of Ddit3 and Hspa5 significantly increased and decreased the number of viable cells, respectively. Moreover, several endoplasmic reticulum (ER stress-related genes including, Ddit3, Atf4 and Hapa5, were observed in these gene networks. These findings will provide further insight into the molecular mechanisms of NaF-induced cell death accompanying ER stress in oral epithelial cells.

  16. Endoplasmic Reticulum Stress Sensor IRE1α Enhances IL-23 Expression by Human Dendritic Cells

    Directory of Open Access Journals (Sweden)

    Saioa Márquez

    2017-06-01

    Full Text Available Human monocyte-derived dendritic cells (DCs exposed to pathogen-associated molecular patterns (PAMPs undergo bioenergetic changes that influence the immune response. We found that stimulation with PAMPs enhanced glycolysis in DCs, whereas oxidative phosphorylation remained unaltered. Glucose starvation and the hexokinase inhibitor 2-deoxy-d-glucose (2-DG modulated cytokine expression in stimulated DCs. Strikingly, IL23A was markedly induced upon 2-DG treatment, but not during glucose deprivation. Since 2-DG can also rapidly inhibit protein N-glycosylation, we postulated that this compound could induce IL-23 in DCs via activation of the endoplasmic reticulum (ER stress response. Indeed, stimulation of DCs with PAMPs in the presence of 2-DG robustly activated inositol-requiring protein 1α (IRE1α signaling and to a lesser extent the PERK arm of the unfolded protein response. Additional ER stressors such as tunicamycin and thapsigargin also promoted IL-23 expression by PAMP-stimulated DCs. Pharmacological, biochemical, and genetic analyses using conditional knockout mice revealed that IL-23 induction in ER stressed DCs stimulated with PAMPs was IRE1α/X-box binding protein 1-dependent upon zymosan stimulation. Interestingly, we further evidenced PERK-mediated and CAAT/enhancer-binding protein β-dependent trans-activation of IL23A upon lipopolysaccharide treatment. Our findings uncover that the ER stress response can potently modulate cytokine expression in PAMP-stimulated human DCs.

  17. Stem cells propagate their DNA by random segregation in the flatworm Macrostomum lignano.

    Directory of Open Access Journals (Sweden)

    Freija Verdoodt

    Full Text Available Adult stem cells are proposed to have acquired special features to prevent an accumulation of DNA-replication errors. Two such mechanisms, frequently suggested to serve this goal are cellular quiescence, and non-random segregation of DNA strands during stem cell division, a theory designated as the immortal strand hypothesis. To date, it has been difficult to test the in vivo relevance of both mechanisms in stem cell systems. It has been shown that in the flatworm Macrostomum lignano pluripotent stem cells (neoblasts are present in adult animals. We sought to address by which means M. lignano neoblasts protect themselves against the accumulation of genomic errors, by studying the exact mode of DNA-segregation during their division. In this study, we demonstrated four lines of in vivo evidence in favor of cellular quiescence. Firstly, performing BrdU pulse-chase experiments, we localized 'Label-Retaining Cells' (LRCs. Secondly, EDU pulse-chase combined with Vasa labeling demonstrated the presence of neoblasts among the LRCs, while the majority of LRCs were differentiated cells. We showed that stem cells lose their label at a slow rate, indicating cellular quiescence. Thirdly, CldU/IdU- double labeling studies confirmed that label-retaining stem cells showed low proliferative activity. Finally, the use of the actin inhibitor, cytochalasin D, unequivocally demonstrated random segregation of DNA-strands in LRCs. Altogether, our data unambiguously demonstrated that the majority of neoblasts in M. lignano distribute their DNA randomly during cell division, and that label-retention is a direct result of cellular quiescence, rather than a sign of co-segregation of labeled strands.

  18. Acidosis Activates Endoplasmic Reticulum Stress Pathways through GPR4 in Human Vascular Endothelial Cells.

    Science.gov (United States)

    Dong, Lixue; Krewson, Elizabeth A; Yang, Li V

    2017-01-27

    Acidosis commonly exists in the tissue microenvironment of various pathophysiological conditions such as tumors, inflammation, ischemia, metabolic disease, and respiratory disease. For instance, the tumor microenvironment is characterized by acidosis and hypoxia due to tumor heterogeneity, aerobic glycolysis (the "Warburg effect"), and the defective vasculature that cannot efficiently deliver oxygen and nutrients or remove metabolic acid byproduct. How the acidic microenvironment affects the function of blood vessels, however, is not well defined. GPR4 (G protein-coupled receptor 4) is a member of the proton-sensing G protein-coupled receptors and it has high expression in endothelial cells (ECs). We have previously reported that acidosis induces a broad inflammatory response in ECs. Acidosis also increases the expression of several endoplasmic reticulum (ER) stress response genes such as CHOP (C/EBP homologous protein) and ATF3 (activating transcription factor 3). In the current study, we have examined acidosis/GPR4- induced ER stress pathways in human umbilical vein endothelial cells (HUVEC) and other types of ECs. All three arms of the ER stress/unfolded protein response (UPR) pathways were activated by acidosis in ECs as an increased expression of phosphorylated eIF2α (eukaryotic initiation factor 2α), phosphorylated IRE1α (inositol-requiring enzyme 1α), and cleaved ATF6 upon acidic pH treatment was observed. The expression of other downstream mediators of the UPR, such as ATF4, ATF3, and spliced XBP-1 (X box-binding protein 1), was also induced by acidosis. Through genetic and pharmacological approaches to modulate the expression level or activity of GPR4 in HUVEC, we found that GPR4 plays an important role in mediating the ER stress response induced by acidosis. As ER stress/UPR can cause inflammation and cell apoptosis, acidosis/GPR4-induced ER stress pathways in ECs may regulate vascular growth and inflammatory response in the acidic microenvironment.

  19. Murine but not human basophil undergoes cell-specific proteolysis of a major endoplasmic reticulum chaperone.

    Directory of Open Access Journals (Sweden)

    Bei Liu

    Full Text Available Basophil has been implicated in anti-parasite defense, allergy and in polarizing T(H2 response. Mouse model has been commonly used to study basophil function although the difference between human and mouse basophils is underappreciated. As an essential chaperone for multiple Toll-like receptors and integrins in the endoplasmic reticulum, gp96 also participates in general protein homeostasis and in the ER unfolded protein response to ensure cell survival during stress. The roles of gp96 in basophil development are unknown.We genetically delete gp96 in mice and examined the expression of gp96 in basophils by Western blot and flow cytometry. We compared the expression pattern of gp96 between human and mouse basophils.We found that gp96 was dispensable for murine basophil development. Moreover, gp96 was cleaved by serine protease(s in murine but not human basophils leading to accumulation of a nun-functional N-terminal ∼50 kDa fragment and striking induction of the unfolded protein response. The alteration of gp96 was unique to basophils and was not observed in any other cell types including mast cells. We also demonstrated that the ectopic expression of a mouse-specific tryptase mMCP11 does not lead to gp96 cleavage in human basophils.Our study revealed a remarkable biochemical event of gp96 silencing in murine but not human basophils, highlighting the need for caution in using mouse models to infer the function of basophils in human immune response. Our study also reveals a novel mechanism of shutting down gp96 post-translationally in regulating its function.

  20. Murine but not human basophil undergoes cell-specific proteolysis of a major endoplasmic reticulum chaperone.

    Science.gov (United States)

    Liu, Bei; Staron, Matthew; Li, Zihai

    2012-01-01

    Basophil has been implicated in anti-parasite defense, allergy and in polarizing T(H)2 response. Mouse model has been commonly used to study basophil function although the difference between human and mouse basophils is underappreciated. As an essential chaperone for multiple Toll-like receptors and integrins in the endoplasmic reticulum, gp96 also participates in general protein homeostasis and in the ER unfolded protein response to ensure cell survival during stress. The roles of gp96 in basophil development are unknown. We genetically delete gp96 in mice and examined the expression of gp96 in basophils by Western blot and flow cytometry. We compared the expression pattern of gp96 between human and mouse basophils. We found that gp96 was dispensable for murine basophil development. Moreover, gp96 was cleaved by serine protease(s) in murine but not human basophils leading to accumulation of a nun-functional N-terminal ∼50 kDa fragment and striking induction of the unfolded protein response. The alteration of gp96 was unique to basophils and was not observed in any other cell types including mast cells. We also demonstrated that the ectopic expression of a mouse-specific tryptase mMCP11 does not lead to gp96 cleavage in human basophils. Our study revealed a remarkable biochemical event of gp96 silencing in murine but not human basophils, highlighting the need for caution in using mouse models to infer the function of basophils in human immune response. Our study also reveals a novel mechanism of shutting down gp96 post-translationally in regulating its function.

  1. Calcium Handling by Endoplasmic Reticulum and Mitochondria in a Cell Model of Huntington’s Disease

    Science.gov (United States)

    De Mario, Agnese; Scarlatti, Chiara; Costiniti, Veronica; Primerano, Simona; Lopreiato, Raffaele; Calì, Tito; Brini, Marisa; Giacomello, Marta; Carafoli, Ernesto

    2016-01-01

    Huntington disease (HD) is caused by the CAG (Q) expansion in exon 1 of the IT15 gene encoding a polyglutamine (poly-Q) stretch of the Huntingtin protein (Htt). In the wild type protein, the repeats specify a stretch of up 34 Q in the N-terminal portion of Htt. In the pathological protein (mHtt) the poly-Q tract is longer. Proteolytic cleavage of the protein liberates an N-terminal fragment containing the expanded poly-Q tract becomes harmful to cells, in particular to striatal neurons. The fragments cause the transcriptional dysfunction of genes that are essential for neuronal survival. Htt, however, could also have non-transcriptional effects, e.g. it could directly alter Ca2+ homeostasis and/or mitochondrial morphology and function. Ca2+ dyshomeostasis and mitochondrial dysfunction are considered important in the molecular aetiology of the disease. Here we have analyzed the effect of the overexpression of Htt fragments (18Q, wild type form, wtHtt and 150Q mutated form, mHtt) on Ca2+ homeostasis in striatal neuronal precursor cells (Q7/7). We have found that the transient overexpression of the Htt fragments increases Ca2+ transients in the mitochondria of cells stimulated with Ca2+-mobilizing agonists. The bulk Ca2+ transients in the cytosol were unaffected, but the Ca2+ content of the endoplasmic reticulum was significantly decreased in the case of mHtt expression. To rule out possible transcriptional effects due to the presence of mHtt, we have measured the mRNA level of a subunit of the respiratory chain complex II, whose expression is commonly altered in many HD models. No effects on the mRNA level was found suggesting that, in our experimental condition, transcriptional action of Htt is not occurring and that the effects on Ca2+ homeostasis were dependent to non-transcriptional mechanisms. PMID:26819834

  2. Calcium Handling by Endoplasmic Reticulum and Mitochondria in a Cell Model of Huntington's Disease.

    Science.gov (United States)

    De Mario, Agnese; Scarlatti, Chiara; Costiniti, Veronica; Primerano, Simona; Lopreiato, Raffaele; Calì, Tito; Brini, Marisa; Giacomello, Marta; Carafoli, Ernesto

    2016-01-06

    Huntington disease (HD) is caused by the CAG (Q) expansion in exon 1 of the IT15 gene encoding a polyglutamine (poly-Q) stretch of the Huntingtin protein (Htt). In the wild type protein, the repeats specify a stretch of up 34 Q in the N-terminal portion of Htt. In the pathological protein (mHtt) the poly-Q tract is longer. Proteolytic cleavage of the protein liberates an N-terminal fragment containing the expanded poly-Q tract becomes harmful to cells, in particular to striatal neurons. The fragments cause the transcriptional dysfunction of genes that are essential for neuronal survival. Htt, however, could also have non-transcriptional effects, e.g. it could directly alter Ca2+ homeostasis and/or mitochondrial morphology and function. Ca2+ dyshomeostasis and mitochondrial dysfunction are considered important in the molecular aetiology of the disease. Here we have analyzed the effect of the overexpression of Htt fragments (18Q, wild type form, wtHtt and 150Q mutated form, mHtt) on Ca2+ homeostasis in striatal neuronal precursor cells (Q7/7). We have found that the transient overexpression of the Htt fragments increases Ca2+ transients in the mitochondria of cells stimulated with Ca2+-mobilizing agonists. The bulk Ca2+ transients in the cytosol were unaffected, but the Ca2+ content of the endoplasmic reticulum was significantly decreased in the case of mHtt expression. To rule out possible transcriptional effects due to the presence of mHtt, we have measured the mRNA level of a subunit of the respiratory chain complex II, whose expression is commonly altered in many HD models. No effects on the mRNA level was found suggesting that, in our experimental condition, transcriptional action of Htt is not occurring and that the effects on Ca2+ homeostasis were dependent to non-transcriptional mechanisms.

  3. Exogenous FABP4 induces endoplasmic reticulum stress in HepG2 liver cells.

    Science.gov (United States)

    Bosquet, Alba; Guaita-Esteruelas, Sandra; Saavedra, Paula; Rodríguez-Calvo, Ricardo; Heras, Mercedes; Girona, Josefa; Masana, Lluís

    2016-06-01

    Fatty acid binding protein 4 (FABP4) is an intracellular fatty acid (FA) carrier protein that is, in part, secreted into circulation. Circulating FABP4 levels are increased in obesity, diabetes and other insulin resistance (IR) diseases. FAs contribute to IR by promoting endoplasmic reticulum stress (ER stress) and altering the insulin signaling pathway. The effect of FABP4 on ER stress in the liver is not known. The aim of this study was to investigate whether exogenous FABP4 (eFABP4) is involved in the lipid-induced ER stress in the liver. HepG2 cells were cultured with eFABP4 (40 ng/ml) with or without linoleic acid (LA, 200 μM) for 18 h. The expression of ER stress-related markers was determined by Western blotting (ATF6, EIF2α, IRE1 and ubiquitin) and real-time PCR (ATF6, CHOP, EIF2α and IRE1). Apoptosis was studied by flow cytometry using Annexin V-FITC and propidium iodide staining. eFABP4 increased the ER stress markers ATF6 and IRE1 in HepG2 cells. This effect led to insulin resistance mediated by changes in AKT and JNK phosphorylation. Furthermore, eFABP4 significantly induced both apoptosis, as assessed by flow cytometry, and CHOP expression, without affecting necrosis and ubiquitination. The presence of LA increased the ER stress response induced by eFABP4. eFABP4, per se, induces ER stress and potentiates the effect of LA in HepG2 cells, suggesting that FABP4 could be a link between obesity-associated metabolic abnormalities and hepatic IR mechanisms. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  4. Prion protein misfolding affects calcium homeostasis and sensitizes cells to endoplasmic reticulum stress.

    Directory of Open Access Journals (Sweden)

    Mauricio Torres

    2010-12-01

    Full Text Available Prion-related disorders (PrDs are fatal neurodegenerative disorders characterized by progressive neuronal impairment as well as the accumulation of an abnormally folded and protease resistant form of the cellular prion protein, termed PrP(RES. Altered endoplasmic reticulum (ER homeostasis is associated with the occurrence of neurodegeneration in sporadic, infectious and familial forms of PrDs. The ER operates as a major intracellular calcium store, playing a crucial role in pathological events related to neuronal dysfunction and death. Here we investigated the possible impact of PrP misfolding on ER calcium homeostasis in infectious and familial models of PrDs. Neuro2A cells chronically infected with scrapie prions showed decreased ER-calcium content that correlated with a stronger upregulation of UPR-inducible chaperones, and a higher sensitivity to ER stress-induced cell death. Overexpression of the calcium pump SERCA stimulated calcium release and increased the neurotoxicity observed after exposure of cells to brain-derived infectious PrP(RES. Furthermore, expression of PrP mutants that cause hereditary Creutzfeldt-Jakob disease or fatal familial insomnia led to accumulation of PrP(RES and their partial retention at the ER, associated with a drastic decrease of ER calcium content and higher susceptibility to ER stress. Finally, similar results were observed when a transmembrane form of PrP was expressed, which is proposed as a neurotoxic intermediate. Our results suggest that alterations in calcium homeostasis and increased susceptibility to ER stress are common pathological features of both infectious and familial PrD models.

  5. Uncovering a Dual Regulatory Role for Caspases During Endoplasmic Reticulum Stress-induced Cell Death.

    Science.gov (United States)

    Anania, Veronica G; Yu, Kebing; Gnad, Florian; Pferdehirt, Rebecca R; Li, Han; Ma, Taylur P; Jeon, Diana; Fortelny, Nikolaus; Forrest, William; Ashkenazi, Avi; Overall, Christopher M; Lill, Jennie R

    2016-07-01

    Many diseases are associated with endoplasmic reticulum (ER) stress, which results from an accumulation of misfolded proteins. This triggers an adaptive response called the "unfolded protein response" (UPR), and prolonged exposure to ER stress leads to cell death. Caspases are reported to play a critical role in ER stress-induced cell death but the underlying mechanisms by which they exert their effect continue to remain elusive. To understand the role caspases play during ER stress, a systems level approach integrating analysis of the transcriptome, proteome, and proteolytic substrate profile was employed. This quantitative analysis revealed transcriptional profiles for most human genes, provided information on protein abundance for 4476 proteins, and identified 445 caspase substrates. Based on these data sets many caspase substrates were shown to be downregulated at the protein level during ER stress suggesting caspase activity inhibits their cellular function. Additionally, RNA sequencing revealed a role for caspases in regulation of ER stress-induced transcriptional pathways and gene set enrichment analysis showed expression of multiple gene targets of essential transcription factors to be upregulated during ER stress upon inhibition of caspases. Furthermore, these transcription factors were degraded in a caspase-dependent manner during ER stress. These results indicate that caspases play a dual role in regulating the cellular response to ER stress through both post-translational and transcriptional regulatory mechanisms. Moreover, this study provides unique insight into progression of the unfolded protein response into cell death, which may help identify therapeutic strategies to treat ER stress-related diseases. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  6. Duplication and segregation of the actin (MreB) cytoskeleton during the prokaryotic cell cycle.

    Science.gov (United States)

    Vats, Purva; Rothfield, Lawrence

    2007-11-06

    The bacterial actin homolog MreB exists as a single-copy helical cytoskeletal structure that extends between the two poles of rod-shaped bacteria. In this study, we show that equipartition of the MreB cytoskeleton into daughter cells is accomplished by division and segregation of the helical MreB array into two equivalent structures located in opposite halves of the predivisional cell. This process ensures that each daughter cell inherits one copy of the MreB cytoskeleton. The process is triggered by the membrane association of the FtsZ cell division protein. The cytoskeletal division and segregation events occur before and independently of cytokinesis and involve specialized MreB structures that appear to be intermediates in this process.

  7. Antitumor agent 25-epi Ritterostatin GN1N induces endoplasmic reticulum stress and autophagy mediated cell death in melanoma cells.

    Science.gov (United States)

    Riaz Ahmed, Kausar Begam; Kanduluru, Ananda Kumar; Feng, Li; Fuchs, Philip L; Huang, Peng

    2017-05-01

    Metastatic melanoma is the most aggressive of all skin cancers and is associated with poor prognosis owing to lack of effective treatments. 25-epi Ritterostatin GN1N is a novel antitumor agent with yet undefined mechanisms of action. We sought to delineate the antitumor mechanisms of 25-epi Ritterostatin GN1N in melanoma cells to determine the potential of this compound as a treatment for melanoma. Activation of the endoplasmic reticulum (ER) stress protein glucose-regulated protein 78 (GRP78) has been associated with increased melanoma progression, oncogenic signaling, drug resistance, and suppression of cell death. We found that 25-epi Ritterostatin GN1N induced cell death in melanoma cells at nanomolar concentrations, and this cell death was characterized by inhibition of GRP78 expression, increased expression of the ER stress marker CHOP, loss of mitochondrial membrane potential, and lipidation of the autophagy marker protein LC3B. Importantly, normal melanocytes exhibited limited sensitivity to 25-epi Ritterostatin GN1N. Subsequent in vivo results demonstrated that 25-epi Ritterostatin GN1N reduced melanoma growth in mouse tumor xenografts and did not affect body weight, suggesting minimal toxicity. In summary, our findings indicate that 25-epi Ritterostatin GN1N causes ER stress and massive autophagy, leading to collapse of mitochondrial membrane potential and cell death in melanoma cells, with minimal effects in normal melanocytes. Thus, 25-epi Ritterostatin GN1N is a promising anticancer agent that warrants further investigation.

  8. Dll1 maintains quiescence of adult neural stem cells and segregates asymmetrically during mitosis.

    Science.gov (United States)

    Kawaguchi, Daichi; Furutachi, Shohei; Kawai, Hiroki; Hozumi, Katsuto; Gotoh, Yukiko

    2013-01-01

    Stem cells often divide asymmetrically to produce one stem cell and one differentiating cell, thus maintaining the stem cell pool. Although neural stem cells (NSCs) in the adult mouse subventricular zone have been suggested to divide asymmetrically, intrinsic cell fate determinants for asymmetric NSC division are largely unknown. Stem cell niches are important for stem cell maintenance, but the niche for the maintenance of adult quiescent NSCs has remained obscure. Here we show that the Notch ligand Delta-like 1 (Dll1) is required to maintain quiescent NSCs in the adult mouse subventricular zone. Dll1 protein is induced in activated NSCs and segregates to one daughter cell during mitosis. Dll1-expressing cells reside in close proximity to quiescent NSCs, suggesting a feedback signal for NSC maintenance by their sister cells and progeny. Our data suggest a model in which NSCs produce their own niche cells for their maintenance through asymmetric Dll1 inheritance at mitosis.

  9. Crosstalk between mitochondrial and sarcoplasmic reticulum Ca2+ cycling modulates cardiac pacemaker cell automaticity.

    Directory of Open Access Journals (Sweden)

    Yael Yaniv

    Full Text Available Mitochondria dynamically buffer cytosolic Ca(2+ in cardiac ventricular cells and this affects the Ca(2+ load of the sarcoplasmic reticulum (SR. In sinoatrial-node cells (SANC the SR generates periodic local, subsarcolemmal Ca(2+ releases (LCRs that depend upon the SR load and are involved in SANC automaticity: LCRs activate an inward Na(+-Ca(2+ exchange current to accelerate the diastolic depolarization, prompting the ensemble of surface membrane ion channels to generate the next action potential (AP.To determine if mitochondrial Ca(2+ (Ca(2+ (m, cytosolic Ca(2+ (Ca(2+ (c-SR-Ca(2+ crosstalk occurs in single rabbit SANC, and how this may relate to SANC normal automaticity.Inhibition of mitochondrial Ca(2+ influx into (Ru360 or Ca(2+ efflux from (CGP-37157 decreased [Ca(2+](m to 80 ± 8% control or increased [Ca(2+](m to 119 ± 7% control, respectively. Concurrent with inhibition of mitochondrial Ca(2+ influx or efflux, the SR Ca(2+ load, and LCR size, duration, amplitude and period (imaged via confocal linescan significantly increased or decreased, respectively. Changes in total ensemble LCR Ca(2+ signal were highly correlated with the change in the SR Ca(2+ load (r(2 = 0.97. Changes in the spontaneous AP cycle length (Ru360, 111 ± 1% control; CGP-37157, 89 ± 2% control in response to changes in [Ca(2+](m were predicted by concurrent changes in LCR period (r(2 = 0.84.A change in SANC Ca(2+ (m flux translates into a change in the AP firing rate by effecting changes in Ca(2+ (c and SR Ca(2+ loading, which affects the characteristics of spontaneous SR Ca(2+ release.

  10. Chemical chaperones reduce ionizing radiation-induced endoplasmic reticulum stress and cell death in IEC-6 cells

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Eun Sang; Lee, Hae-June; Lee, Yoon-Jin [Division of Radiation Effects, Korea Institute of Radiological and Medical Sciences, Seoul 139-706 (Korea, Republic of); Jeong, Jae-Hoon [Division of Radiotherapy, Korea Institute of Radiological and Medical Sciences, Seoul 139-706 (Korea, Republic of); Kang, Seongman [Division of Life Sciences, Korea University, Seoul 136-701 (Korea, Republic of); Lim, Young-Bin, E-mail: yblim@kirams.re.kr [Division of Radiation Effects, Korea Institute of Radiological and Medical Sciences, Seoul 139-706 (Korea, Republic of)

    2014-07-25

    Highlights: • UPR activation precedes caspase activation in irradiated IEC-6 cells. • Chemical ER stress inducers radiosensitize IEC-6 cells. • siRNAs that targeted ER stress responses ameliorate IR-induced cell death. • Chemical chaperons prevent cell death in irradiated IEC-6 cells. - Abstract: Radiotherapy, which is one of the most effective approaches to the treatment of various cancers, plays an important role in malignant cell eradication in the pelvic area and abdomen. However, it also generates some degree of intestinal injury. Apoptosis in the intestinal epithelium is the primary pathological factor that initiates radiation-induced intestinal injury, but the mechanism by which ionizing radiation (IR) induces apoptosis in the intestinal epithelium is not clearly understood. Recently, IR has been shown to induce endoplasmic reticulum (ER) stress, thereby activating the unfolded protein response (UPR) signaling pathway in intestinal epithelial cells. However, the consequences of the IR-induced activation of the UPR signaling pathway on radiosensitivity in intestinal epithelial cells remain to be determined. In this study, we investigated the role of ER stress responses in IR-induced intestinal epithelial cell death. We show that chemical ER stress inducers, such as tunicamycin or thapsigargin, enhanced IR-induced caspase 3 activation and DNA fragmentation in intestinal epithelial cells. Knockdown of Xbp1 or Atf6 with small interfering RNA inhibited IR-induced caspase 3 activation. Treatment with chemical chaperones prevented ER stress and subsequent apoptosis in IR-exposed intestinal epithelial cells. Our results suggest a pro-apoptotic role of ER stress in IR-exposed intestinal epithelial cells. Furthermore, inhibiting ER stress may be an effective strategy to prevent IR-induced intestinal injury.

  11. Pancreatic endoplasmic reticulum kinase activation promotes medulloblastoma cell migration and invasion through induction of vascular endothelial growth factor A.

    Directory of Open Access Journals (Sweden)

    Stephanie Jamison

    Full Text Available Evidence is accumulating that activation of the pancreatic endoplasmic reticulum kinase (PERK in response to endoplasmic reticulum (ER stress adapts tumor cells to the tumor microenvironment and enhances tumor angiogenesis by inducing vascular endothelial growth factor A (VEGF-A. Recent studies suggest that VEGF-A can act directly on certain tumor cell types in an autocrine manner, via binding to VEGF receptor 2 (VEGFR2, to promote tumor cell migration and invasion. Although several reports show that PERK activation increases VEGF-A expression in medulloblastoma, the most common solid malignancy of childhood, the role that either PERK or VEGF-A plays in medulloblastoma remains elusive. In this study, we mimicked the moderate enhancement of PERK activity observed in tumor patients using a genetic approach and a pharmacologic approach, and found that moderate activation of PERK signaling facilitated medulloblastoma cell migration and invasion and increased the production of VEGF-A. Moreover, using the VEGFR2 inhibitor SU5416 and the VEGF-A neutralizing antibody to block VEGF-A/VEGFR2 signaling, our results suggested that tumor cell-derived VEGF-A promoted medulloblastoma cell migration and invasion through VEGFR2 signaling, and that both VEGF-A and VEGFR2 were required for the promoting effects of PERK activation on medulloblastoma cell migration and invasion. Thus, these findings suggest that moderate PERK activation promotes medulloblastoma cell migration and invasion through enhancement of VEGF-A/VEGFR2 signaling.

  12. Role of endoplasmic reticulum stress in the loss of retinal ganglion cells in diabetic retinopathy

    Institute of Scientific and Technical Information of China (English)

    Liping Yang; Lemeng Wu; Dongmei Wang; Ying Li; Hongliang Dou; Mark OMTso; Zhizhong Ma

    2013-01-01

    Endoplasmic reticulum stress is closely involved in the early stage of diabetic retinopathy. In the present study, a streptozotocin-induced diabetic animal model was given an intraperitoneal injection of tauroursodeoxycholic acid. Results from immunofluorescent co-localization experiments showed that both caspase-12 protein and c-Jun N-terminal kinase 1 phosphorylation levels significantly in-creased, which was associated with retinal ganglion celldeath in diabetic retinas. The C/ERB ho-mologous protein pathway directly contributed to glial reactivity, and was subsequently responsible for neuronal loss and vascular abnormalities in diabetic retinopathy. Our experimental findings in-dicate that endoplasmic reticulum stress plays an important role in diabetes-induced retinal neu-ronal loss and vascular abnormalities, and that inhibiting the activation of the endoplasmic reticulum stress pathway provides effective protection against diabetic retinopathy.

  13. Light and Electrically Induced Phase Segregation and Its Impact on the Stability of Quadruple Cation High Bandgap Perovskite Solar Cells.

    Science.gov (United States)

    Duong, The; Mulmudi, Hemant Kumar; Wu, YiLiang; Fu, Xiao; Shen, Heping; Peng, Jun; Wu, Nandi; Nguyen, Hieu T; Macdonald, Daniel; Lockrey, Mark; White, Thomas P; Weber, Klaus; Catchpole, Kylie

    2017-08-16

    Perovskite material with a bandgap of 1.7-1.8 eV is highly desirable for the top cell in a tandem configuration with a lower bandgap bottom cell, such as a silicon cell. This can be achieved by alloying iodide and bromide anions, but light-induced phase-segregation phenomena are often observed in perovskite films of this kind, with implications for solar cell efficiency. Here, we investigate light-induced phase segregation inside quadruple-cation perovskite material in a complete cell structure and find that the magnitude of this phenomenon is dependent on the operating condition of the solar cell. Under short-circuit and even maximum power point conditions, phase segregation is found to be negligible compared to the magnitude of segregation under open-circuit conditions. In accordance with the finding, perovskite cells based on quadruple-cation perovskite with 1.73 eV bandgap retain 94% of the original efficiency after 12 h operation at the maximum power point, while the cell only retains 82% of the original efficiency after 12 h operation at the open-circuit condition. This result highlights the need to have standard methods including light/dark and bias condition for testing the stability of perovskite solar cells. Additionally, phase segregation is observed when the cell was forward biased at 1.2 V in the dark, which indicates that photoexcitation is not required to induce phase segregation.

  14. Curcumin induces apoptotic cell death of activated human CD4+ T cells via increasing endoplasmic reticulum stress and mitochondrial dysfunction.

    Science.gov (United States)

    Zheng, Min; Zhang, Qinggao; Joe, Yeonsoo; Lee, Bong Hee; Ryu, Do Gon; Kwon, Kang Beom; Ryter, Stefan W; Chung, Hun Taeg

    2013-03-01

    Curcumin, a natural polyphenolic antioxidant compound, exerts well-known anti-inflammatory and immunomodulatory effects, the latter which can influence the activation of immune cells including T cells. Furthermore, curcumin can inhibit the expression of pro-inflammatory cytokines and chemokines, through suppression of the NF-κB signaling pathway. The beneficial effects of curcumin in diseases such as arthritis, allergy, asthma, atherosclerosis, diabetes and cancer may be due to its immunomodulatory properties. We studied the potential of curcumin to modulate CD4+ T cells-mediated autoimmune disease, by examining the effects of this compound on human CD4+ lymphocyte activation. Stimulation of human T cells with PHA or CD3/CD28 induced IL-2 mRNA expression and activated the endoplasmic reticulum (ER) stress response. The treatment of T cells with curcumin induced the unfolded protein response (UPR) signaling pathway, initiated by the phosphorylation of PERK and IRE1. Furthermore, curcumin increased the expression of the ER stress associated transcriptional factors XBP-1, cleaved p50ATF6α and C/EBP homologous protein (CHOP) in human CD4+ and Jurkat T cells. In PHA-activated T cells, curcumin further enhanced PHA-induced CHOP expression and reduced the expression of the anti-apoptotic protein Bcl-2. Finally, curcumin treatment induced apoptotic cell death in activated T cells via eliciting an excessive ER stress response, which was reversed by the ER-stress inhibitor 4-phenylbutyric acid or transfection with CHOP-specific siRNA. These results suggest that curcumin can impact both ER stress and mitochondria functional pathways, and thereby could be used as a promising therapy in the context of Th1-mediated autoimmune diseases. Copyright © 2013 Elsevier B.V. All rights reserved.

  15. Acrolein activates cell survival and apoptotic death responses involving the endoplasmic reticulum in A549 lung cells.

    Science.gov (United States)

    Tanel, André; Pallepati, Pragathi; Bettaieb, Ahmed; Morin, Patrick; Averill-Bates, Diana A

    2014-05-01

    Acrolein, a highly reactive α,β-unsaturated aldehyde, is a product of endogenous lipid peroxidation. It is a ubiquitous environmental pollutant that is generated mainly by smoke, overheated cooking oil and vehicle exhaust. Acrolein damages cellular proteins, which could lead to accumulation of aberrantly-folded proteins in the endoplasmic reticulum (ER). This study determines the mechanisms involved in acrolein-induced apoptosis mediated by the ER and possible links with the ER stress response in human A549 lung cells. The exposure of cells to acrolein (15-50μM) for shorter times of 15 to 30min activated several ER stress markers. These included the ER chaperone protein BiP and the three ER sensors: (i) the survival/rescue molecules protein kinase RNA (PKR)-like ER kinase (PERK) and eukaryotic initiation factor 2 alpha (eIF2α) were phosphorylated; (ii) cleavage of activating transcription factor 6 (ATF6) occurred, and (iii) inositol-requiring protein-1 alpha (IRE1α) was phosphorylated. Acrolein (25-50μM) caused apoptotic cell death mediated by the ER after 2h, which was characterised by the induction of CHOP and activation of ER proteases calpain and caspase-4. Calpain and caspase-7 were the initiating factors for caspase-4 activation in acrolein-induced apoptosis. These results increase our knowledge about cellular responses to acrolein in lung cells, which have implications for human health. Copyright © 2014 Elsevier B.V. All rights reserved.

  16. P190B RhoGAP Regulates Chromosome Segregation in Cancer Cells

    International Nuclear Information System (INIS)

    Hwang, Melissa; Peddibhotla, Sirisha; McHenry, Peter; Chang, Peggy; Yochum, Zachary; Park, Ko Un; Sears, James Cooper; Vargo-Gogola, Tracy

    2012-01-01

    Rho GTPases are overexpressed and hyperactivated in many cancers, including breast cancer. Rho proteins, as well as their regulators and effectors, have been implicated in mitosis, and their altered expression promotes mitotic defects and aneuploidy. Previously, we demonstrated that p190B Rho GTPase activating protein (RhoGAP) deficiency inhibits ErbB2-induced mammary tumor formation in mice. Here we describe a novel role for p190B as a regulator of mitosis. We found that p190B localized to centrosomes during interphase and mitosis, and that it is differentially phosphorylated during mitosis. Knockdown of p190B expression in MCF-7 and Hela cells increased the incidence of aberrant microtubule-kinetochore attachments at metaphase, lagging chromosomes at anaphase, and micronucleation, all of which are indicative of aneuploidy. Cell cycle analysis of p190B deficient MCF-7 cells revealed a significant increase in apoptotic cells with a concomitant decrease in cells in G1 and S phase, suggesting that p190B deficient cells die at the G1 to S transition. Chemical inhibition of the Rac GTPase during mitosis reduced the incidence of lagging chromosomes in p190B knockdown cells to levels detected in control cells, suggesting that aberrant Rac activity in the absence of p190B promotes chromosome segregation defects. Taken together, these data suggest that p190B regulates chromosome segregation and apoptosis in cancer cells. We propose that disruption of mitosis may be one mechanism by which p190B deficiency inhibits tumorigenesis

  17. P190B RhoGAP Regulates Chromosome Segregation in Cancer Cells

    Energy Technology Data Exchange (ETDEWEB)

    Hwang, Melissa [Department of Biochemistry and Molecular Biology and the Indiana University Simon Cancer Center, Indiana University School of Medicine, 1234 Notre Dame Avenue, South Bend, IN 46617 (United States); Peddibhotla, Sirisha [Department of Molecular and Human Genetics, Baylor College of Medicine, John P. McGovern Campus, NABS-0250, Houston, TX 77030 (United States); McHenry, Peter [Department of Biology, Southwestern Adventist University, 100 W. Hillcrest, Keene, TX 76059 (United States); Chang, Peggy; Yochum, Zachary; Park, Ko Un; Sears, James Cooper; Vargo-Gogola, Tracy, E-mail: vargo-gogola.1@nd.edu [Department of Biochemistry and Molecular Biology and the Indiana University Simon Cancer Center, Indiana University School of Medicine, 1234 Notre Dame Avenue, South Bend, IN 46617 (United States)

    2012-04-25

    Rho GTPases are overexpressed and hyperactivated in many cancers, including breast cancer. Rho proteins, as well as their regulators and effectors, have been implicated in mitosis, and their altered expression promotes mitotic defects and aneuploidy. Previously, we demonstrated that p190B Rho GTPase activating protein (RhoGAP) deficiency inhibits ErbB2-induced mammary tumor formation in mice. Here we describe a novel role for p190B as a regulator of mitosis. We found that p190B localized to centrosomes during interphase and mitosis, and that it is differentially phosphorylated during mitosis. Knockdown of p190B expression in MCF-7 and Hela cells increased the incidence of aberrant microtubule-kinetochore attachments at metaphase, lagging chromosomes at anaphase, and micronucleation, all of which are indicative of aneuploidy. Cell cycle analysis of p190B deficient MCF-7 cells revealed a significant increase in apoptotic cells with a concomitant decrease in cells in G1 and S phase, suggesting that p190B deficient cells die at the G1 to S transition. Chemical inhibition of the Rac GTPase during mitosis reduced the incidence of lagging chromosomes in p190B knockdown cells to levels detected in control cells, suggesting that aberrant Rac activity in the absence of p190B promotes chromosome segregation defects. Taken together, these data suggest that p190B regulates chromosome segregation and apoptosis in cancer cells. We propose that disruption of mitosis may be one mechanism by which p190B deficiency inhibits tumorigenesis.

  18. Morphology versus Vertical Phase Segregation in Solvent Annealed Small Molecule Bulk Heterojunction Organic Solar Cells

    Directory of Open Access Journals (Sweden)

    Alexander Kovalenko

    2015-01-01

    Full Text Available The deep study of solvent annealed small molecules bulk heterojunction organic solar cells based on DPP(TBFu2 : PC60BM blend is carried out. To reveal the reason of the solvent annealing advantage over the thermal one, capacitance-voltage measurements were applied. It was found that controlling the vertical phase segregation in the solar cells a high fullerene population in the vicinity of the cathode could be achieved. This results in increase of the shunt resistance of the cell, thus improving the light harvesting efficiency.

  19. A mathematical model for eph/ephrin-directed segregation of intermingled cells.

    Directory of Open Access Journals (Sweden)

    Rotem Aharon

    Full Text Available Eph receptors, the largest family of receptor tyrosine kinases, control cell-cell adhesion/de-adhesion, cell morphology and cell positioning through interaction with cell surface ephrin ligands. Bi-directional signalling from the Eph and ephrin complexes on interacting cells have a significant role in controlling normal tissue development and oncogenic tissue patterning. Eph-mediated tissue patterning is based on the fine-tuned balance of adhesion and de-adhesion reactions between distinct Eph- and ephrin-expressing cell populations, and adhesion within like populations (expressing either Eph or ephrin. Here we develop a stochastic, Lagrangian model that is based on Eph/ephrin biology: incorporating independent Brownian motion to describe cell movement and a deterministic term (the drift term to represent repulsive and adhesive interactions between neighbouring cells. Comparison between the experimental and computer simulated Eph/ephrin cell patterning events shows that the model recapitulates the dynamics of cell-cell segregation and cell cluster formation. Moreover, by modulating the term for Eph/ephrin-mediated repulsion, the model can be tuned to match the actual behaviour of cells with different levels of Eph expression or activity. Together the results of our experiments and modelling suggest that the complexity of Eph/ephrin signalling mechanisms that control cell-cell interactions can be described well by a mathematical model with a single term balancing adhesion and de-adhesion between interacting cells. This model allows reliable prediction of Eph/ephrin-dependent control of cell patterning behaviour.

  20. Dipeptidyl peptidase-4 inhibitor, vildagliptin, inhibits pancreatic beta cell apoptosis in association with its effects suppressing endoplasmic reticulum stress in db/db mice.

    Science.gov (United States)

    Wu, Yan-ju; Guo, Xin; Li, Chun-jun; Li, Dai-qing; Zhang, Jie; Yang, Yiping; Kong, Yan; Guo, Hang; Liu, De-min; Chen, Li-ming

    2015-02-01

    Vildagliptin promotes beta cell survival by inhibiting cell apoptosis. It has been suggested that chronic ER (endoplasmic reticulum) stress triggers beta cell apoptosis. The objective of the study is to explore whether the pro-survival effect of vildagliptin is associated with attenuation of endoplasmic reticulum stress in islets of db/db mice. Vildagliptin was orally administered to db/db mice for 6 weeks, followed by evaluation of beta cell apoptosis by caspase3 activity and TUNEL staining method. Endoplasmic reticulum stress markers were determined with quantitative RT-PCR, immunohistochemistry and immunoblot analysis. After 6 weeks of treatment, vildagliptin treatment increased plasma active GLP-1 levels (22.63±1.19 vs. 11.69±0.44, Pvildagliptin treatment down-regulated several genes related to endoplasmic reticulum stress including TRIB3 (tribbles homolog 3) (15.9±0.4 vs. 33.3±1.7, ×10⁻³, PVildagliptin promoted beta cell survival in db/db mice in association with down-regulating markers of endoplasmic reticulum stress including TRIB3, ATF-4 as well as CHOP. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

  1. The Effect of Bornyl cis-4-Hydroxycinnamate on Melanoma Cell Apoptosis Is Associated with Mitochondrial Dysfunction and Endoplasmic Reticulum Stress

    Directory of Open Access Journals (Sweden)

    Tzu-Yen Yang

    2018-05-01

    Full Text Available Bornyl cis-4-hydroxycinnamate, an active compound isolated from Piper betle stems, was investigated in terms of its effects on A2058 and A375 melanoma cell proliferation and protein expression in this study. We used flow cytometric analysis to examine the early stages of apoptosis induced by bornyl cis-4-hydroxycinnamate in the two melanoma cell lines and employed comparative proteomic analysis to investigate the effects of this compound on protein expression in A375 cells. Master maps generated by PDQuest software from two-dimensional electrophoresis (2-DE analysis of A375 cells showed that the expression levels of 35 proteins were significantly altered, with 18 proteins upregulated and 17 downregulated. The proteomics study identified several proteins that are involved in mitochondrial dysfunction and endoplasmic reticulum stress (ER stress, in addition to apoptosis-associated proteins, including prohibitin, hypoxia-upregulated protein 1, stress 70 protein, 78 kDa glucose-regulated protein (GRP78, and protein deglycase DJ-1 (protein DJ-1 in melanoma cells exposed to bornyl cis-4-hydroxycinnamate. The treatment also resulted in a marked decline of the mitochondrial membrane potential, in cytochrome C release into the cytosol, in the activation of Bcl-2-associated X protein (Bax, Bcl-2-associated death promoter protein (Bad, caspase-3, and caspase-9, and in the decreased expression of p-Bad, B-cell lymphoma 2 (Bcl-2, Bcl-xl, and induced myeloid leukemia cell differentiation protein-1 (Mcl-1, indicating that apoptosis induced by bornyl cis-4-hydroxycinnamate was mediated by the mitochondria through the caspase-dependent pathway. Also, salubrinal (an eukaryotic initiation factor 2α inhibitor; eIF2α inhibitor was able to protect the cells from bornyl cis-4-hydroxycinnamate-induced apoptosis. Bornyl cis-4-hydroxycinnamate-related cell death also implied that the protein kinase R-like endoplasmic reticulum kinase (PERK–eIF2α–ATF4–CHOP signal

  2. The Effect of Bornyl cis-4-Hydroxycinnamate on Melanoma Cell Apoptosis Is Associated with Mitochondrial Dysfunction and Endoplasmic Reticulum Stress

    Science.gov (United States)

    Yang, Tzu-Yen; Wu, Yu-Jen; Chang, Chi-I; Wu, Mei-Li

    2018-01-01

    Bornyl cis-4-hydroxycinnamate, an active compound isolated from Piper betle stems, was investigated in terms of its effects on A2058 and A375 melanoma cell proliferation and protein expression in this study. We used flow cytometric analysis to examine the early stages of apoptosis induced by bornyl cis-4-hydroxycinnamate in the two melanoma cell lines and employed comparative proteomic analysis to investigate the effects of this compound on protein expression in A375 cells. Master maps generated by PDQuest software from two-dimensional electrophoresis (2-DE) analysis of A375 cells showed that the expression levels of 35 proteins were significantly altered, with 18 proteins upregulated and 17 downregulated. The proteomics study identified several proteins that are involved in mitochondrial dysfunction and endoplasmic reticulum stress (ER stress), in addition to apoptosis-associated proteins, including prohibitin, hypoxia-upregulated protein 1, stress 70 protein, 78 kDa glucose-regulated protein (GRP78), and protein deglycase DJ-1 (protein DJ-1) in melanoma cells exposed to bornyl cis-4-hydroxycinnamate. The treatment also resulted in a marked decline of the mitochondrial membrane potential, in cytochrome C release into the cytosol, in the activation of Bcl-2-associated X protein (Bax), Bcl-2-associated death promoter protein (Bad), caspase-3, and caspase-9, and in the decreased expression of p-Bad, B-cell lymphoma 2 (Bcl-2), Bcl-xl, and induced myeloid leukemia cell differentiation protein-1 (Mcl-1), indicating that apoptosis induced by bornyl cis-4-hydroxycinnamate was mediated by the mitochondria through the caspase-dependent pathway. Also, salubrinal (an eukaryotic initiation factor 2α inhibitor; eIF2α inhibitor) was able to protect the cells from bornyl cis-4-hydroxycinnamate-induced apoptosis. Bornyl cis-4-hydroxycinnamate-related cell death also implied that the protein kinase R-like endoplasmic reticulum kinase (PERK)–eIF2α–ATF4–CHOP signal pathways

  3. Intravenous Lipid Infusion Induces Endoplasmic Reticulum Stress in Endothelial Cells and Blood Mononuclear Cells of Healthy Adults.

    Science.gov (United States)

    Tampakakis, Emmanouil; Tabit, Corey E; Holbrook, Monika; Linder, Erika A; Berk, Brittany D; Frame, Alissa A; Bretón-Romero, Rosa; Fetterman, Jessica L; Gokce, Noyan; Vita, Joseph A; Hamburg, Naomi M

    2016-01-11

    Endoplasmic reticulum (ER) stress and the subsequent unfolded protein response may initially be protective, but when prolonged, have been implicated in atherogenesis in diabetic conditions. Triglycerides and free fatty acids (FFAs) are elevated in patients with diabetes and may contribute to ER stress. We sought to evaluate the effect of acute FFA elevation on ER stress in endothelial and circulating white cells. Twenty-one healthy subjects were treated with intralipid (20%; 45 mL/h) plus heparin (12 U/kg/h) infusion for 5 hours. Along with increased triglyceride and FFA levels, intralipid/heparin infusion reduced the calf reactive hyperemic response without a change in conduit artery flow-mediated dilation consistent with microvascular dysfunction. To investigate the short-term effects of elevated triglycerides and FFA, we measured markers of ER stress in peripheral blood mononuclear cells (PBMCs) and vascular endothelial cells (VECs). In VECs, activating transcription factor 6 (ATF6) and phospho-inositol requiring kinase 1 (pIRE1) proteins were elevated after infusion (both P<0.05). In PBMCs, ATF6 and spliced X-box-binding protein 1 (XBP-1) gene expression increased by 2.0- and 2.5-fold, respectively (both P<0.05), whereas CHOP and GADD34 decreased by ≈67% and 74%, respectively (both P<0.01). ATF6 and pIRE1 protein levels also increased (both P<0.05), and confocal microscopy revealed the nuclear localization of ATF6 after infusion, suggesting activation. Along with microvascular dysfunction, intralipid infusion induced an early protective ER stress response evidenced by activation of ATF6 and IRE1 in both leukocytes and endothelial cells. Our results suggest a potential link between metabolic disturbances and ER stress that may be relevant to vascular disease. © 2016 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley Blackwell.

  4. Understanding odor information segregation in the olfactory bulb by means of mitral and tufted cells.

    Directory of Open Access Journals (Sweden)

    Davide Polese

    Full Text Available Odor identification is one of the main tasks of the olfactory system. It is performed almost independently from the concentration of the odor providing a robust recognition. This capacity to ignore concentration information does not preclude the olfactory system from estimating concentration itself. Significant experimental evidence has indicated that the olfactory system is able to infer simultaneously odor identity and intensity. However, it is still unclear at what level or levels of the olfactory pathway this segregation of information occurs. In this work, we study whether this odor information segregation is performed at the input stage of the olfactory bulb: the glomerular layer. To this end, we built a detailed neural model of the glomerular layer based on its known anatomical connections and conducted two simulated odor experiments. In the first experiment, the model was exposed to an odor stimulus dataset composed of six different odorants, each one dosed at six different concentrations. In the second experiment, we conducted an odor morphing experiment where a sequence of binary mixtures going from one odor to another through intermediate mixtures was presented to the model. The results of the experiments were visualized using principal components analysis and analyzed with hierarchical clustering to unveil the structure of the high-dimensional output space. Additionally, Fisher's discriminant ratio and Pearson's correlation coefficient were used to quantify odor identity and odor concentration information respectively. Our results showed that the architecture of the glomerular layer was able to mediate the segregation of odor information obtaining output spiking sequences of the principal neurons, namely the mitral and external tufted cells, strongly correlated with odor identity and concentration, respectively. An important conclusion is also that the morphological difference between the principal neurons is not key to achieve odor

  5. Thiamine deficiency induces endoplasmic reticulum stress and oxidative stress in human neurons derived from induced pluripotent stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Xin; Xu, Mei; Frank, Jacqueline A. [Department of Pharmacology and Nutritional Sciences, University of Kentucky College of Medicine, Lexington, KY 40536 (United States); Ke, Zun-ji [Department of Biochemistry, Shanghai University of Traditional Chinese Medicine, Shanghai, China 201203 (China); Luo, Jia, E-mail: jialuo888@uky.edu [Department of Pharmacology and Nutritional Sciences, University of Kentucky College of Medicine, Lexington, KY 40536 (United States); Department of Biochemistry, Shanghai University of Traditional Chinese Medicine, Shanghai, China 201203 (China)

    2017-04-01

    Thiamine (vitamin B1) deficiency (TD) plays a major role in the etiology of Wernicke's encephalopathy (WE) which is a severe neurological disorder. TD induces selective neuronal cell death, neuroinflammation, endoplasmic reticulum (ER) stress and oxidative stress in the brain which are commonly observed in many aging-related neurodegenerative diseases, such as Alzheimer's disease (AD), Parkinson's disease (PD), Huntington's disease (HD) and progressive supranuclear palsy (PSP). However, the underlying cellular and molecular mechanisms remain unclear. The progress in this line of research is hindered due to the lack of appropriate in vitro models. The neurons derived for the human induced pluripotent stem cells (hiPSCs) provide a relevant and powerful tool for the research in pharmaceutical and environmental neurotoxicity. In this study, we for the first time used human induced pluripotent stem cells (hiPSCs)-derived neurons (iCell neurons) to investigate the mechanisms of TD-induced neurodegeneration. We showed that TD caused a concentration- and duration-dependent death of iCell neurons. TD induced ER stress which was evident by the increase in ER stress markers, such as GRP78, XBP-1, CHOP, ATF-6, phosphorylated eIF2α, and cleaved caspase-12. TD also triggered oxidative stress which was shown by the increase in the expression 2,4-dinitrophenyl (DNP) and 4-hydroxynonenal (HNE). ER stress inhibitors (STF-083010 and salubrinal) and antioxidant N-acetyl cysteine (NAC) were effective in alleviating TD-induced death of iCell neurons, supporting the involvement of ER stress and oxidative stress. It establishes that the iCell neurons are a novel tool to investigate cellular and molecular mechanisms for TD-induced neurodegeneration. - Highlights: • Thiamine deficiency (TD) causes death of human neurons in culture. • TD induces both endoplasmic reticulum (ER) stress and oxidative stress. • Alleviating ER stress and oxidative stress reduces TD

  6. Thiamine deficiency induces endoplasmic reticulum stress and oxidative stress in human neurons derived from induced pluripotent stem cells

    International Nuclear Information System (INIS)

    Wang, Xin; Xu, Mei; Frank, Jacqueline A.; Ke, Zun-ji; Luo, Jia

    2017-01-01

    Thiamine (vitamin B1) deficiency (TD) plays a major role in the etiology of Wernicke's encephalopathy (WE) which is a severe neurological disorder. TD induces selective neuronal cell death, neuroinflammation, endoplasmic reticulum (ER) stress and oxidative stress in the brain which are commonly observed in many aging-related neurodegenerative diseases, such as Alzheimer's disease (AD), Parkinson's disease (PD), Huntington's disease (HD) and progressive supranuclear palsy (PSP). However, the underlying cellular and molecular mechanisms remain unclear. The progress in this line of research is hindered due to the lack of appropriate in vitro models. The neurons derived for the human induced pluripotent stem cells (hiPSCs) provide a relevant and powerful tool for the research in pharmaceutical and environmental neurotoxicity. In this study, we for the first time used human induced pluripotent stem cells (hiPSCs)-derived neurons (iCell neurons) to investigate the mechanisms of TD-induced neurodegeneration. We showed that TD caused a concentration- and duration-dependent death of iCell neurons. TD induced ER stress which was evident by the increase in ER stress markers, such as GRP78, XBP-1, CHOP, ATF-6, phosphorylated eIF2α, and cleaved caspase-12. TD also triggered oxidative stress which was shown by the increase in the expression 2,4-dinitrophenyl (DNP) and 4-hydroxynonenal (HNE). ER stress inhibitors (STF-083010 and salubrinal) and antioxidant N-acetyl cysteine (NAC) were effective in alleviating TD-induced death of iCell neurons, supporting the involvement of ER stress and oxidative stress. It establishes that the iCell neurons are a novel tool to investigate cellular and molecular mechanisms for TD-induced neurodegeneration. - Highlights: • Thiamine deficiency (TD) causes death of human neurons in culture. • TD induces both endoplasmic reticulum (ER) stress and oxidative stress. • Alleviating ER stress and oxidative stress reduces TD

  7. Endoplasmic reticulum stress-induced resistance to doxorubicin is reversed by paeonol treatment in human hepatocellular carcinoma cells.

    Directory of Open Access Journals (Sweden)

    Lulu Fan

    Full Text Available BACKGROUND: Endoplasmic reticulum stress (ER stress is generally activated in solid tumors and results in tumor cell anti-apoptosis and drug resistance. Paeonol (Pae, 2-hydroxy-4-methoxyacetophenone, is a natural product extracted from the root of Paeonia Suffruticosa Andrew. Although Pae displays anti-neoplastic activity and increases the efficacy of chemotherapeutic drugs in various cell lines and in animal models, studies related to the effect of Pae on ER stress-induced resistance to chemotherapeutic agents in hepatocellular carcinoma (HCC are poorly understood. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we investigated the effect of the endoplasmic reticulum (ER stress response during resistance of human hepatocellular carcinoma cells to doxorubicin. Treatment with the ER stress-inducer tunicamycin (TM before the addition of doxorubicin reduced the rate of apoptosis induced by doxorubicin. Interestingly, co-pretreatment with tunicamycin and Pae significantly increased apoptosis induced by doxorubicin. Furthermore, induction of ER stress resulted in increasing expression of COX-2 concomitant with inactivation of Akt and up-regulation of the pro-apoptotic transcription factor CHOP (GADD153 in HepG2 cells. These cellular changes in gene expression and Akt activation may be an important resistance mechanism against doxorubicin in hepatocellular carcinoma cells undergoing ER stress. However, co-pretreatment with tunicamycin and Pae decreased the expression of COX-2 and levels of activation of Akt as well as increasing the levels of CHOP in HCC cells. CONCLUSIONS/SIGNIFICANCE: Our results demonstrate that Pae reverses ER stress-induced resistance to doxorubicin in human hepatocellular carcinoma cells by targeting COX-2 mediated inactivation of PI3K/AKT/CHOP.

  8. N-acetylcysteine protects against cadmium-induced germ cell apoptosis by inhibiting endoplasmic reticulum stress in testes.

    Science.gov (United States)

    Ji, Yan-Li; Wang, Hua; Zhang, Cheng; Zhang, Ying; Zhao, Mei; Chen, Yuan-Hua; Xu, De-Xiang

    2013-03-01

    Cadmium (Cd) is a reproductive toxicant that induces germ cell apoptosis in the testes. Previous studies have demonstrated that endoplasmic reticulum (ER) stress is involved in Cd-induced germ cell apoptosis. The aim of the present study was to investigate the effects of N-acetylcysteine (NAC), an antioxidant, on Cd-induced ER stress and germ cell apoptosis in the testes. Male CD-1 mice were intraperitoneally injected with CdCl2 (2.0 mg kg(-1)). As expected, acute Cd exposure induced germ cell apoptosis in the testes, as determined by terminal dUTP nick-end labelling (TUNEL). However, the administration of NAC alleviated Cd-induced germ cell apoptosis in the testes. Further analysis showed that NAC attenuated the Cd-induced upregulation of testicular glucose-regulated protein 78 (GRP78), an important ER molecular chaperone. Moreover, NAC inhibited the Cd-induced phosphorylation of testicular eukaryotic translation initiation factor 2α (eIF2α), a downstream target of the double-stranded RNA-activated kinase-like ER kinase (PERK) pathway. In addition, NAC blocked the Cd-induced activation of testicular X binding protein (XBP)-1, indicating that NAC attenuates the Cd-induced ER stress and the unfolded protein response (UPR). Interestingly, NAC almost completely prevented the Cd-induced elevation of C/EBP homologous protein (CHOP) and phosphorylation of c-Jun N-terminal kinase (JNK), two components of the ER stress-mediated apoptotic pathway. In conclusion, NAC protects against Cd-induced germ cell apoptosis by inhibiting endoplasmic reticulum stress in the testes.

  9. High incidence of non-random template strand segregation and asymmetric fate determination in dividing stem cells and their progeny.

    Science.gov (United States)

    Conboy, Michael J; Karasov, Ariela O; Rando, Thomas A

    2007-05-01

    Decades ago, the "immortal strand hypothesis" was proposed as a means by which stem cells might limit acquiring mutations that could give rise to cancer, while continuing to proliferate for the life of an organism. Originally based on observations in embryonic cells, and later studied in terms of stem cell self-renewal, this hypothesis has remained largely unaccepted because of few additional reports, the rarity of the cells displaying template strand segregation, and alternative interpretations of experiments involving single labels or different types of labels to follow template strands. Using sequential pulses of halogenated thymidine analogs (bromodeoxyuridine [BrdU], chlorodeoxyuridine [CldU], and iododeoxyuridine [IdU]), and analyzing stem cell progeny during induced regeneration in vivo, we observed extraordinarily high frequencies of segregation of older and younger template strands during a period of proliferative expansion of muscle stem cells. Furthermore, template strand co-segregation was strongly associated with asymmetric cell divisions yielding daughters with divergent fates. Daughter cells inheriting the older templates retained the more immature phenotype, whereas daughters inheriting the newer templates acquired a more differentiated phenotype. These data provide compelling evidence of template strand co-segregation based on template age and associated with cell fate determination, suggest that template strand age is monitored during stem cell lineage progression, and raise important caveats for the interpretation of label-retaining cells.

  10. High incidence of non-random template strand segregation and asymmetric fate determination in dividing stem cells and their progeny.

    Directory of Open Access Journals (Sweden)

    Michael J Conboy

    2007-05-01

    Full Text Available Decades ago, the "immortal strand hypothesis" was proposed as a means by which stem cells might limit acquiring mutations that could give rise to cancer, while continuing to proliferate for the life of an organism. Originally based on observations in embryonic cells, and later studied in terms of stem cell self-renewal, this hypothesis has remained largely unaccepted because of few additional reports, the rarity of the cells displaying template strand segregation, and alternative interpretations of experiments involving single labels or different types of labels to follow template strands. Using sequential pulses of halogenated thymidine analogs (bromodeoxyuridine [BrdU], chlorodeoxyuridine [CldU], and iododeoxyuridine [IdU], and analyzing stem cell progeny during induced regeneration in vivo, we observed extraordinarily high frequencies of segregation of older and younger template strands during a period of proliferative expansion of muscle stem cells. Furthermore, template strand co-segregation was strongly associated with asymmetric cell divisions yielding daughters with divergent fates. Daughter cells inheriting the older templates retained the more immature phenotype, whereas daughters inheriting the newer templates acquired a more differentiated phenotype. These data provide compelling evidence of template strand co-segregation based on template age and associated with cell fate determination, suggest that template strand age is monitored during stem cell lineage progression, and raise important caveats for the interpretation of label-retaining cells.

  11. Downregulation of miR-205 modulates cell susceptibility to oxidative and endoplasmic reticulum stresses in renal tubular cells.

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    Shiyo Muratsu-Ikeda

    Full Text Available BACKGROUND: Oxidative stress and endoplasmic reticulum (ER stress play a crucial role in tubular damage in both acute kidney injury (AKI and chronic kidney disease (CKD. While the pathophysiological contribution of microRNAs (miRNA to renal damage has also been highlighted, the effect of miRNA on renal damage under oxidative and ER stresses conditions remains elusive. METHODS: We assessed changes in miRNA expression in the cultured renal tubular cell line HK-2 under hypoxia-reoxygenation-induced oxidative stress or ER stress using miRNA microarray assay and real-time RT-PCR. The pathophysiological effect of miRNA was evaluated by cell survival rate, intracellular reactive oxygen species (ROS level, and anti-oxidant enzyme expression in miRNA-inhibited HK-2 or miRNA-overexpressed HK-2 under these stress conditions. The target gene of miRNA was identified by 3'-UTR-luciferase assay. RESULTS: We identified 8 and 10 miRNAs whose expression was significantly altered by oxidative and ER stresses, respectively. Among these, expression of miR-205 was markedly decreased in both stress conditions. Functional analysis revealed that decreased miR-205 led to an increase in cell susceptibility to oxidative and ER stresses, and that this increase was associated with the induction of intracellular ROS and suppression of anti-oxidant enzymes. While increased miR-205 by itself made no change in cell growth or morphology, cell viability under oxidative or ER stress conditions was partially restored. Further, miR-205 bound to the 3'-UTR of the prolyl hydroxylase 1 (PHD1/EGLN2 gene and suppressed the transcription level of EGLN2, which modulates both intracellular ROS level and ER stress state. CONCLUSIONS: miR-205 serves a protective role against both oxidative and ER stresses via the suppression of EGLN2 and subsequent decrease in intracellular ROS. miR-205 may represent a novel therapeutic target in AKI and CKD associated with oxidative or ER stress in tubules.

  12. Oxidative stress induces monocyte necrosis with enrichment of cell-bound albumin and overexpression of endoplasmic reticulum and mitochondrial chaperones.

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    Haiping Tang

    Full Text Available In the present study, monocytes were treated with 5-azacytidine (azacytidine, gossypol or hydrogen peroxide to induce cell death through oxidative stress. A shift from apoptotic to necrotic cell death occurred when monocytes were treated with 100 µM azacytidine for more than 12 hours. Necrotic monocytes exhibited characteristics, including enrichment of cell-bound albumin and up-regulation of endoplasmic reticulum (ER- and mitochondrial-specific chaperones to protect mitochondrial integrity, which were not observed in other necrotic cells, including HUH-7, A2780, A549 and HOC1a. Our results show that the cell-bound albumin originates in the culture medium rather than from monocyte-derived hepatocytes, and that HSP60 is a potential binding partner of the cell-bound albumin. Proteomic analysis shows that HSP60 and protein disulfide isomerase are the most abundant up-regulated mitochondrial and ER-chaperones, and that both HSP60 and calreticulin are ubiquitinated in necrotic monocytes. In contrast, expression levels of the cytosolic chaperones HSP90 and HSP71 were down-regulated in the azacytidine-treated monocytes, concomitant with an increase in the levels of these chaperones in the cell culture medium. Collectively, our results demonstrates that chaperones from different organelles behave differently in necrotic monocytes, ER- and mitochondrial chaperones being retained and cytosolic and nuclear chaperones being released into the cell culture medium through the ruptured cell membrane. HSP60 may serve as a new target for development of myeloid leukemia treatment.

  13. Chromosome Mis-segregation Generates Cell-Cycle-Arrested Cells with Complex Karyotypes that Are Eliminated by the Immune System.

    Science.gov (United States)

    Santaguida, Stefano; Richardson, Amelia; Iyer, Divya Ramalingam; M'Saad, Ons; Zasadil, Lauren; Knouse, Kristin A; Wong, Yao Liang; Rhind, Nicholas; Desai, Arshad; Amon, Angelika

    2017-06-19

    Aneuploidy, a state of karyotype imbalance, is a hallmark of cancer. Changes in chromosome copy number have been proposed to drive disease by modulating the dosage of cancer driver genes and by promoting cancer genome evolution. Given the potential of cells with abnormal karyotypes to become cancerous, do pathways that limit the prevalence of such cells exist? By investigating the immediate consequences of aneuploidy on cell physiology, we identified mechanisms that eliminate aneuploid cells. We find that chromosome mis-segregation leads to further genomic instability that ultimately causes cell-cycle arrest. We further show that cells with complex karyotypes exhibit features of senescence and produce pro-inflammatory signals that promote their clearance by the immune system. We propose that cells with abnormal karyotypes generate a signal for their own elimination that may serve as a means for cancer cell immunosurveillance. Copyright © 2017 Elsevier Inc. All rights reserved.

  14. Endoplasmic reticulum-derived reactive oxygen species (ROS) is involved in toxicity of cell wall stress to Candida albicans.

    Science.gov (United States)

    Yu, Qilin; Zhang, Bing; Li, Jianrong; Zhang, Biao; Wang, Honggang; Li, Mingchun

    2016-10-01

    The cell wall is an important cell structure in both fungi and bacteria, and hence becomes a common antimicrobial target. The cell wall-perturbing agents disrupt synthesis and function of cell wall components, leading to cell wall stress and consequent cell death. However, little is known about the detailed mechanisms by which cell wall stress renders fungal cell death. In this study, we found that ROS scavengers drastically attenuated the antifungal effect of cell wall-perturbing agents to the model fungal pathogen Candida albicans, and these agents caused remarkable ROS accumulation and activation of oxidative stress response (OSR) in this fungus. Interestingly, cell wall stress did not cause mitochondrial dysfunction and elevation of mitochondrial superoxide levels. Furthermore, the iron chelator 2,2'-bipyridyl (BIP) and the hydroxyl radical scavengers could not attenuate cell wall stress-caused growth inhibition and ROS accumulation. However, cell wall stress up-regulated expression of unfold protein response (UPR) genes, enhanced protein secretion and promoted protein folding-related oxidation of Ero1, an important source of ROS production. These results indicated that oxidation of Ero1 in the endoplasmic reticulum (ER), rather than mitochondrial electron transport and Fenton reaction, contributed to cell wall stress-related ROS accumulation and consequent growth inhibition. Our findings uncover a novel link between cell wall integrity (CWI), ER function and ROS production in fungal cells, and shed novel light on development of strategies promoting the antifungal efficacy of cell wall-perturbing agents against fungal infections. Copyright © 2016 Elsevier Inc. All rights reserved.

  15. Endoplasmic reticulum stress and IRE-1 signaling cause apoptosis in colon cancer cells in response to andrographolide treatment.

    Science.gov (United States)

    Banerjee, Aditi; Ahmed, Hafiz; Yang, Peixin; Czinn, Steven J; Blanchard, Thomas G

    2016-07-05

    The plant metabolite andrographolide induces cell cycle arrest and apoptosis in cancer cells. The mechanism(s) by which andrographolide induces apoptosis however, have not been elucidated. The present study was performed to determine the molecular events that promote apoptosis in andrographolide treated cells using T84, HCT116 and COLO 205 colon cancer cell lines. Andrographolide was determined to limit colony formation and Ki67 expression, alter nuclear morphology, increase cytoplasmic histone-associated-DNA-fragments, and increase cleaved caspase-3 levels. Andrographolide also induced significantly higher expression of endoplasmic reticulum (ER) stress proteins GRP-78 and IRE-1 by 48 h but not PERK or ATF6. Apoptosis signaling molecules BAX, spliced XBP-1 and CHOP were also significantly increased. Moreover, chemical inhibition of ER stress or IRE-1 depletion with siRNA in andrographolide treated cells significantly limited expression of IRE-1 and CHOP as determined by immunofluorescence staining, real time PCR, or immunobloting. This was accompanied by a decreased BAX/Bcl-2 ratio. Andrographolide significantly promotes cancer cell death compared to normal cells. These data demonstrate that andrographolide associated ER stress contributes to apoptosis through the activation of a pro-apoptotic GRP-78/IRE-1/XBP-1/CHOP signaling pathway.

  16. Control over the morphology and segregation of Zebrafish germ cell granules during embryonic development

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    Nakkrasae La-Iad

    2008-05-01

    Full Text Available Abstract Background Zebrafish germ cells contain granular-like structures, organized around the cell nucleus. These structures share common features with polar granules in Drosophila, germinal granules in Xenopus and chromatoid bodies in mice germ cells, such as the localization of the zebrafish Vasa, Piwi and Nanos proteins, among others. Little is known about the structure of these granules as well as their segregation in mitosis during early germ-cell development. Results Using transgenic fish expressing a fluorescently labeled novel component of Zebrafish germ cell granules termed Granulito, we followed the morphology and distribution of the granules. We show that whereas these granules initially exhibit a wide size variation, by the end of the first day of development they become a homogeneous population of medium size granules. We investigated this resizing event and demonstrated the role of microtubules and the minus-end microtubule dependent motor protein Dynein in the process. Last, we show that the function of the germ cell granule resident protein the Tudor domain containing protein-7 (Tdrd7 is required for determination of granule morphology and number. Conclusion Our results suggest that Zebrafish germ cell granules undergo a transformation process, which involves germ cell specific proteins as well as the microtubular network.

  17. A novel benzofuran derivative, ACDB, induces apoptosis of human chondrosarcoma cells through mitochondrial dysfunction and endoplasmic reticulum stress.

    Science.gov (United States)

    Su, Chen-Ming; Chen, Chien-Yu; Lu, Tingting; Sun, Yi; Li, Weimin; Huang, Yuan-Li; Tsai, Chun-Hao; Chang, Chih-Shiang; Tang, Chih-Hsin

    2016-12-13

    Chondrosarcoma is one of the bone tumor with high mortality in respond to poor radiation and chemotherapy treatment. Here, we analyze the antitumor activity of a novel benzofuran derivative, 2-amino-3-(2-chlorophenyl)-6-(4-dimethylaminophenyl)benzofuran-4-yl acetate (ACDB), in human chondrosarcoma cells. ACDB increased the cell apoptosis of human chondrosarcomas without harm in chondrocytes. ACDB also enhanced endoplasmic reticulum (ER) stress, which was characterized by varieties in the cytosolic calcium levels and induced the expression of glucose-regulated protein (GRP) and calpain. Furthermore, the ACDB-induced chondrosarcoma apoptosis was associated with the upregulation of the B cell lymphoma-2 (Bcl-2) family members including pro- and anti-apoptotic proteins, downregulation of dysfunctional mitochondria that released cytochrome C, and subsequent activation of caspases-3. In addition, the ACDB-mediated cellular apoptosis was suppressed by transfecting cells with glucose-regulated protein (GRP) and calpain siRNA or treating cells with ER stress chelators and caspase inhibitors. Interestingly, animal experiments illustrated a reduction in the tumor volume following ACDB treatment. Together, these results suggest that ACDB may be a novel tumor suppressor of chondrosarcoma, and this study demonstrates that the novel antitumor agent, ACDB, induced apoptosis by mitochondrial dysfunction and ER stress in human chondrosarcoma cells in vitro and in vivo.

  18. Angiogenic Gene Signature Derived from Subtype Specific Cell Models Segregate Proneural and Mesenchymal Glioblastoma

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    Aman Sharma

    2017-07-01

    Full Text Available Intertumoral molecular heterogeneity in glioblastoma identifies four major subtypes based on expression of molecular markers. Among them, the two clinically interrelated subtypes, proneural and mesenchymal, are the most aggressive with proneural liable for conversion to mesenchymal upon therapy. Using two patient-derived novel primary cell culture models (MTA10 and KW10, we developed a minimal but unique four-gene signature comprising genes vascular endothelial growth factor A (VEGF-A, vascular endothelial growth factor B (VEGF-B and angiopoietin 1 (ANG1, angiopoietin 2 (ANG2 that effectively segregated the proneural (MTA10 and mesenchymal (KW10 glioblastoma subtypes. The cell culture preclassified as mesenchymal showed elevated expression of genes VEGF-A, VEGF-B and ANG1, ANG2 as compared to the other cell culture model that mimicked the proneural subtype. The differentially expressed genes in these two cell culture models were confirmed by us using TCGA and Verhaak databases and we refer to it as a minimal multigene signature (MMS. We validated this MMS on human glioblastoma tissue sections with the use of immunohistochemistry on preclassified (YKL-40 high or mesenchymal glioblastoma and OLIG2 high or proneural glioblastoma tumor samples (n = 30. MMS segregated mesenchymal and proneural subtypes with 83% efficiency using a simple histopathology scoring approach (p = 0.008 for ANG2 and p = 0.01 for ANG1. Furthermore, MMS expression negatively correlated with patient survival. Importantly, MMS staining demonstrated spatiotemporal heterogeneity within each subclass, adding further complexity to subtype identification in glioblastoma. In conclusion, we report a novel and simple sequencing-independent histopathology-based biomarker signature comprising genes VEGF-A, VEGF-B and ANG1, ANG2 for subtyping of proneural and mesenchymal glioblastoma.

  19. The effect of ZnS segregation on Zn-rich CZTS thin film solar cells

    International Nuclear Information System (INIS)

    Li, Wei; Chen, Jian; Yan, Chang; Hao, Xiaojing

    2015-01-01

    Highlights: • Secondary phase segregation in CZTS based solar cells has been studied by TEM. • A “Zn layer exchange” behaviour was found in sulphurisation of Zn/SnCu stacked layers. • XAS reveals a large spike-like CBO (>0.86 eV) between CZTS and ZnS. • Larger ZnS secondary phase proportion increases solar cell’s V oc but limits J sc . - Abstract: Analysis of ZnS segregation behaviour and its influence on the device performance has been made on the Zn-rich Cu 2 ZnSnS 4 thin film solar cells. Cross-sectional transmission electron microscopy images reveal that ZnS is the main secondary phase in the Cu 2 ZnSnS 4 layer obtained from a sulphurised Zn/CuSn metallic stack. The excess Zn diffuses from back contact region to top surface of Cu 2 ZnSnS 4 layer accumulating in the form of ZnS. The solar cell with a higher Zn concentration shows a large quantity of isolated ZnS grains at Cu 2 ZnSnS 4 top surface which is close to CdS/Cu 2 ZnSnS 4 heterojunction interface. Soft X-ray absorption spectroscopy indicates a large spike-like conduction band offset between Cu 2 ZnSnS 4 and ZnS. Consequently, such much ZnS precipitates would increase series resistance and generate lower short-circuit current and external quantum efficiency. However, appropriate amount of ZnS at the space charge region of the solar cell has beneficial effects by reducing the heterojunction interface recombination. Therefore, an improved open-circuit voltage and a higher shunt resistance are achieved. This paper provides a possible method to intentionally segregate ZnS at the space charge region by depositing the Zn layer at the bottom of co-sputtered CuSn layer. Although it is difficult to synthesis a pure phase Cu 2 ZnSnS 4 absorber, we can utilise the ZnS secondary phase to improve the Cu 2 ZnSnS 4 solar performance by controlling the Zn-excess amount

  20. Coupled cell-free synthesis, segregation, and core glycosylation of a secretory protein.

    Science.gov (United States)

    Lingappa, V R; Lingappa, J R; Prasad, R; Ebner, K E; Blobel, G

    1978-05-01

    mRNA from rat mammary glands 13-15 days post partum was translated in a wheat germ cell-free system either in the absence or in the presence of ribosome-denuded membranes prepared from isolated rough microsomes of dog pancreas. Newly synthesized alpha-lactalbumin was identified by immunoprecipitation with a monospecific rabbit antiserum against rat alpha-lactalbumin and was characterized by partial amino-terminal sequence determination and by lectin affinity chromatography. In the absence of membranes a presumably unglycosylated form of alpha-lactalbumin was synthesized that bound neither to concanavalin A-Sepharose nor to Ricinus communis lectin-agarose and that contained an amino-terminal signal peptide region comprising 19 amino acid residues. In the presence of membranes a processed form was synthesized that lacked the signal peptide portion and that had an amino-terminal sequence identical to that of mature alpha-lactalbumin. Furthermore, this processed form was found to be segregated, presumably within the microsomal vesicles, because it was resistant to post-translational proteolysis. It was also found to be glycosylated, and because it bound to concanavalin A-Sepharose, from which it could be eluted specifically by alpha-methyl mannoside, but not to R. communis lectin-agarose, it was presumably core-glycosylated. Processing, segregation, and core glycosylation were observed to proceed only when membranes were present during translation and not when they were added after translation.

  1. Lipocalin 2, a new GADD153 target gene, as an apoptosis inducer of endoplasmic reticulum stress in lung cancer cells

    International Nuclear Information System (INIS)

    Hsin, I-Lun; Hsiao, Yueh-Chieh; Wu, Ming-Fang; Jan, Ming-Shiou; Tang, Sheau-Chung; Lin, Yu-Wen; Hsu, Chung-Ping; Ko, Jiunn-Liang

    2012-01-01

    Endoplasmic reticulum (ER) stress is activated under severe cellular conditions. GADD153, a member of the C/EBP family, is an unfolded protein response (UPR) responsive transcription factor. Increased levels of lipocalin 2, an acute phase protein, have been found in several epithelial cancers. The aim of this study is to investigate the function of lipocalin 2 in lung cancer cells under ER stress. Treatment with thapsigargin, an ER stress activator, led to increases in cytotoxicity, ER stress, apoptosis, and lipocalin 2 expression in A549 cells. GADD153 silencing decreased lipocalin 2 expression in A549 cells. On chromatin immunoprecipitation assay, ER stress increased GADD153 DNA binding to lipocalin 2 promoter. Furthermore, silencing of lipocalin 2 mitigated ER stress-mediated apoptosis in A549 cells. Our findings demonstrated that lipocalin 2 is a new GADD153 target gene that mediates ER stress-induced apoptosis. Highlights: ► We demonstrate that Lipocalin 2 is a new GADD153 target gene. ► Lipocalin 2 mediates ER stress-induced apoptosis. ► ER stress-induced lipocalin 2 expression is calcium-independent in A549 cells. ► Lipocalin 2 dose not play a major role in ER stress-induced autophagy.

  2. Statins Prevent Dextrose-Induced Endoplasmic Reticulum Stress and Oxidative Stress in Endothelial and HepG2 Cells.

    Science.gov (United States)

    Kojanian, Hagop; Szafran-Swietlik, Anna; Onstead-Haas, Luisa M; Haas, Michael J; Mooradian, Arshag D

    Statins have favorable effects on endothelial function partly because of their capacity to reduce oxidative stress. However, antioxidant vitamins, unlike statins, are not as cardioprotective, and this paradox has been explained by failure of vitamin antioxidants to ameliorate endoplasmic reticulum (ER) stress. To determine whether statins prevent dextrose-induced ER stress in addition to their antioxidative effects, human umbilical vein endothelial cells and HepG2 hepatocytes were treated with 27.5 mM dextrose in the presence of simvastatin (lipophilic statin that is a prodrug) and pravastatin (water-soluble active drug), and oxidative stress, ER stress, and cell death were measured. Superoxide generation was measured using 2-methyl-6-(4-methoxyphenyl)-3,7-dihydroimidazo[1,2-A]pyrazin-3-one hydrochloride. ER stress was measured using the placental alkaline phosphatase assay and Western blot of glucose-regulated protein 75, c-jun-N-terminal kinase, phospho-JNK, eukaryotic initiating factor 2α and phospho-eIF2α, and X-box binding protein 1 mRNA splicing. Cell viability was measured by propidium iodide staining. Superoxide anion production, ER stress, and cell death induced by 27.5 mM dextrose were inhibited by therapeutic concentrations of simvastatin and pravastatin. The salutary effects of statins on endothelial cells in reducing both ER stress and oxidative stress observed with pravastatin and the prodrug simvastatin suggest that the effects may be independent of cholesterol-lowering activity.

  3. Violacein induces death of resistant leukaemia cells via kinome reprogramming, endoplasmic reticulum stress and Golgi apparatus collapse.

    Directory of Open Access Journals (Sweden)

    Karla C S Queiroz

    Full Text Available It is now generally recognised that different modes of programmed cell death (PCD are intimately linked to the cancerous process. However, the mechanism of PCD involved in cancer chemoprevention is much less clear and may be different between types of chemopreventive agents and tumour cell types involved. Therefore, from a pharmacological view, it is crucial during the earlier steps of drug development to define the cellular specificity of the candidate as well as its capacity to bypass dysfunctional tumoral signalling pathways providing insensitivity to death stimuli. Studying the cytotoxic effects of violacein, an antibiotic dihydro-indolone synthesised by an Amazon river Chromobacterium, we observed that death induced in CD34(+/c-Kit(+/P-glycoprotein(+/MRP1(+ TF1 leukaemia progenitor cells is not mediated by apoptosis and/or autophagy, since biomarkers of both types of cell death were not significantly affected by this compound. To clarify the working mechanism of violacein, we performed kinome profiling using peptide arrays to yield comprehensive descriptions of cellular kinase activities. Pro-death activity of violacein is actually carried out by inhibition of calpain and DAPK1 and activation of PKA, AKT and PDK, followed by structural changes caused by endoplasmic reticulum stress and Golgi apparatus collapse, leading to cellular demise. Our results demonstrate that violacein induces kinome reprogramming, overcoming death signaling dysfunctions of intrinsically resistant human leukaemia cells.

  4. Calreticulin is a fine tuning molecule in epibrassinolide-induced apoptosis through activating endoplasmic reticulum stress in colon cancer cells.

    Science.gov (United States)

    Obakan-Yerlikaya, Pinar; Arisan, Elif Damla; Coker-Gurkan, Ajda; Adacan, Kaan; Ozbey, Utku; Somuncu, Berna; Baran, Didem; Palavan-Unsal, Narcin

    2017-06-01

    Epibrassinolide (EBR), a member of brassinostreoids plant hormones with cell proliferation promoting role in plants, is a natural polyhydroxysteroid with structural similarity to steroid hormones of vertebrates. EBR has antiproliferative and apoptosis-inducing effect in various cancer cells. Although EBR has been shown to affect survival and mitochondria-mediated apoptosis pathways in a p53-independent manner, the exact molecular targets of EBR are still under investigation. Our recent SILAC (Stable Isotope Labeling by Amino Acids in Cell Culture) data showed that the most significantly altered protein after EBR treatment was calreticulin (CALR). CALR, a chaperone localized in endoplasmic reticulum (ER) lumen, plays role in protein folding and buffering Ca 2+ ions. The alteration of CALR may cause ER stress and unfolded protein response correspondingly the induction of apoptosis. Unfolded proteins are conducted to 26S proteasomal degradation following ubiquitination. Our study revealed that EBR treatment caused ER stress and UPR by altering CALR expression causing caspase-dependent apoptosis in HCT 116, HT29, DLD-1, and SW480 colon cancer cells. Furthermore, 48 h EBR treatment did not caused UPR in Fetal Human Colon cells (FHC) and Mouse Embryonic Fibroblast cells (MEF). In addition our findings showed that HCT 116 colon cancer cells lacking Bax and Puma expression still undergo UPR and related apoptosis. CALR silencing and rapamycin co-treatment prevented EBR-induced UPR and apoptosis, whereas 26S proteasome inhibition further increased the effect of EBR in colon cancer cells. All these findings showed that EBR is an ER stress and apoptotic inducer in colon cancer cells without affecting non-malignant cells. © 2017 Wiley Periodicals, Inc.

  5. Activation of the endoplasmic reticulum stress response by the amyloid-beta 1-40 peptide in brain endothelial cells.

    Science.gov (United States)

    Fonseca, Ana Catarina R G; Ferreiro, Elisabete; Oliveira, Catarina R; Cardoso, Sandra M; Pereira, Cláudia F

    2013-12-01

    Neurovascular dysfunction arising from endothelial cell damage is an early pathogenic event that contributes to the neurodegenerative process occurring in Alzheimer's disease (AD). Since the mechanisms underlying endothelial dysfunction are not fully elucidated, this study was aimed to explore the hypothesis that brain endothelial cell death is induced upon the sustained activation of the endoplasmic reticulum (ER) stress response by amyloid-beta (Aβ) peptide, which deposits in the cerebral vessels in many AD patients and transgenic mice. Incubation of rat brain endothelial cells (RBE4 cell line) with Aβ1-40 increased the levels of several markers of ER stress-induced unfolded protein response (UPR), in a time-dependent manner, and affected the Ca(2+) homeostasis due to the release of Ca(2+) from this intracellular store. Finally, Aβ1-40 was shown to activate both mitochondria-dependent and -independent apoptotic cell death pathways. Enhanced release of cytochrome c from mitochondria and activation of the downstream caspase-9 were observed in cells treated with Aβ1-40 concomitantly with caspase-12 activation. Furthermore, Aβ1-40 activated the apoptosis effectors' caspase-3 and promoted the translocation of apoptosis-inducing factor (AIF) to the nucleus demonstrating the involvement of caspase-dependent and -independent mechanisms during Aβ-induced endothelial cell death. In conclusion, our data demonstrate that ER stress plays a significant role in Aβ1-40-induced apoptotic cell death in brain endothelial cells suggesting that ER stress-targeted therapeutic strategies might be useful in AD to counteract vascular defects and ultimately neurodegeneration. © 2013.

  6. Statolith sedimentation kinetics and force transduction to the cortical endoplasmic reticulum in gravity-sensing Arabidopsis columella cells.

    Science.gov (United States)

    Leitz, Guenther; Kang, Byung-Ho; Schoenwaelder, Monica E A; Staehelin, L Andrew

    2009-03-01

    The starch statolith hypothesis of gravity sensing in plants postulates that the sedimentation of statoliths in specialized statocytes (columella cells) provides the means for converting the gravitational potential energy into a biochemical signal. We have analyzed the sedimentation kinetics of statoliths in the central S2 columella cells of Arabidopsis thaliana. The statoliths can form compact aggregates with gap sizes between statoliths approaching sedimentation phase, the statoliths tend to move at a distance to the cortical endoplasmic reticulum (ER) boundary and interact only transiently with the ER. Statoliths moved by laser tweezers against the ER boundary experience an elastic lift force upon release from the optical trap. High-resolution electron tomography analysis of statolith-to-ER contact sites indicate that the weight of statoliths is sufficient to locally deform the ER membranes that can potentially activate mechanosensitive ion channels. We suggest that in root columella cells, the transduction of the kinetic energy of sedimenting statoliths into a biochemical signal involves a combination of statolith-driven motion of the cytosol, statolith-induced deformation of the ER membranes, and a rapid release of kinetic energy from the ER during reorientation to activate mechanosensitive sites within the central columella cells.

  7. Changes of MODY signal pathway genes in the endoplasmic reticulum stress in INS-1-3 cells.

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    Yanan Dong

    Full Text Available Metabolic disturbances induce endoplasmic reticulum stress (ERS in pancreatic beta cells. This study aims to investigate whether a common pathway exists in the ERS induced by various chemicals, including high levels of glucose and palmitate in INS-1-3 cells.ERS in INS-1-3 cells was induced by exposure cells to thapsigargin (TG, tunicamycin (TM or palmitic acid (PA +high glucose (HG. Digital gene expression (DGE profiling technique was used to detect differentially expressed genes. The profile of gene expression was detected by gene oncology (GO function and pathway enrichment analysis. Nkx6.1 over-expression was established in INS-1-3 cell lines by lentivirus infection to revert the inhibition of Nkx6.1 expression found in the situation of ERS. Real time reverse transcription polymerase chain reaction (RT-PCR was used to verify the expression changes of key genes. Cell viability was measured by 3-(4, 5-dimethylthiazol-2-yl-2, 5-diphenyltetrazolium bromide (MTT assay. The apoptosis was determined by flow cytometry. INS-1-3 cell function was measured by glucose stimulated insulin secretion test(GSIS.As compared to control, DGE demonstrated that there were 135, 57 and 74 differentially expressed genes in TM, TG and HG+PA groups, respectively. Those differentially expressed genes were enriched to ERS, antigen processing and presentation, protein export pathways, and interestingly, the maturity onset diabetes of the young (MODY pathway. Nkx6.1 is one of common down-regulated gene in MODY signaling pathway among TM, TG and HG+PA groups. Over-expression of Nkx6.1 ameliorated glucolipotoxicity induced apoptosis rate by 45.4%, and increased proliferation by 40.9%. At the same time, GSIS increased by 1.82 folds.MODY pathway genes expression was changed in the state of ERS. Over-expression of Nkx6.1 protected the INS-1-3 cells from glucolipotoxicity.

  8. Induction of endoplasmic reticulum stress by deletion of Grp78 depletes Apc mutant intestinal epithelial stem cells.

    Science.gov (United States)

    van Lidth de Jeude, J F; Meijer, B J; Wielenga, M C B; Spaan, C N; Baan, B; Rosekrans, S L; Meisner, S; Shen, Y H; Lee, A S; Paton, J C; Paton, A W; Muncan, V; van den Brink, G R; Heijmans, J

    2017-06-15

    Intestinal epithelial stem cells are highly sensitive to differentiation induced by endoplasmic reticulum (ER) stress. Colorectal cancer develops from mutated intestinal epithelial stem cells. The most frequent initiating mutation occurs in Apc, which results in hyperactivated Wnt signalling. This causes hyperproliferation and reduced sensitivity to chemotherapy, but whether these mutated stem cells are sensitive to ER stress induced differentiation remains unknown. Here we examined this by generating mice in which both Apc and ER stress repressor chaperone Grp78 can be conditionally deleted from the intestinal epithelium. For molecular studies, we used intestinal organoids derived from these mice. Homozygous loss of Apc alone resulted in crypt elongation, activation of the Wnt signature and accumulation of intestinal epithelial stem cells, as expected. This phenotype was however completely rescued on activation of ER stress by additional deletion of Grp78. In these Apc-Grp78 double mutant animals, stem cells were rapidly lost and repopulation occurred by non-mutant cells that had escaped recombination, suggesting that Apc-Grp78 double mutant stem cells had lost self-renewal capacity. Although in Apc-Grp78 double mutant mice the Wnt signature was lost, these intestines exhibited ubiquitous epithelial presence of nuclear β-catenin. This suggests that ER stress interferes with Wnt signalling downstream of nuclear β-catenin. In conclusion, our findings indicate that ER stress signalling results in loss of Apc mutated intestinal epithelial stem cells by interference with the Wnt signature. In contrast to many known inhibitors of Wnt signalling, ER stress acts downstream of β-catenin. Therefore, ER stress poses a promising target in colorectal cancers, which develop as a result of Wnt activating mutations.

  9. Role of prostate apoptosis response 4 in translocation of GRP78 from the endoplasmic reticulum to the cell surface of trophoblastic cells.

    Directory of Open Access Journals (Sweden)

    Marie Cohen

    Full Text Available Glucose-regulated protein 78 (GRP78 is an endoplasmic reticulum (ER molecular chaperone that belongs to the heat shock protein 70 family. GRP78 is also present on the cell surface membrane of trophoblastic cells, where it is associated with invasive or fusion properties of these cells. Impaired mechanism of GRP78 relocation from ER to the cell surface was observed in preeclamptic cytotrophoblastic cells (CTB and could take part in the pathogenesis of preeclampsia. In this study, we have investigated whether prostate apoptosis response 4 (Par-4, a protein identified as a partner of GRP78 relocation to the cell surface in prostate cancer cells, is present in trophoblastic cells and is involved in the translocation of GRP78 to the cell surface of CTB. Par-4 is indeed present in trophoblastic cells and its expression correlates with expression of membrane GRP78. Moreover, overexpression of Par-4 led to an increase of cell surface expression of GRP78 and decreased Par-4 gene expression reduced cell surface localization of GRP78 confirming a role of Par-4 in relocation of GRP78 from ER to the cell surface. Accordingly, invasive property was modified in these cells. In conclusion, we show that Par-4 is expressed in trophoblastic cells and is involved in transport of GRP78 to the cell surface and thus regulates invasive property of extravillous CTB.

  10. Induction of Pro-Apoptotic Endoplasmic Reticulum Stress in Multiple Myeloma Cells by NEO214, Perillyl Alcohol Conjugated to Rolipram.

    Science.gov (United States)

    Chen, Thomas C; Chan, Nymph; Labib, Shirin; Yu, Jiali; Cho, Hee-Yeon; Hofman, Florence M; Schönthal, Axel H

    2018-01-17

    Despite the introduction of new therapies for multiple myeloma (MM), many patients are still dying from this disease and novel treatments are urgently needed. We have designed a novel hybrid molecule, called NEO214, that was generated by covalent conjugation of the natural monoterpene perillyl alcohol (POH), an inducer of endoplasmic reticulum (ER) stress, to rolipram (Rp), an inhibitor of phosphodiesterase-4 (PDE4). Its potential anticancer effects were investigated in a panel of MM cell lines. We found that NEO214 effectively killed MM cells in vitro with a potency that was over an order of magnitude stronger than that of its individual components, either alone or in combination. The cytotoxic mechanism of NEO214 involved severe ER stress and prolonged induction of CCAAT/enhancer-binding protein homologous protein (CHOP), a key pro-apoptotic component of the ER stress response. These effects were prevented by salubrinal, a pharmacologic inhibitor of ER stress, and by CHOP gene knockout. Conversely, combination of NEO214 with bortezomib, a drug in clinical use for patients with MM, resulted in synergistic enhancement of MM cell death. Combination with the adenylate cyclase stimulant forskolin did not enhance NEO214 impact, indicating that cyclic adenosine 3',5'-monophosphate (AMP) pathways might play a lesser role. Our study introduces the novel agent NEO214 as a potent inducer of ER stress with significant anti-MM activity in vitro. It should be further investigated as a potential MM therapy aimed at exploiting this tumor's distinct sensitivity to ER stress.

  11. Copper induces hepatocyte injury due to the endoplasmic reticulum stress in cultured cells and patients with Wilson disease

    International Nuclear Information System (INIS)

    Oe, Shinji; Miyagawa, Koichiro; Honma, Yuichi; Harada, Masaru

    2016-01-01

    Copper is an essential trace element, however, excess copper is harmful to human health. Excess copper-derived oxidants contribute to the progression of Wilson disease, and oxidative stress induces accumulation of abnormal proteins. It is known that the endoplasmic reticulum (ER) plays an important role in proper protein folding, and that accumulation of misfolded proteins disturbs ER homeostasis resulting in ER stress. However, copper-induced ER homeostasis disturbance has not been fully clarified. We treated human hepatoma cell line (Huh7) and immortalized-human hepatocyte cell line (OUMS29) with copper and chemical chaperones, including 4-phenylbutyrate and ursodeoxycholic acid. We examined copper-induced oxidative stress, ER stress and apoptosis by immunofluorescence microscopy and immunoblot analyses. Furthermore, we examined the effects of copper on carcinogenesis. Excess copper induced not only oxidative stress but also ER stress. Furthermore, excess copper induced DNA damage and reduced cell proliferation. Chemical chaperones reduced this copper-induced hepatotoxicity. Excess copper induced hepatotoxicity via ER stress. We also confirmed the abnormality of ultra-structure of the ER of hepatocytes in patients with Wilson disease. These findings show that ER stress plays a pivotal role in Wilson disease, and suggests that chemical chaperones may have beneficial effects in the treatment of Wilson disease.

  12. Copper induces hepatocyte injury due to the endoplasmic reticulum stress in cultured cells and patients with Wilson disease

    Energy Technology Data Exchange (ETDEWEB)

    Oe, Shinji, E-mail: ooes@med.uoeh-u.ac.jp; Miyagawa, Koichiro, E-mail: koichiro@med.uoeh-u.ac.jp; Honma, Yuichi, E-mail: y-homma@med.uoeh-u.ac.jp; Harada, Masaru, E-mail: msrharada@med.uoeh-u.ac.jp

    2016-09-10

    Copper is an essential trace element, however, excess copper is harmful to human health. Excess copper-derived oxidants contribute to the progression of Wilson disease, and oxidative stress induces accumulation of abnormal proteins. It is known that the endoplasmic reticulum (ER) plays an important role in proper protein folding, and that accumulation of misfolded proteins disturbs ER homeostasis resulting in ER stress. However, copper-induced ER homeostasis disturbance has not been fully clarified. We treated human hepatoma cell line (Huh7) and immortalized-human hepatocyte cell line (OUMS29) with copper and chemical chaperones, including 4-phenylbutyrate and ursodeoxycholic acid. We examined copper-induced oxidative stress, ER stress and apoptosis by immunofluorescence microscopy and immunoblot analyses. Furthermore, we examined the effects of copper on carcinogenesis. Excess copper induced not only oxidative stress but also ER stress. Furthermore, excess copper induced DNA damage and reduced cell proliferation. Chemical chaperones reduced this copper-induced hepatotoxicity. Excess copper induced hepatotoxicity via ER stress. We also confirmed the abnormality of ultra-structure of the ER of hepatocytes in patients with Wilson disease. These findings show that ER stress plays a pivotal role in Wilson disease, and suggests that chemical chaperones may have beneficial effects in the treatment of Wilson disease.

  13. Redox signaling via the molecular chaperone BiP protects cells against endoplasmic reticulum-derived oxidative stress

    Science.gov (United States)

    Wang, Jie; Pareja, Kristeen A; Kaiser, Chris A; Sevier, Carolyn S

    2014-01-01

    Oxidative protein folding in the endoplasmic reticulum (ER) has emerged as a potentially significant source of cellular reactive oxygen species (ROS). Recent studies suggest that levels of ROS generated as a byproduct of oxidative folding rival those produced by mitochondrial respiration. Mechanisms that protect cells against oxidant accumulation within the ER have begun to be elucidated yet many questions still remain regarding how cells prevent oxidant-induced damage from ER folding events. Here we report a new role for a central well-characterized player in ER homeostasis as a direct sensor of ER redox imbalance. Specifically we show that a conserved cysteine in the lumenal chaperone BiP is susceptible to oxidation by peroxide, and we demonstrate that oxidation of this conserved cysteine disrupts BiP's ATPase cycle. We propose that alteration of BiP activity upon oxidation helps cells cope with disruption to oxidative folding within the ER during oxidative stress. DOI: http://dx.doi.org/10.7554/eLife.03496.001 PMID:25053742

  14. Beta Blockers Suppress Dextrose-Induced Endoplasmic Reticulum Stress, Oxidative Stress, and Apoptosis in Human Coronary Artery Endothelial Cells.

    Science.gov (United States)

    Haas, Michael J; Kurban, William; Shah, Harshit; Onstead-Haas, Luisa; Mooradian, Arshag D

    Beta blockers are known to have favorable effects on endothelial function partly because of their capacity to reduce oxidative stress. To determine whether beta blockers can also prevent dextrose-induced endoplasmic reticulum (ER) stress in addition to their antioxidative effects, human coronary artery endothelial cells and hepatocyte-derived HepG2 cells were treated with 27.5 mM dextrose for 24 hours in the presence of carvedilol (a lipophilic beta blockers with alpha blocking activity), propranolol (a lipophilic nonselective beta blockers), and atenolol (a water-soluble selective beta blockers), and ER stress, oxidative, stress and cell death were measured. ER stress was measured using the placental alkaline phosphatase assay and Western blot analysis of glucose regulated protein 78, c-Jun-N-terminal kinase (JNK), phospho-JNK, eukaryotic initiating factor 2α (eIF2α), and phospho-eIF2α and measurement of X-box binding protein 1 (XBP1) mRNA splicing using reverse transcriptase-polymerase chain reaction. Superoxide (SO) generation was measured using the superoxide-reactive probe 2-methyl-6-(4-methoxyphenyl)-3,7-dihydroimidazo[1,2-A]pyrazin-3-one hydrochloride (MCLA) chemiluminescence. Cell viability was measured by propidium iodide staining method. The ER stress, SO production, and cell death induced by 27.5 mM dextrose were inhibited by all 3 beta blockers tested. The antioxidative and ER stress reducing effects of beta blockers were also observed in HepG2 cells. The salutary effects of beta blockers on endothelial cells in reducing both ER stress and oxidative stress may contribute to the cardioprotective effects of these agents.

  15. Thiamine deficiency induces endoplasmic reticulum stress and oxidative stress in human neurons derived from induced pluripotent stem cells.

    Science.gov (United States)

    Wang, Xin; Xu, Mei; Frank, Jacqueline A; Ke, Zun-Ji; Luo, Jia

    2017-04-01

    Thiamine (vitamin B1) deficiency (TD) plays a major role in the etiology of Wernicke's encephalopathy (WE) which is a severe neurological disorder. TD induces selective neuronal cell death, neuroinflammation, endoplasmic reticulum (ER) stress and oxidative stress in the brain which are commonly observed in many aging-related neurodegenerative diseases, such as Alzheimer's disease (AD), Parkinson's disease (PD), Huntington's disease (HD) and progressive supranuclear palsy (PSP). However, the underlying cellular and molecular mechanisms remain unclear. The progress in this line of research is hindered due to the lack of appropriate in vitro models. The neurons derived for the human induced pluripotent stem cells (hiPSCs) provide a relevant and powerful tool for the research in pharmaceutical and environmental neurotoxicity. In this study, we for the first time used human induced pluripotent stem cells (hiPSCs)-derived neurons (iCell neurons) to investigate the mechanisms of TD-induced neurodegeneration. We showed that TD caused a concentration- and duration-dependent death of iCell neurons. TD induced ER stress which was evident by the increase in ER stress markers, such as GRP78, XBP-1, CHOP, ATF-6, phosphorylated eIF2α, and cleaved caspase-12. TD also triggered oxidative stress which was shown by the increase in the expression 2,4-dinitrophenyl (DNP) and 4-hydroxynonenal (HNE). ER stress inhibitors (STF-083010 and salubrinal) and antioxidant N-acetyl cysteine (NAC) were effective in alleviating TD-induced death of iCell neurons, supporting the involvement of ER stress and oxidative stress. It establishes that the iCell neurons are a novel tool to investigate cellular and molecular mechanisms for TD-induced neurodegeneration. Copyright © 2017 Elsevier Inc. All rights reserved.

  16. Compound K induced apoptosis via endoplasmic reticulum Ca2+ release through ryanodine receptor in human lung cancer cells

    Directory of Open Access Journals (Sweden)

    Dong-Hyun Shin

    2018-04-01

    Full Text Available Background: Extended endoplasmic reticulum (ER stress may initiate apoptotic pathways in cancer cells, and ER stress has been reported to possibly increase tumor death in cancer therapy. We previously reported that caspase-8 played an important role in compound K-induced apoptosis via activation of caspase-3 directly or indirectly through Bid cleavage, cytochrome c release, and caspase-9 activation in HL-60 human leukemia cells. The mechanisms leading to apoptosis in A549 and SK-MES-1 human lung cancer cells and the role of ER stress have not yet been understood. Methods: The apoptotic effects of compound K were analyzed using flow cytometry, and the changes in protein levels were determined using Western blot analysis. The intracellular calcium levels were monitored by staining with Fura-2/AM and Fluo-3/AM. Results: Compound K-induced ER stress was confirmed through increased phosphorylation of eIF2α and protein levels of GRP78/BiP, XBP-1S, and IRE1α in human lung cancer cells. Moreover, compound-K led to the accumulation of intracellular calcium and an increase in m-calpain activities that were both significantly inhibited by pretreatment either with BAPTA-AM (an intracellular Ca2+ chelator or dantrolene (an RyR channel antagonist. These results were correlated with the outcome that compound K induced ER stress-related apoptosis through caspase-12, as z-ATAD-fmk (a specific inhibitor of caspase-12 partially ameliorated this effect. Interestingly, 4-PBA (ER stress inhibitor dramatically improved the compound K-induced apoptosis. Conclusion: Cell survival and intracellular Ca2+ homeostasis during ER stress in human lung cancer cells are important factors in the induction of the compound K-induced apoptotic pathway. Keywords: apoptosis, calcium, compound K, ER stress, lung cancer cells

  17. The invasion of tobacco mosaic virus RNA induces endoplasmic reticulum stress-related autophagy in HeLa cells

    Science.gov (United States)

    Li, Li; Wang, Li; Xiao, Ruijing; Zhu, Guoguo; Li, Yan; Liu, Changxuan; Yang, Ru; Tang, Zhiqing; Li, Jie; Huang, Wei; Chen, Lang; Zheng, Xiaoling; He, Yuling; Tan, Jinquan

    2011-01-01

    The ability of human cells to defend against viruses originating from distant species has long been ignored. Owing to the pressure of natural evolution and human exploration, some of these viruses may be able to invade human beings. If their ‘fresh’ host had no defences, the viruses could cause a serious pandemic, as seen with HIV, SARS (severe acute respiratory syndrome) and avian influenza virus that originated from chimpanzees, the common palm civet and birds, respectively. It is unknown whether the human immune system could tolerate invasion with a plant virus. To model such an alien virus invasion, we chose TMV (tobacco mosaic virus) and used human epithelial carcinoma cells (HeLa cells) as its ‘fresh’ host. We established a reliable system for transfecting TMV-RNA into HeLa cells and found that TMV-RNA triggered autophagy in HeLa cells as shown by the appearance of autophagic vacuoles, the conversion of LC3-I (light chain protein 3-I) to LC3-II, the up-regulated expression of Beclin1 and the accumulation of TMV protein on autophagosomal membranes. We observed suspected TMV virions in HeLa cells by TEM (transmission electron microscopy). Furthermore, we found that TMV-RNA was translated into CP (coat protein) in the ER (endoplasmic reticulum) and that TMV-positive RNA translocated from the cytoplasm to the nucleolus. Finally, we detected greatly increased expression of GRP78 (78 kDa glucose-regulated protein), a typical marker of ERS (ER stress) and found that the formation of autophagosomes was closely related to the expanded ER membrane. Taken together, our data indicate that HeLa cells used ERS and ERS-related autophagy to defend against TMV-RNA. PMID:21729006

  18. Effect of tauroursodeoxycholic acid on PUFA levels and inflammation in an animal and cell model of hepatic endoplasmic reticulum stress.

    Science.gov (United States)

    Aslan, M; Kıraç, E; Yılmaz, Ö; Ünal, B; Konuk, E K; Özcan, F; Tuzcu, H

    2017-01-01

    The aim of this study was to evaluate hepatic polyunsaturated fatty acids (PUFAs) and inflammatory response in an animal and cell model of endoplasmic reticulum (ER) stress. Rats were divided into control, tunicamycin (TM)-treated, and TM + tauroursodeoxycholic acid (TUDCA)-treated groups. Hepatic ER stress was induced by TM and the ER stress inhibitor TUDCA was injected 30 min before induction of ER stress. Liver THLE-3 cells were treated with TM and TUDCA was administered in advance to decrease cytotoxic effects. Necroinflammation was evaluated in liver sections, while cell viability was determined using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay kit. ER stress was confirmed by immunofluorescence and Western blot analysis of C/EBP-homologous protein and 78-kDa glucose-regulated protein. Arachidonic acid (C20:4n-6), dihomo-γ-linolenic acid (C20:3n-6), eicosapentaenoic acid (C20:5n-3), and docosahexaenoic acid (C22:6n-3) in liver tissue and THLE-3 cells were determined by liquid chromatography tandem mass spectrometry (LC-MS/MS). Phospholipase A2 (PLA2), cyclooxygenase (COX), and prostaglandin E2 (PGE2) were measured in tissue and cell samples. Hepatic ER stress was accomplished by TM and was alleviated by TUDCA. TM treatment significantly decreased PUFAs in both liver and THLE-3 cells compared to controls. PLA2, COX, and PGE2 levels were significantly increased in TM-treated rats and THLE-3 cells compared to controls. TUDCA leads to a partial restoration of liver PUFA levels and decreased PLA2, COX, and PGE2. This study reports decreased PUFA levels in ER stress and supports the use of omega-3 fatty acids in liver diseases demonstrating ER stress.

  19. Mechanisms of mtDNA segregation and mitochondrial signalling in cells with the pathogenic A3243G mutation

    NARCIS (Netherlands)

    Jahangir Tafrechi, Roshan Sakineh

    2008-01-01

    Using newly developed single cell A3243G mutation load assays a novel mechanism of mtDNA segregation was identified in which the multi-copy mtDNA nucleoid takes a central position. Furthermore, likely due to low level changes in gene expression, no genes or gene sets could be identified with gene

  20. Cell membrane thermo-stability studies through joint segregation analysis in various wheat populations

    International Nuclear Information System (INIS)

    Ullah, K.; Khan, N.U.; Gul, S.; Khan, S.J.

    2014-01-01

    Using joint segregation analysis (JSA) technique as statistical approach, mixed inheritance analysis for cell plasma membrane as membrane thermal stability (MTS) was assayed in two parental lines (P1, P2) and their four populations (F1, BC1, BC2, F2) of four wheat crosses, viz., Hashim-08 * LU-26, Farid-06 * Shafaq, Parula * Blue Silver and TD-1 * D-97603 at Faculty of Agriculture, Gomal University, Dera Ismail Khan, Pakistan during crop season 2011-12. Results revealed that MTS was under control of two mixed groups of genes i.e., additive-dominant-epistatic major genes plus additive-dominant-epistasis of polygenes (model E) in Hashim-08 * LU-26 and Farid-06 * Shafaq crosses, respectively. In cross Parula * Blue Silver, it was governed by mixed genes i.e. one major-gene and additive-dominance-epistatic polygenes (model D). However, in cross TD-1 * D-97603, the MTS was under the influence of mixed epistasis of two major genes plus polygenes (model E-1). Polygene variation and polygene heritability were higher than major gene variation and heritability in crosses Hashim-08 * LU-26 and Farid-06 * Shafaq. In crosses Parula * Blue Silver and TD-1 * D-97603, the major gene variation and heritability were higher than polygene variation and heritability, indicating maximum contribution of the major genes. While in cross TD-1 * D-97603, epistatic components were also positive and due to which the polygene heritability was almost zero. Moderate to high environmental variation in the MTS for segregating generations revealed that the said trait was highly persuaded by the environment. However, the genetic behavior of the MTS suggested that early selection for MTS in the crosses Hashim-08 * LU-26 and Farid-06 * Shafaq would be efficient. Whereas, the delayed selection in crosses Parula * Blue Silver and TD-1 * D-97603 until the accumulation of maximum favorable genes will be effective. (author)

  1. Kaempferol induces apoptosis in HepG2 cells via activation of the endoplasmic reticulum stress pathway.

    Science.gov (United States)

    Guo, Haiqing; Ren, Feng; Zhang, Li; Zhang, Xiangying; Yang, Rongrong; Xie, Bangxiang; Li, Zhuo; Hu, Zhongjie; Duan, Zhongping; Zhang, Jing

    2016-03-01

    Kaempferol is a flavonoid compound that has gained importance due to its antitumor properties; however, the underlying mechanisms remain to be fully understood. The present study aimed to investigate the molecular mechanisms of the antitumor function of kaempferol in HepG2 hepatocellular carcinoma cells. Kaempferol was determined to reduce cell viability, increase lactate dehydrogenase activity and induce apoptosis in a concentration‑ and time‑dependent manner in HepG2 cells. Additionally, kaempferol‑induced apoptosis possibly acts via the endoplasmic reticulum (ER) stress pathway, due to the significant increase in the protein expression levels of glucose‑regulated protein 78, glucose‑regulated protein 94, protein kinase R‑like ER kinase, inositol‑requiring enzyme 1α, partial activating transcription factor 6 cleavage, caspase‑4, C/EBP homologous protein (CHOP) and cleaved caspase‑3. The pro‑apoptotic activity of kaempferol was determined to be due to induction of the ER stress‑CHOP pathway, as: i) ER stress was blocked by 4‑phenyl butyric acid (4‑PBA) pretreatment and knockdown of CHOP with small interfering RNA, which resulted in alleviation of kaempferol‑induced HepG2 cell apoptosis; and ii) transfection with plasmid overexpressing CHOP reversed the protective effect of 4‑PBA in kaempferol‑induced HepG2 cells and increased the apoptotic rate. Thus, kaempferol promoted HepG2 cell apoptosis via induction of the ER stress‑CHOP signaling pathway. These observations indicate that kaempferol may be used as a potential chemopreventive treatment strategy for patients with hepatocellular carcinoma.

  2. Monoamine oxidase-dependent endoplasmic reticulum-mitochondria dysfunction and mast cell degranulation lead to adverse cardiac remodeling in diabetes.

    Science.gov (United States)

    Deshwal, Soni; Forkink, Marleen; Hu, Chou-Hui; Buonincontri, Guido; Antonucci, Salvatore; Di Sante, Moises; Murphy, Michael P; Paolocci, Nazareno; Mochly-Rosen, Daria; Krieg, Thomas; Di Lisa, Fabio; Kaludercic, Nina

    2018-02-19

    Monoamine oxidase (MAO) inhibitors ameliorate contractile function in diabetic animals, but the mechanisms remain unknown. Equally elusive is the interplay between the cardiomyocyte alterations induced by hyperglycemia and the accompanying inflammation. Here we show that exposure of primary cardiomyocytes to high glucose and pro-inflammatory stimuli leads to MAO-dependent increase in reactive oxygen species that causes permeability transition pore opening and mitochondrial dysfunction. These events occur upstream of endoplasmic reticulum (ER) stress and are abolished by the MAO inhibitor pargyline, highlighting the role of these flavoenzymes in the ER/mitochondria cross-talk. In vivo, streptozotocin administration to mice induced oxidative changes and ER stress in the heart, events that were abolished by pargyline. Moreover, MAO inhibition prevented both mast cell degranulation and altered collagen deposition, thereby normalizing diastolic function. Taken together, these results elucidate the mechanisms underlying MAO-induced damage in diabetic cardiomyopathy and provide novel evidence for the role of MAOs in inflammation and inter-organelle communication. MAO inhibitors may be considered as a therapeutic option for diabetic complications as well as for other disorders in which mast cell degranulation is a dominant phenomenon.

  3. Non-SMC condensin I complex proteins control chromosome segregation and survival of proliferating cells in the zebrafish neural retina

    Directory of Open Access Journals (Sweden)

    Harris William A

    2009-07-01

    Full Text Available Abstract Background The condensation of chromosomes and correct sister chromatid segregation during cell division is an essential feature of all proliferative cells. Structural maintenance of chromosomes (SMC and non-SMC proteins form the condensin I complex and regulate chromosome condensation and segregation during mitosis. However, due to the lack of appropriate mutants, the function of the condensin I complex during vertebrate development has not been described. Results Here, we report the positional cloning and detailed characterization of retinal phenotypes of a zebrafish mutation at the cap-g locus. High resolution live imaging reveals that the progression of mitosis between prometa- to telophase is delayed and that sister chromatid segregation is impaired upon loss of CAP-G. CAP-G associates with chromosomes between prometa- and telophase of the cell cycle. Loss of the interaction partners CAP-H and CAP-D2 causes cytoplasmic mislocalization of CAP-G throughout mitosis. DNA content analysis reveals increased genomic imbalances upon loss of non-SMC condensin I subunits. Within the retina, loss of condensin I function causes increased rates of apoptosis among cells within the proliferative ciliary marginal zone (CMZ whereas postmitotic retinal cells are viable. Inhibition of p53-mediated apoptosis partially rescues cell numbers in cap-g mutant retinae and allows normal layering of retinal cell types without alleviating their aberrant nuclear sizes. Conclusion Our findings indicate that the condensin I complex is particularly important within rapidly amplifying progenitor cell populations to ensure faithful chromosome segregation. In contrast, differentiation of postmitotic retinal cells is not impaired upon polyploidization.

  4. An isoform of myosin XI is responsible for the translocation of endoplasmic reticulum in tobacco cultured BY-2 cells.

    Science.gov (United States)

    Yokota, Etsuo; Ueda, Shunpei; Tamura, Kentaro; Orii, Hidefumi; Uchi, Satoko; Sonobe, Seiji; Hara-Nishimura, Ikuko; Shimmen, Teruo

    2009-01-01

    The involvement of myosin XI in generating the motive force for cytoplasmic streaming in plant cells is becoming evident. For a comprehensive understanding of the physiological roles of myosin XI isoforms, it is necessary to elucidate the properties and functions of each isoform individually. In tobacco cultured BY-2 cells, two types of myosins, one composed of 175 kDa heavy chain (175 kDa myosin) and the other of 170 kDa heavy chain (170 kDa myosin), have been identified biochemically and immunocytochemically. From sequence analyses of cDNA clones encoding heavy chains of 175 kDa and 170 kDa myosin, both myosins have been classified as myosin XI. Immunocytochemical studies using a polyclonal antibody against purified 175 kDa myosin heavy chain showed that the 175 kDa myosin is distributed throughout the cytoplasm as fine dots in interphase BY-2 cells. During mitosis, some parts of 175 kDa myosin were found to accumulate in the pre-prophase band (PPB), spindle, the equatorial plane of a phragmoplast and on the circumference of daughter nuclei. In transgenic BY-2 cells, in which an endoplasmic reticulum (ER)-specific retention signal, HDEL, tagged with green fluorescent protein (GFP) was stably expressed, ER showed a similar behaviour to that of 175 kDa myosin. Furthermore, this myosin was co-fractionated with GFP-ER by sucrose density gradient centrifugation. From these findings, it was suggested that the 175 kDa myosin is a molecular motor responsible for translocating ER in BY-2 cells.

  5. Sodium 4-phenylbutyrate acts as a chemical chaperone on misfolded myocilin to rescue cells from endoplasmic reticulum stress and apoptosis.

    Science.gov (United States)

    Yam, Gary Hin-Fai; Gaplovska-Kysela, Katarina; Zuber, Christian; Roth, Jürgen

    2007-04-01

    To evaluate the effect of chemical chaperones on the trafficking of secretion-incompetent primary open-angle glaucoma-associated mutant myocilin and the possibility to rescue cells coexpressing mutant and wild-type myocilin from endoplasmic reticulum (ER) stress and apoptosis. CHO-K1, HEK293 and human trabecular meshwork cells were transfected to express wild-type or mutant (C245Y, G364V, P370L, Y437H) myocilin-green fluorescent protein fusion protein and were treated or not with various chemical chaperones (glycerol, dimethylsulfoxide, or sodium 4-phenylbutyrate) for different time periods. The secretion, Triton X-100 solubility, and intracellular distribution of wild-type and mutant myocilin were analyzed by immunoprecipitation, Western blotting, and confocal double immunofluorescence. The effect of sodium 4-phenylbutyrate on ER stress proteins and apoptosis was examined in cells coexpressing mutant and wild-type myocilin. Treatment with sodium 4-phenylbutyrate, but not with glycerol or dimethylsulfoxide, reduced the amount of detergent-insoluble myocilin aggregates, diminished myocilin interaction with calreticulin, and restored the secretion of mutant myocilin. Heteromeric complexes formed by mutant and wild-type myocilin induced the ER stress-associated phosphorylated form of ER-localized eukaryotic initiation factor (eIF)-2alpha kinase and the active form of caspase 3, which resulted in an increased rate of apoptosis. Sodium 4-phenylbutyrate treatment of cells coexpressing mutant and wild-type myocilin relieved ER stress and significantly reduced the rate of apoptosis. These findings indicate that sodium 4-phenylbutyrate protects cells from the deleterious effects of ER-retained aggregated mutant myocilin. These data point to the possibility of a chemical chaperone treatment for myocilin-caused primary open-angle glaucoma.

  6. TRPC1, STIM1, and ORAI influence signal-regulated intracellular and endoplasmic reticulum calcium dynamics in human myometrial cells.

    Science.gov (United States)

    Murtazina, Dilyara A; Chung, Daesuk; Ulloa, Aida; Bryan, Emily; Galan, Henry L; Sanborn, Barbara M

    2011-08-01

    To explore the relationship between signal-stimulated increases in intracellular calcium ([Ca(2+)](i)) and depletion and refilling of the endoplasmic reticulum (ER) Ca(2+) stores ([Ca(2+)](L)) in human myometrial cells, we measured simultaneous changes in [Ca(2+)](i) and [Ca(2+)](L) using Fura-2 and Mag-fluo-4, respectively, in PHM1-41 immortalized and primary cells derived from pregnant myometrium and in primary cells derived from nonpregnant tissue. Signal- and extracellular Ca(2+)-dependent increases in [Ca(2+)](i) (SRCE) and ER refilling stimulated by oxytocin and cyclopiazonic acid were not inhibited by voltage-operated channel blocker nifedipine or mibefradil, inhibition of Na(+)/Ca(2+) exchange with KB-R7943, or zero extracellular Na(+) in PHM1-41 cells. Gadolinium-inhibited oxytocin- and cyclopiazonic acid-induced SRCE and slowed ER store refilling. TRPC1 mRNA knockdown specifically inhibited oxytocin-stimulated SRCE but had no statistically significant effect on ER store refilling and no effect on either parameter following cyclopiazonic acid treatment. Dominant negative STIMΔERM expression attenuated oxytocin- and thapsigargin-stimulated SRCE. Both STIM1 and ORAI1-ORAI3 mRNA knockdowns significantly attenuated oxytocin- and cyclopiazonic acid-stimulated SRCE. The data also suggest that reduction in STIM1 or ORAI1-ORAI3 mRNA can impede the rate of ER store refilling following removal of SERCA inhibition. These data provide evidence for both distinct and overlapping influences of TRPC1, STIM1, and ORAI1-ORAI3 on SRCE and ER store refilling in human myometrial cells that may contribute to the regulation of myometrial Ca(2+) dynamics. These findings have important implications for understanding the control of myometrial Ca(2+) dynamics in relation to myometrial contractile function.

  7. The cell wall and endoplasmic reticulum stress responses are coordinately regulated in Saccharomyces cerevisiae

    OpenAIRE

    Krysan, Damian J

    2009-01-01

    The unfolded protein response (UPR) is an intracellular signaling pathway that regulates the cellular response to the accumulation of misfolded proteins in eukaryotes. Our group has demonstrated that cell wall stress activates UPR in yeast through signals transmitted by the cell wall integrity (CWI) mitogen-activated protein (MAP) kinase cascade. The UPR is required to maintain cell wall integrity; mutants lacking a functional UPR have defects in cell wall biosynthesis and are hypersensitive ...

  8. Resveratrol production in bioreactor: Assessment of cell physiological states and plasmid segregational stability

    Directory of Open Access Journals (Sweden)

    Margarida S. Afonso

    2015-03-01

    Full Text Available Resveratrol is a plant secondary metabolite commonly found in peanuts and grapevines with significant health benefits. Recombinant organisms can produce large amounts of resveratrol and, in this work, Escherichia coli BW27784 was used to produce resveratrol in bioreactors while monitoring cell physiology and plasmid stability through flow cytometry and real-time qPCR, respectively. Initially, the influence of culture conditions and precursor addition was evaluated in screening assays and the data gathered was used to perform the bioreactor assays, allowing the production of 160 μg/mL of resveratrol. Cellular physiology and plasmid instability affected the final resveratrol production, with lower viability and plasmid copy numbers associated with lower yields. In sum, this study describes new tools to monitor the bioprocess, evaluating the effect of culture conditions, and its correlation with cell physiology and plasmid segregational stability, in order to define a viable and scalable bioprocess to fulfill the need for larger quantities of resveratrol.

  9. A CO-FISH assay to assess sister chromatid segregation patterns in mitosis of mouse embryonic stem cells.

    Science.gov (United States)

    Sauer, Stephan; Burkett, Sandra S; Lewandoski, Mark; Klar, Amar J S

    2013-05-01

    Sister chromatids contain identical DNA sequence but are chiral with respect to both their helical handedness and their replication history. Emerging evidence from various model organisms suggests that certain stem cells segregate sister chromatids nonrandomly to either maintain genome integrity or to bias cellular differentiation in asymmetric cell divisions. Conventional methods for tracing of old vs. newly synthesized DNA strands generally lack resolution for individual chromosomes and employ halogenated thymidine analogs with profound cytotoxic effects on rapidly dividing cells. Here, we present a modified chromosome orientation fluorescence in situ hybridization (CO-FISH) assay, where identification of individual chromosomes and their replication history is achieved in subsequent hybridization steps with chromosome-specific DNA probes and PNA telomere probes. Importantly, we tackle the issue of BrdU cytotoxicity and show that our method is compatible with normal mouse ES cell biology, unlike a recently published related protocol. Results from our CO-FISH assay show that mitotic segregation of mouse chromosome 7 is random in ES cells, which contrasts previously published results from our laboratory and settles a controversy. Our straightforward protocol represents a useful resource for future studies on chromatid segregation patterns of in vitro-cultured cells from distinct model organisms.

  10. Endoplasmic Reticulum-Targeted Subunit Toxins Provide a New Approach to Rescue Misfolded Mutant Proteins and Revert Cell Models of Genetic Diseases

    OpenAIRE

    Adnan, Humaira; Zhang, Zhenbo; Park, Hyun-Joo; Tailor, Chetankumar; Che, Clare; Kamani, Mustafa; Spitalny, George; Binnington, Beth; Lingwood, Clifford

    2016-01-01

    Many germ line diseases stem from a relatively minor disturbance in mutant protein endoplasmic reticulum (ER) 3D assembly. Chaperones are recruited which, on failure to correct folding, sort the mutant for retrotranslocation and cytosolic proteasomal degradation (ER-associated degradation-ERAD), to initiate/exacerbate deficiency-disease symptoms. Several bacterial (and plant) subunit toxins, retrograde transport to the ER after initial cell surface receptor binding/internalization. The A subu...

  11. Kaempferol Inhibits Endoplasmic Reticulum Stress-Associated Mucus Hypersecretion in Airway Epithelial Cells And Ovalbumin-Sensitized Mice.

    Science.gov (United States)

    Park, Sin-Hye; Gong, Ju-Hyun; Choi, Yean-Jung; Kang, Min-Kyung; Kim, Yun-Ho; Kang, Young-Hee

    2015-01-01

    Mucus hypersecretion is an important pathological feature of chronic airway diseases, such as asthma and pulmonary diseases. MUC5AC is a major component of the mucus matrix forming family of mucins in the airways. The initiation of endoplasmic reticulum (ER)-mediated stress responses contributes to the pathogenesis of airway diseases. The present study investigated that ER stress was responsible for airway mucus production and this effect was blocked by the flavonoid kaempferol. Oral administration of ≥10 mg/kg kaempferol suppressed mucus secretion and goblet cell hyperplasia observed in the bronchial airway and lung of BALB/c mice sensitized with ovalbumin (OVA). TGF-β and tunicamycin promoted MUC5AC induction after 72 h in human bronchial airway epithelial BEAS-2B cells, which was dampened by 20 μM kaempferol. Kaempferol inhibited tunicamycin-induced ER stress of airway epithelial cells through disturbing the activation of the ER transmembrane sensor ATF6 and IRE1α. Additionally, this compound demoted the induction of ER chaperones such as GRP78 and HSP70 and the splicing of XBP-1 mRNA by tunicamycin. The in vivo study further revealed that kaempferol attenuated the induction of XBP-1 and IRE1α in epithelial tissues of OVA-challenged mice. TGF-β and tunicamycin induced TRAF2 with JNK activation and such induction was deterred by kaempferol. The inhibition of JNK activation encumbered the XBP-1 mRNA splicing and MUC5AC induction by tunicamycin and TGF-β. These results demonstrate that kaempferol alleviated asthmatic mucus hypersecretion through blocking bronchial epithelial ER stress via the inhibition of IRE1α-TRAF2-JNK activation. Therefore, kaempferol may be a potential therapeutic agent targeting mucus hypersecretion-associated pulmonary diseases.

  12. Plasmid segregation mechanisms

    DEFF Research Database (Denmark)

    Ebersbach, G.; Gerdes, Kenn

    2005-01-01

    Bacterial plasmids encode partitioning (par) loci that ensure ordered plasmid segregation prior to cell division. par loci come in two types: those that encode actin-like ATPases and those that encode deviant Walker-type ATPases. ParM, the actin-like ATPase of plasmid R1, forms dynamic filaments...... that segregate plasmids paired at mid-cell to daughter cells. Like microtubules, ParM filaments exhibit dynamic instability (i.e., catastrophic decay) whose regulation is an important component of the DNA segregation process. The Walker box ParA ATPases are related to MinD and form highly dynamic, oscillating...... filaments that are required for the subcellular movement and positioning of plasmids. The role of the observed ATPase oscillation is not yet understood. However, we propose a simple model that couples plasmid segregation to ParA oscillation. The model is consistent with the observed movement...

  13. Myosin XI-dependent formation of tubular structures from endoplasmic reticulum isolated from tobacco cultured BY-2 cells.

    Science.gov (United States)

    Yokota, Etsuo; Ueda, Haruko; Hashimoto, Kohsuke; Orii, Hidefumi; Shimada, Tomoo; Hara-Nishimura, Ikuko; Shimmen, Teruo

    2011-05-01

    The reticular network of the endoplasmic reticulum (ER) consists of tubular and lamellar elements and is arranged in the cortical region of plant cells. This network constantly shows shape change and remodeling motion. Tubular ER structures were formed when GTP was added to the ER vesicles isolated from tobacco (Nicotiana tabacum) cultured BY-2 cells expressing ER-localized green fluorescent protein. The hydrolysis of GTP during ER tubule formation was higher than that under conditions in which ER tubule formation was not induced. Furthermore, a shearing force, such as the flow of liquid, was needed for the elongation/extension of the ER tubule. The shearing force was assumed to correspond to the force generated by the actomyosin system in vivo. To confirm this hypothesis, the S12 fraction was prepared, which contained both cytosol and microsome fractions, including two classes of myosins, XI (175-kD myosin) and VIII (BY-2 myosin VIII-1), and ER-localized green fluorescent protein vesicles. The ER tubules and their mesh-like structures were arranged in the S12 fraction efficiently by the addition of ATP, GTP, and exogenous filamentous actin. The tubule formation was significantly inhibited by the depletion of 175-kD myosin from the S12 fraction but not BY-2 myosin VIII-1. Furthermore, a recombinant carboxyl-terminal tail region of 175-kD myosin also suppressed ER tubule formation. The tips of tubules moved along filamentous actin during tubule elongation. These results indicated that the motive force generated by the actomyosin system contributes to the formation of ER tubules, suggesting that myosin XI is responsible not only for the transport of ER in cytoplasm but also for the reticular organization of cortical ER.

  14. Pentylenetetrazol modulates redox system by inducing addicsin translocation from endoplasmic reticulum to plasma membrane in NG108-15 cells

    Directory of Open Access Journals (Sweden)

    Mitsushi J. Ikemoto

    2017-09-01

    Full Text Available Addicsin (Arl6ip5 is a multifunctional physiological and pathophysiological regulator that exerts its effects by readily forming homo- and hetero-complexes with various functional factors. In particular, addicsin acts as a negative modulator of neural glutamate transporter excitatory amino acid carrier 1 (EAAC1 and participates in the regulation of intracellular glutathione (GSH content by negatively modulating EAAC1-mediated cysteine and glutamate uptake. Addicsin is considered to play a crucial role in the onset of neurodegenerative diseases including epilepsy. However, the molecular dynamics of addicsin remains largely unknown. Here, we report the dynamics of addicsin in NG108-15 cells upon exposure to pentylenetetrazol (PTZ, a representative epileptogenic agent acting on the gamma-Aminobutyric acid A (GABAA receptor. Fluorescent immunostaining analysis demonstrated that addicsin drastically changed its localization from the endoplasmic reticulum (ER to the plasma membrane within 1 h of PTZ exposure in a dose-dependent manner. Moreover, addicsin was co-localized with the plasma membrane markers EAAC1 and Na+/K+ ATPase alpha-3 upon PTZ stimulation. This translocation was significantly inhibited by a non-competitive GABAA receptor antagonist, picrotoxin, but not by a competitive GABAA receptor antagonist, bicuculline. Furthermore, lactate dehydrogenase (LDH assay and 2,2-diphenyl-1-picrylhydrazyl (DPPH radical-scavenging assay showed that PTZ-induced addicsin translocation was accompanied by a decrease of radical-scavenging activity and an increase of cytotoxicity in a PTZ dose-dependent manner. These findings suggest that PTZ induces the translocation of addicsin from the ER to the plasma membrane and modulates the redox system by regulating EAAC1-mediated GSH synthesis, which leads to the activation of cell death signaling.

  15. Brucella suis vaccine strain 2 induces endoplasmic reticulum stress that affects intracellular replication in goat trophoblast cells in vitro

    Directory of Open Access Journals (Sweden)

    Xiangguo eWang

    2016-02-01

    Full Text Available Brucella has been reported to impair placental trophoblasts, a cellular target where Brucella efficiently replicates in association with the endoplasmic reticulum (ER, and ultimately trigger abortion in pregnant animals. However, the precise effects of Brucella on trophoblast cells remain unclear. Here, we describe the infection and replication of Brucella suis vaccine strain 2 (B.suis.S2 in goat trophoblast cells (GTCs and the cellular and molecular responses induced in vitro. Our studies demonstrated that B.suis.S2 was able to infect and proliferate to high titers, hamper the proliferation of GTCs and induce apoptosis due to ER stress. Tunicamycin (Tm, a pharmacological chaperone that strongly mounts ER stress-induced apoptosis, inhibited B.suis.S2 replication in GTCs. In addition, 4 phenyl butyric acid (4-PBA, a pharmacological chaperone that alleviates ER stress-induced apoptosis, significantly enhanced B.suis.S2 replication in GTCs. The Unfolded Protein Response (UPR chaperone molecule GRP78 also promoted B.suis.S2 proliferation in GTCs by inhibiting ER stress-induced apoptosis. We also discovered that the IRE1 pathway, but not the PERK or ATF6 pathway, was activated in the process. However, decreasing the expression of phosphoIRE1α and IRE1α proteins with Irestatin 9389 (IRE1 antagonist in GTCs did not affect the proliferation of B.suis.S2. Although GTC implantation was not affected upon B.suis.S2 infection, progesterone secretion was suppressed, and prolactin and estrogen secretion increased; these effects were accompanied by changes in the expression of genes encoding key steroidogenic enzymes. This study systematically explored the mechanisms of abortion in Brucella infection from the viewpoint of pathogen invasion, ER stress and reproductive endocrinology. Our findings may provide new insight for understanding the mechanisms involved in goat abortions caused by Brucella infection.

  16. Acute Endoplasmic Reticulum Stress-Independent Unconventional Splicing of XBP1 mRNA in the Nucleus of Mammalian Cells

    Directory of Open Access Journals (Sweden)

    Yuanyuan Wang

    2015-06-01

    Full Text Available The regulation of expression of X-box-binding protein-1 (XBP1, a transcriptional factor, involves an unconventional mRNA splicing that removes the 26 nucleotides intron. In contrast to the conventional splicing that exclusively takes place in the nucleus, determining the location of unconventional splicing still remains controversial. This study was designed to examine whether the unconventional spicing of XBP1 mRNA could occur in the nucleus and its possible biological relevance. We use RT-PCR reverse transcription system and the expand high fidelity PCR system to detect spliced XBP1 mRNA, and fraction cells to determine the location of the unconventional splicing of XBP1 mRNA. We employ reporter constructs to show the presence of unconventional splicing machinery in mammal cells independently of acute endoplasmic reticulum (ER stress. Our results reveal the presence of basal unconventional splicing of XBP1 mRNA in the nucleus that also requires inositol-requiring transmembrane kinase and endonuclease 1α (IRE1α and can occur independently of acute ER stress. Furthermore, we confirm that acute ER stress induces the splicing of XBP1 mRNA predominantly occurring in the cytoplasm, but it also promotes the splicing in the nucleus. The deletion of 5′-nucleotides in XBP1 mRNA significantly increases its basal unconventional splicing, suggesting that the secondary structure of XBP1 mRNA may determine the location of unconventional splicing. These results suggest that the unconventional splicing of XBP1 mRNA can take place in the nucleus and/or cytoplasm, which possibly depends on the elaborate regulation. The acute ER stress-independent unconventional splicing in the nucleus is most likely required for the maintaining of day-to-day folding protein homeostasis.

  17. Curcumin induces osteoblast differentiation through mild-endoplasmic reticulum stress-mediated such as BMP2 on osteoblast cells.

    Science.gov (United States)

    Son, Hyo-Eun; Kim, Eun-Jung; Jang, Won-Gu

    2018-01-15

    Curcumin (diferuloylmethane or [1E,6E]-1,7-bis[4-hydroxy-3-methoxyphenyl]-1,6heptadiene-3,5-dione) is a phenolic natural product derived from the rhizomes of the turmeric plant, Curcuma longa. It is reported to have various biological actions such as anti-oxidative, anti-inflammatory, and anti-cancer effects. However, the molecular mechanism of osteoblast differentiation by curcumin has not yet been reported. The cytotoxicity of curcumin was identified using the 3-(4,5-dimethylthiazol-2yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Expression of osteogenic markers and endoplasmic reticulum (ER) stress markers in C3H1-T1/2 cells were measured using reverse-transcriptase polymerase chain reaction (RT-PCR) and Western blotting. Alkaline phosphatase (ALP) staining was performed to assess ALP activity in C3H10T1/2 cells. Transcriptional activity was detected using a luciferase reporter assay. Curcumin increased the expression of genes such as distal-less homeobox 5 (Dlx5), runt-related transcription factor 2 (Runx2), ALP, and osteocalcin (OC), which subsequently induced osteoblast differentiation in C3H10T1/2 cells. In addition, ALP activity and mineralization was found to be increased by curcumin treatment. Curcumin also induced mild ER stress similar to bone morphogenetic protein 2 (BMP2) function in osteoblast cells. Next, we confirmed that curcumin increased mild ER stress and osteoblast differentiation similar to BMP2 in C3H10T1/2 mesenchymal stem cells. Transient transfection studies also showed that curcumin increased ATF6-Luc activity, while decreasing the activities of CREBH-Luc and SMILE-Luc. In addition, similar to BMP2, curcumin induced the phosphorylation of Smad 1/5/9. Overall, these results demonstrate that curcumin-induced mild ER stress increases osteoblast differentiation via ATF6 expression in C3H10T1/2 cells. Copyright © 2017. Published by Elsevier Inc.

  18. Asymmetric segregation and self-renewal of hematopoietic stem and progenitor cells with endocytic Ap2a2.

    Science.gov (United States)

    Ting, Stephen B; Deneault, Eric; Hope, Kristin; Cellot, Sonia; Chagraoui, Jalila; Mayotte, Nadine; Dorn, Jonas F; Laverdure, Jean-Philippe; Harvey, Michael; Hawkins, Edwin D; Russell, Sarah M; Maddox, Paul S; Iscove, Norman N; Sauvageau, Guy

    2012-03-15

    The stem cell-intrinsic model of self-renewal via asymmetric cell division (ACD) posits that fate determinants be partitioned unequally between daughter cells to either activate or suppress the stemness state. ACD is a purported mechanism by which hematopoietic stem cells (HSCs) self-renew, but definitive evidence for this cellular process remains open to conjecture. To address this issue, we chose 73 candidate genes that function within the cell polarity network to identify potential determinants that may concomitantly alter HSC fate while also exhibiting asymmetric segregation at cell division. Initial gene-expression profiles of polarity candidates showed high and differential expression in both HSCs and leukemia stem cells. Altered HSC fate was assessed by our established in vitro to in vivo screen on a subcohort of candidate polarity genes, which revealed 6 novel positive regulators of HSC function: Ap2a2, Gpsm2, Tmod1, Kif3a, Racgap1, and Ccnb1. Interestingly, live-cell videomicroscopy of the endocytic protein AP2A2 shows instances of asymmetric segregation during HSC/progenitor cell cytokinesis. These results contribute further evidence that ACD is functional in HSC self-renewal, suggest a role for Ap2a2 in HSC activity, and provide a unique opportunity to prospectively analyze progeny from HSC asymmetric divisions.

  19. Intracellular insulin-receptor dissociation and segregation in a rat fibroblast cell line transfected with a human insulin receptor gene

    International Nuclear Information System (INIS)

    Levy, J.R.; Olefsky, J.M.

    1988-01-01

    The cellular processing of insulin and insulin receptors was studied using a rat fibroblast cell line that had been transfected with a normal human insulin receptor gene, expressing approximately 500 times the normal number of native fibroblasts insulin receptors. These cells bind and internalize insulin normally. Biochemically assays based on the selective precipitation by polyethylene glycol of intact insulin-receptor complexes but not of free intracellular insulin were developed to study the time course of intracellular insulin-receptor dissociation. Fibroblasts were incubated with radiolabeled insulin at 4 0 C, and internalization of insulin-receptor complexes was initiated by warming the cells to 37 0 C. Within 2 min, 90% of the internalized radioactivity was composed of intact insulin-receptor complexes. The dissociation of insulin from internalized insulin-receptor complexes was markedly inhibited by monensin and chloroquine. Furthermore, chloroquine markedly increased the number of cross-linkable intracellular insulin-receptor complexes, as analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis autoradiography. These findings suggest that acidification of intracellular vesicles is responsible for insulin-receptor dissociation. Physical segregation of dissociated intracellular insulin from its receptor was monitored. The results are consistent with the view that segregation of insulin and receptor occurs 5-10 min after initiation of dissociation. These studies demonstrate the intracellular itinerary of insulin-receptor complexes, including internalization, dissociation of insulin from the internalized receptor within an acidified compartment, segregation of insulin from the receptor, and subsequent ligand degradation

  20. Wise retained in the endoplasmic reticulum inhibits Wnt signaling by reducing cell surface LRP6.

    Science.gov (United States)

    Guidato, Sonia; Itasaki, Nobue

    2007-10-15

    The Wnt signaling pathway is tightly regulated by extracellular and intracellular modulators. Wise was isolated as a secreted protein capable of interacting with the Wnt co-receptor LRP6. Studies in Xenopus embryos revealed that Wise either enhances or inhibits the Wnt pathway depending on the cellular context. Here we show that the cellular localization of Wise has distinct effects on the Wnt pathway readout. While secreted Wise either synergizes or inhibits the Wnt signals depending on the partner ligand, ER-retained Wise consistently blocks the Wnt pathway. ER-retained Wise reduces LRP6 on the cell surface, making cells less susceptible to the Wnt signal. This study provides a cellular mechanism for the action of Wise and introduces the modulation of cellular susceptibility to Wnt signals as a novel mechanism of the regulation of the Wnt pathway.

  1. Kaempferol induces hepatocellular carcinoma cell death via endoplasmic reticulum stress-CHOP-autophagy signaling pathway

    OpenAIRE

    Guo, Haiqing; Lin, Wei; Zhang, Xiangying; Zhang, Xiaohui; Hu, Zhongjie; Li, Liying; Duan, Zhongping; Zhang, Jing; Ren, Feng

    2017-01-01

    Kaempferol is a flavonoid compound that has gained widespread attention due to its antitumor functions. However, the underlying mechanisms are still not clear. The present study investigated the effect of kaempferol on hepatocellular carcinoma and its underlying mechanisms. Kaempferol induced autophagy in a concentration- and time-dependent manner in HepG2 or Huh7 cells, which was evidenced by the significant increase of autophagy-related genes. Inhibition of autophagy pathway, through 3-meth...

  2. Multimodal nanoparticles as alignment and correlation markers in fluorescence/soft X-ray cryo-microscopy/tomography of nucleoplasmic reticulum and apoptosis in mammalian cells

    International Nuclear Information System (INIS)

    Hagen, Christoph; Werner, Stephan; Carregal-Romero, Susana; Malhas, Ashraf N.; Klupp, Barbara G.; Guttmann, Peter; Rehbein, Stefan; Henzler, Katja; Mettenleiter, Thomas C.

    2014-01-01

    Correlative fluorescence and soft X-ray cryo-microscopy/tomography on flat sample holders is perfectly suited to study the uncompromised physiological status of adherent cells at its best possible preservation by imaging after fast cryo-immobilization. To understand the mechanism by which herpesviruses induce nucleoplasmic reticulum, i.e. invaginations of the nuclear envelope, during their egress from the host cell nucleus, morphologically similar structures found in laminopathies and after chemical induction were investigated as a potentially more easily accessible model system. For example, anti-retroviral protease inhibitors like Saquinavir also induce invaginations of the nuclear membranes. With the help of newly designed multimodal nanoparticles as alignment and correlation markers, and by optimizing fluorescence cryo-microscopy data acquisition, an elaborate three-dimensional network of nucleoplasmic reticulum was demonstrated in nuclei of Saquinavir-treated rabbit kidney cells expressing a fluorescently labeled inner nuclear membrane protein. In part of the protease inhibitor-treated samples, nuclei exhibited dramatic ultrastructural changes indicative of programmed cell death/apoptosis. This unexpected observation highlights another unique feature of soft X-ray microscopy, i.e. high absorption contrast information not relying on labeled cellular components, at a 3D resolution of approximately 40 nm (half-pitch) and through a sample thickness of several micrometers. These properties make it a valuable part of the cell biology imaging toolbox to visualize the cellular ultrastructure in its completeness. - Highlights: • Nucleoplasmic reticulum was demonstrated in nuclei of Saquinavir-treated cells. • New polyelectrolyte-Qdot ® 605 coated gold beads were employed as fiducials. • Saquinavir can induce a strong apoptotic phenotype in the nucleus. • CryoXT is an auspicious imaging technique in apoptosis research

  3. The role of meiotic cohesin REC8 in chromosome segregation in {gamma} irradiation-induced endopolyploid tumour cells

    Energy Technology Data Exchange (ETDEWEB)

    Erenpreisa, Jekaterina [Latvian Biomedicine Research and Study Centre, Riga, LV-1067 (Latvia); Cragg, Mark S. [Tenovus Laboratory, Cancer Sciences Division, Southampton University School of Medicine, General Hospital, Southampton SO16 6YD (United Kingdom); Salmina, Kristine [Latvian Biomedicine Research and Study Centre, Riga, LV-1067 (Latvia); Hausmann, Michael [Kirchhoff Inst. fuer Physik, Univ. of Heidelberg, D-69120 Heidelberg (Germany); Scherthan, Harry, E-mail: scherth@web.de [Inst. fuer Radiobiologie der Bundeswehr in Verbindung mit der Univ. Ulm, D-80937 Munich (Germany); MPI for Molec. Genetics, 14195 Berlin (Germany)

    2009-09-10

    Escape from mitotic catastrophe and generation of endopolyploid tumour cells (ETCs) represents a potential survival strategy of tumour cells in response to genotoxic treatments. ETCs that resume the mitotic cell cycle have reduced ploidy and are often resistant to these treatments. In search for a mechanism for genome reduction, we previously observed that ETCs express meiotic proteins among which REC8 (a meiotic cohesin component) is of particular interest, since it favours reductional cell division in meiosis. In the present investigation, we induced endopolyploidy in p53-dysfunctional human tumour cell lines (Namalwa, WI-L2-NS, HeLa) by gamma irradiation, and analysed the sub-cellular localisation of REC8 in the resulting ETCs. We observed by RT-PCR and Western blot that REC8 is constitutively expressed in these tumour cells, along with SGOL1 and SGOL2, and that REC8 becomes modified after irradiation. REC8 localised to paired sister centromeres in ETCs, the former co-segregating to opposite poles. Furthermore, REC8 localised to the centrosome of interphase ETCs and to the astral poles in anaphase cells where it colocalised with the microtubule-associated protein NuMA. Altogether, our observations indicate that radiation-induced ETCs express features of meiotic cell divisions and that these may facilitate chromosome segregation and genome reduction.

  4. The role of meiotic cohesin REC8 in chromosome segregation in γ irradiation-induced endopolyploid tumour cells

    International Nuclear Information System (INIS)

    Erenpreisa, Jekaterina; Cragg, Mark S.; Salmina, Kristine; Hausmann, Michael; Scherthan, Harry

    2009-01-01

    Escape from mitotic catastrophe and generation of endopolyploid tumour cells (ETCs) represents a potential survival strategy of tumour cells in response to genotoxic treatments. ETCs that resume the mitotic cell cycle have reduced ploidy and are often resistant to these treatments. In search for a mechanism for genome reduction, we previously observed that ETCs express meiotic proteins among which REC8 (a meiotic cohesin component) is of particular interest, since it favours reductional cell division in meiosis. In the present investigation, we induced endopolyploidy in p53-dysfunctional human tumour cell lines (Namalwa, WI-L2-NS, HeLa) by gamma irradiation, and analysed the sub-cellular localisation of REC8 in the resulting ETCs. We observed by RT-PCR and Western blot that REC8 is constitutively expressed in these tumour cells, along with SGOL1 and SGOL2, and that REC8 becomes modified after irradiation. REC8 localised to paired sister centromeres in ETCs, the former co-segregating to opposite poles. Furthermore, REC8 localised to the centrosome of interphase ETCs and to the astral poles in anaphase cells where it colocalised with the microtubule-associated protein NuMA. Altogether, our observations indicate that radiation-induced ETCs express features of meiotic cell divisions and that these may facilitate chromosome segregation and genome reduction.

  5. The role of meiotic cohesin REC8 in chromosome segregation in gamma irradiation-induced endopolyploid tumour cells.

    Science.gov (United States)

    Erenpreisa, Jekaterina; Cragg, Mark S; Salmina, Kristine; Hausmann, Michael; Scherthan, Harry

    2009-09-10

    Escape from mitotic catastrophe and generation of endopolyploid tumour cells (ETCs) represents a potential survival strategy of tumour cells in response to genotoxic treatments. ETCs that resume the mitotic cell cycle have reduced ploidy and are often resistant to these treatments. In search for a mechanism for genome reduction, we previously observed that ETCs express meiotic proteins among which REC8 (a meiotic cohesin component) is of particular interest, since it favours reductional cell division in meiosis. In the present investigation, we induced endopolyploidy in p53-dysfunctional human tumour cell lines (Namalwa, WI-L2-NS, HeLa) by gamma irradiation, and analysed the sub-cellular localisation of REC8 in the resulting ETCs. We observed by RT-PCR and Western blot that REC8 is constitutively expressed in these tumour cells, along with SGOL1 and SGOL2, and that REC8 becomes modified after irradiation. REC8 localised to paired sister centromeres in ETCs, the former co-segregating to opposite poles. Furthermore, REC8 localised to the centrosome of interphase ETCs and to the astral poles in anaphase cells where it colocalised with the microtubule-associated protein NuMA. Altogether, our observations indicate that radiation-induced ETCs express features of meiotic cell divisions and that these may facilitate chromosome segregation and genome reduction.

  6. Excessive L-cysteine induces vacuole-like cell death by activating endoplasmic reticulum stress and mitogen-activated protein kinase signaling in intestinal porcine epithelial cells.

    Science.gov (United States)

    Ji, Yun; Wu, Zhenlong; Dai, Zhaolai; Sun, Kaiji; Zhang, Qing; Wu, Guoyao

    2016-01-01

    High intake of dietary cysteine is extremely toxic to animals and the underlying mechanism remains largely unknown. This study was conducted to test the hypothesis that excessive L-cysteine induces cell death by activating endoplasmic reticulum (ER) stress and mitogen-activated protein kinase (MAPK) signaling in intestinal porcine epithelial cells. Jejunal enterocytes were cultured in the presence of 0-10 mmol/L L-cysteine. Cell viability, morphologic alterations, mRNA levels for genes involved in ER stress, protein abundances for glucose-regulated protein 78, C/EBP homologous protein (CHOP), alpha subunit of eukaryotic initiation factor-2 (eIF2α), extracellular signal-regulated kinase (ERK1/2), p38 MAPK, and c-Jun N-terminal protein kinase (JNK1/2) were determined. The results showed that L-cysteine (5-10 mmol/L) reduced cell viability (P L-cysteine were not affected by the autophagy inhibitor 3-methyladenine. The protein abundances for CHOP, phosphorylated (p)-eIF2α, p-JNK1/2, p-p38 MAPK, and the spliced form of XBP-1 mRNA were enhanced (P L-cysteine induces vacuole-like cell death via the activation of ER stress and MAPK signaling in small intestinal epithelial cells. These signaling pathways may be potential targets for developing effective strategies to prevent the toxicity of dietary cysteine.

  7. Silica nanoparticles mediated neuronal cell death in corpus striatum of rat brain: implication of mitochondrial, endoplasmic reticulum and oxidative stress

    Science.gov (United States)

    Parveen, Arshiya; Rizvi, Syed Husain Mustafa; Mahdi, Farzana; Tripathi, Sandeep; Ahmad, Iqbal; Shukla, Rajendra K.; Khanna, Vinay K.; Singh, Ranjana; Patel, Devendra K.; Mahdi, Abbas Ali

    2014-11-01

    Extensive uses of silica nanoparticles (SiNPs) in biomedical and industrial fields have increased the risk of exposure, resulting concerns about their safety. We focussed on some of the safety aspects by studying neurobehavioural impairment, oxidative stress (OS), neurochemical and ultrastructural changes in corpus striatum (CS) of male Wistar rats exposed to 80-nm SiNPs. Moreover, its role in inducing mitochondrial and endoplasmic reticulum (ER) stress-mediated neuronal apoptosis was also investigated. The results demonstrated impairment in neurobehavioural indices, and a significant increase in lipid peroxide levels (LPO), hydrogen peroxide (H2O2), superoxide (O2 -) and protein carbonyl content, whereas there was a significant decrease in the activities of the enzymes, manganese superoxide dismutase (Mn SOD), glutathione peroxidase (GPx), catalase (CAT) and reduced glutathione (GSH) content, suggesting impaired antioxidant defence system. Protein (cytochrome c, Bcl-2, Bax, p53, caspase-3, caspase 12 and CHOP/Gadd153) and mRNA (Bcl-2, Bax, p53 and CHOP/Gadd153, cytochrome c) expression studies of mitochondrial and ER stress-related apoptotic factors suggested that both the cell organelles were involved in OS-mediated apoptosis in treated rat brain CS. Moreover, electron microscopic studies clearly showed mitochondrial and ER dysfunction. In conclusion, the result of the study suggested that subchronic SiNPs' exposure has the potential to alter the behavioural activity and also to bring about changes in biochemical, neurochemical and ultrastructural profiles in CS region of rat brain. Furthermore, we also report SiNPs-induced apoptosis in CS, through mitochondrial and ER stress-mediated signalling.

  8. Activation of PERK signaling through fluoride-mediated endoplasmic reticulum stress in OS732 cells

    International Nuclear Information System (INIS)

    Xu, Hui; Zhou, Yu-lai; Zhang, Xiu-yun; Lu, Peng; Li, Guang-sheng

    2010-01-01

    Our proteomical analysis of osteoblasts exposed to fluoride revealed a distinctive upregulation of proteins in osteoblast. These upregulated proteins play key roles in the protein folding. The PRK-like ER kinase (PERK) signaling, one branch of unfolded protein response (UPR) to combat ER stress, is a transcription factor needed for osteoblast proliferation and differentiation. The mechanism of skeletal fluorosis by which fluoride regulates osteoblast is not fully defined. Here we studied the effect of fluoride on PERK signaling genes and x-box binding protein 1 (xbp-1) in OS7232 cells (human osteoblast-like cell line). Meantime, genes associated with bone turnover were examined in this study. We found that early and continuous fluoride exposure increased the binding immunoglobulin protein (BiP) expression and activated the PERK signaling pathway, resulting in activation of transcription factor 4 (ATF4) and nuclear factor erythroid 2-related factor 2 (Nrf2). The altered expression of cbfa1, osteoprotegerin (OPG)/nuclear factor kappa B ligand (RANKL) were viewed in this study. These results showed fluoride impelled a distinctive ER stress response in OS732 cells, primarily by activating PERK and PERK-dependent signaling. Little effects were viewed for activating xbp-1, a common target of the other two canonical sensors of ER stress, ATF6 and IRE1. In this study the altered expression of bone turnover genes were consistent with activation of ER stress and PERK signaling. This study proved that PERK signaling play major roles in action of fluoride on osteoblast, and suggested that bone response in skeletal fluorosis may be due in part to PERK signaling pathway.

  9. Involvement of endoplasmic reticulum stress in albuminuria induced inflammasome activation in renal proximal tubular cells.

    Directory of Open Access Journals (Sweden)

    Li Fang

    Full Text Available Albuminuria contributes to the progression of tubulointerstitial fibrosis. Although it has been demonstrated that ongoing albuminuria leads to tubular injury manifested by the overexpression of numerous proinflammatory cytokines, the mechanism remains largely unknown. In this study, we found that the inflammasome activation which has been recognized as one of the cornerstones of intracellular surveillance system was associated with the severity of albuminuria in the renal biopsies specimens. In vitro, bovine serum albumin (BSA could also induce the activation of NLRP3 inflammasome in the cultured kidney epithelial cells (NRK-52E. Since there was a significant overlap of NLRP3 with the ER marker calreticulin, the ER stress provoked by BSA seemed to play a crucial role in the activation of inflammasome. Here, we demonstrated that the chemical chaperone taurine-conjugated ursodeoxycholic acid (TUDCA which was proved to be an enhancer for the adaptive capacity of ER could attenuate the inflammasome activation induced by albuminuria not only in vitro but also in diabetic nephropathy. Taken together, these data suggested that ER stress seemed to play an important role in albuminuria-induced inflammasome activation, elimination of ER stress via TUDCA might hold promise as a novel avenue for preventing inflammasome activation ameliorating kidney epithelial cells injury induced by albuminuria.

  10. Function of endoplasmic reticulum calcium ATPase in innate immunity-mediated programmed cell death

    Science.gov (United States)

    Zhu, Xiaohong; Caplan, Jeffrey; Mamillapalli, Padmavathi; Czymmek, Kirk; Dinesh-Kumar, Savithramma P

    2010-01-01

    Programmed cell death (PCD) initiated at the pathogen-infected sites during the plant innate immune response is thought to prevent the development of disease. Here, we describe the identification and characterization of an ER-localized type IIB Ca2+-ATPase (NbCA1) that function as a regulator of PCD. Silencing of NbCA1 accelerates viral immune receptor N- and fungal-immune receptor Cf9-mediated PCD, as well as non-host pathogen Pseudomonas syringae pv. tomato DC3000 and the general elicitor cryptogein-induced cell death. The accelerated PCD rescues loss-of-resistance phenotype of Rar1, HSP90-silenced plants, but not SGT1-silenced plants. Using a genetically encoded calcium sensor, we show that downregulation of NbCA1 results in the modulation of intracellular calcium signalling in response to cryptogein elicitor. We further show that NbCAM1 and NbrbohB function as downstream calcium decoders in N-immune receptor-mediated PCD. Our results indicate that ER-Ca2+-ATPase is a component of the calcium efflux pathway that controls PCD during an innate immune response. PMID:20075858

  11. Inhibition of the sarco/endoplasmic reticulum (ER) Ca2+-ATPase by thapsigargin analogs induces cell death via ER Ca2+ depletion and the unfolded protein response

    DEFF Research Database (Denmark)

    Sehgal, Pankaj; Szalai, Paula; Olesen, Claus

    2017-01-01

    Calcium (Ca2+) is a fundamental regulator of cell signaling and function. Thapsigargin (Tg) blocks the sarco/endoplasmic reticulum (ER) Ca2+-ATPase (SERCA), disrupts Ca2+ homeostasis, and causes cell death. However, the exact mechanisms whereby SERCA-inhibition induces cell death are incompletely...... extensive drainage of the ER Ca2+ stores. This Ca2+ depletion was followed by markedly reduced cell proliferation rates and morphological changes that developed over 2–4 days and culminated in cell death. Interestingly, these changes were not accompanied by bulk increases in cytosolic Ca2+ levels. Moreover...... and their detrimental effects on cell viability. Furthermore, caspase activation and cell death were associated with a sustained unfolded protein response (UPR). We conclude that ER Ca2+ drainage and sustained UPR activation are key for initiation of apoptosis at low concentrations of Tg and Tg analogs, whereas high...

  12. Insulin-like growth factor stimulation increases radiosensitivity of a pancreatic cancer cell line through endoplasmic reticulum stress under hypoxic conditions

    International Nuclear Information System (INIS)

    Isohashi, Fumiaki; Endo, Hiroko; Mukai, Mutsuko; Inoue, Masahiro; Inoue, Takehiro

    2008-01-01

    Tumor hypoxia is an obstacle to radiotherapy. Radiosensitivity under hypoxic conditions is determined by molecular oxygen levels, as well as by various biological cellular responses. The insulin-like growth factor (IGF) signaling pathway is a widely recognized survival signal that confers radioresistance. However, under hypoxic conditions the role of IGF signaling in radiosensitivity is still poorly understood. Here, we demonstrate that IGF-II stimulation decreases clonogenic survival under hypoxic conditions in the pancreatic cancer cell lines AsPC-1 and Panc-1, and in the human breast cancer cell line MCF-7. IGF treatment under hypoxic conditions suppressed increased radiation sensitivity in these cell lines by pharmacologically inhibiting the phosphoinositide 3-kinase-mammalian target of rapamycin pathway, a major IGF signal-transduction pathway. Meanwhile, IGF-II induced the endoplasmic reticulum stress response under hypoxia, including increased protein levels of CHOP and ATF4, mRNA levels of CHOP, GADD34, and BiP as well as splicing levels of XBP-1. The response was suppressed by inhibiting phosphoinositide 3-kinase and mammalian target of rapamycin activity. Overexpression of CHOP in AsPC-1 cells increased radiation sensitivity by IGF-II simulation under hypoxic conditions, whereas suppression of CHOP expression levels with small hairpin RNA or a dominant negative form of a proline-rich extensin-like receptor protein kinase in hypoxia decreased IGF-induced radiosensitivity. IGF-induced endoplasmic reticulum stress contributed to radiosensitization independent of cell cycle status. Taken together, IGF stimulation increased radiosensitivity through the endoplasmic reticulum stress response under hypoxic conditions. (author)

  13. Vildagliptin preserves the mass and function of pancreatic β cells via the developmental regulation and suppression of oxidative and endoplasmic reticulum stress in a mouse model of diabetes

    Science.gov (United States)

    Hamamoto, S; Kanda, Y; Shimoda, M; Tatsumi, F; Kohara, K; Tawaramoto, K; Hashiramoto, M; Kaku, K

    2013-01-01

    Aim We investigated the molecular mechanisms by which vildagliptin preserved pancreatic β cell mass and function. Methods Morphological, biochemical and gene expression profiles of the pancreatic islets were investigated in male KK-Ay-TaJcl(KK-Ay) and C57BL/6JJcl (B6) mice aged 8 weeks which received either vildagliptin or a vehicle for 4 weeks. Results Body weight, food intake, fasting blood glucose, plasma insulin and active glucagon-like peptide-1 were unchanged with vildagliptin treatment in both mice. In KK-Ay mice treated with vildagliptin, increased plasma triglyceride (TG) level and islet TG content were decreased, insulin sensitivity significantly improved, and the glucose tolerance ameliorated with increases in plasma insulin levels. Furthermore, vildagliptin increased glucose-stimulated insulin secretion, islet insulin content and pancreatic β cell mass in both strains. By vildagliptin, the expression of genes involved in cell differentiation/proliferation was upregulated in both strains, those related to apoptosis, endoplasmic reticulum stress and lipid synthesis was decreased and those related to anti-apoptosis and anti-oxidative stress was upregulated, in KK-Ay mice. The morphological results were consistent with the gene expression profiles. Conclusion Vildagliptin increases β cell mass by not only directly affecting cell kinetics but also by indirectly reducing cell apoptosis, oxidative stress and endoplasmic reticulum stress in diabetic mice. PMID:22950702

  14. Fisetin induces apoptosis and endoplasmic reticulum stress in human non-small cell lung cancer through inhibition of the MAPK signaling pathway.

    Science.gov (United States)

    Kang, Kyoung Ah; Piao, Mei Jing; Madduma Hewage, Susara Ruwan Kumara; Ryu, Yea Seong; Oh, Min Chang; Kwon, Taeg Kyu; Chae, Sungwook; Hyun, Jin Won

    2016-07-01

    Fisetin (3,3',4',7-tetrahydroxyflavone), a dietary flavonoid compound, is currently being investigated for its anticancer effect in various cancer models, including lung cancer. Recent studies show that fisetin induces cell growth inhibition and apoptosis in the human non-small cell lung cancer line NCI-H460. In this study, we investigated whether fisetin can induce endoplasmic reticulum (ER) stress-mediated apoptosis in NCI-H460 cells. Fisetin induced mitochondrial reactive oxygen species (ROS) and characteristic signs of ER stress: ER staining; mitochondrial Ca(2+) overload; expression of ER stress-related proteins; glucose-regulated protein (GRP)-78, phosphorylation of protein kinase RNA (PKR)-like endoplasmic reticulum kinase (PERK) and phosphorylation of eukaryotic initiation factor-2 α subunit; cleavage of activating transcription factor-6; phosphorylation of inositol-requiring kinase-1 and splicing of X-box transcription factor-1; induction of C/EBP homologous protein and cleaved caspase-12. siRNA-mediated knockdown of CHOP and ATF-6 attenuated fisetin-induced apoptotic cell death. In addition, fisetin induced phosphorylation of ERK, JNK, and p38 MAPK. Moreover, silencing of the MAPK signaling pathway prevented apoptotic cell death. In summary, our results indicate that, in NCI-H460 cells, fisetin induces apoptosis and ER stress that is mediated by induction of the MAPK signaling pathway.

  15. Cigarette smoke induces endoplasmic reticulum stress and the unfolded protein response in normal and malignant human lung cells

    Directory of Open Access Journals (Sweden)

    Yang Jin

    2008-08-01

    Full Text Available Abstract Background Although lung cancer is among the few malignancies for which we know the primary etiological agent (i.e., cigarette smoke, a precise understanding of the temporal sequence of events that drive tumor progression remains elusive. In addition to finding that cigarette smoke (CS impacts the functioning of key pathways with significant roles in redox homeostasis, xenobiotic detoxification, cell cycle control, and endoplasmic reticulum (ER functioning, our data highlighted a defensive role for the unfolded protein response (UPR program. The UPR promotes cell survival by reducing the accumulation of aberrantly folded proteins through translation arrest, production of chaperone proteins, and increased degradation. Importance of the UPR in maintaining tissue health is evidenced by the fact that a chronic increase in defective protein structures plays a pathogenic role in diabetes, cardiovascular disease, Alzheimer's and Parkinson's syndromes, and cancer. Methods Gene and protein expression changes in CS exposed human cell cultures were monitored by high-density microarrays and Western blot analysis. Tissue arrays containing samples from 110 lung cancers were probed with antibodies to proteins of interest using immunohistochemistry. Results We show that: 1 CS induces ER stress and activates components of the UPR; 2 reactive species in CS that promote oxidative stress are primarily responsible for UPR activation; 3 CS exposure results in increased expression of several genes with significant roles in attenuating oxidative stress; and 4 several major UPR regulators are increased either in expression (i.e., BiP and eIF2α or phosphorylation (i.e., phospho-eIF2α in a majority of human lung cancers. Conclusion These data indicate that chronic ER stress and recruitment of one or more UPR effector arms upon exposure to CS may play a pivotal role in the etiology or progression of lung cancers, and that phospho-eIF2α and BiP may have

  16. The potato virus x TGBp2 protein association with the endoplasmic reticulum plays a role in but is not sufficient for viral cell-to-cell movement

    International Nuclear Information System (INIS)

    Mitra, Ruchira; Krishnamurthy, Konduru; Blancaflor, Elison; Payton, Mark; Nelson, Richard S.; Verchot-Lubicz, Jeanmarie

    2003-01-01

    Potato virus X (PVX) TGBp1, TGBp2, TGBp3, and coat protein are required for virus cell-to-cell movement. Plasmids expressing GFP fused to TGBp2 were bombarded to leaf epidermal cells and GFP:TGBp2 moved cell to cell in Nicotiana benthamiana leaves but not in Nicotiana tabacum leaves. GFP:TGBp2 movement was observed in TGBp1-transgenic N. tabacum, indicating that TGBp2 requires TGBp1 to promote its movement in N. tabacum. In this study, GFP:TGBp2 was detected in a polygonal pattern that resembles the endoplasmic reticulum (ER) network. Amino acid sequence analysis revealed TGBp2 has two putative transmembrane domains. Two mutations separately introduced into the coding sequences encompassing the putative transmembrane domains within the GFP:TGBp2 plasmids and PVX genome, disrupted membrane binding of GFP:TGBp2, inhibited GFP:TGBp2 movement in N. benthamiana and TGBp1-expressing N. tabacum, and inhibited PVX movement. A third mutation, lying outside the transmembrane domains, had no effect on GFP:TGBp2 ER association or movement in N. benthamiana but inhibited GFP:TGBp2 movement in TGBp1-expressing N. tabacum and PVX movement in either Nicotiana species. Thus, ER association of TGBp2 may be required but not be sufficient for virus movement. TGBp2 likely provides an activity for PVX movement beyond ER association

  17. Scanning force microscopy study of phase segregation in fuel cell membrane materials as a function of solvent polarity and relative humidity

    Energy Technology Data Exchange (ETDEWEB)

    Hawley, Marilyn Emily [Los Alamos National Laboratory; Kim, Yu S [Los Alamos National Laboratory; Hjelm, Rex P [Los Alamos National Laboratory

    2010-01-01

    Scanning force microscopy (SFM) phase imaging provides a powerful method for directly studying and comparing phase segregation in fuel cell membrane materials due to different preparation and under different temperature and hwnidity exposures. In this work, we explored two parameters that can influence phase segregation: the properties of the solvents used in casting membrane films and how these solvents alter phase segregation after exposure to boiling water as a function of time. SFM was used under ambient conditions to image phase segregation in Nafion samples prepared using five different solvents. Samples were then subjected to water vapor maintained at 100C for periods ranging from 30 minutes to three hours and re-imaged using the same phase imaging conditions. SFM shows what appears to be an increase in phase segregation as a function of solvent polarity that changes as a function of water exposure.

  18. Initiation of GalNAc-type O-glycosylation in the endoplasmic reticulum promotes cancer cell invasiveness

    DEFF Research Database (Denmark)

    Gill, David J; Tham, Keit Min; Chia, Joanne

    2013-01-01

    Invasiveness underlies cancer aggressiveness and is a hallmark of malignancy. Most malignant tumors have elevated levels of Tn, an O-GalNAc glycan. Mechanisms underlying Tn up-regulation and its effects remain unclear. Here we show that Golgi-to-endoplasmic reticulum relocation of polypeptide N-a...

  19. Targeting cell migration and the endoplasmic reticulum stress response with calmodulin antagonists: a clinically tested small molecule phenocopy of SEC62 gene silencing in human tumor cells

    International Nuclear Information System (INIS)

    Linxweiler, Maximilian; Greiner, Markus; Schorr, Stefan; Schäuble, Nico; Jung, Martin; Linxweiler, Johannes; Langer, Frank; Schäfers, Hans-Joachim; Cavalié, Adolfo; Zimmermann, Richard

    2013-01-01

    Tumor cells benefit from their ability to avoid apoptosis and invade other tissues. The endoplasmic reticulum (ER) membrane protein Sec62 is a key player in these processes. Sec62 is essential for cell migration and protects tumor cells against thapsigargin-induced ER stress, which are both linked to cytosolic Ca 2+ . SEC62 silencing leads to elevated cytosolic Ca 2+ and increased ER Ca 2+ leakage after thapsigargin treatment. Sec62 protein levels are significantly increased in different tumors, including prostate, lung and thyroid cancer. In lung cancer, the influence of Sec62 protein levels on patient survival was analyzed using the Kaplan-Meier method and log-rank test. To elucidate the underlying pathophysiological functions of Sec62, Ca 2+ imaging techniques, real-time cell analysis and cell migration assays were performed. The effects of treatment with the calmodulin antagonists, trifluoperazine (TFP) and ophiobolin A, on cellular Ca 2+ homeostasis, cell growth and cell migration were compared with the effects of siRNA-mediated Sec62 depletion or the expression of a mutated SEC62 variant in vitro. Using Biacore analysis we examined the Ca 2+ -sensitive interaction of Sec62 with the Sec61 complex. Sec62 overproduction significantly correlated with reduced patient survival. Therefore, Sec62 is not only a predictive marker for this type of tumor, but also an interesting therapeutic target. The present study suggests a regulatory function for Sec62 in the major Ca 2+ leakage channel in the ER, Sec61, by a direct and Ca 2+ -sensitive interaction. A Ca 2+ -binding motif in Sec62 is essential for its molecular function. Treatment of cells with calmodulin antagonists mimicked Sec62 depletion by inhibiting cell migration and rendering the cells sensitive to thapsigargin treatment. Targeting tumors that overproduce Sec62 with calmodulin antagonists in combination with targeted thapsigargin analogues may offer novel personalized therapeutic options

  20. Paxillin associates with poly(A)-binding protein 1 at the dense endoplasmic reticulum and the leading edge of migrating cells.

    Science.gov (United States)

    Woods, Alison J; Roberts, Marnie S; Choudhary, Jyoti; Barry, Simon T; Mazaki, Yuichi; Sabe, Hisataka; Morley, Simon J; Critchley, David R; Norman, Jim C

    2002-02-22

    Using mass spectrometry we have identified proteins which co-immunoprecipitate with paxillin, an adaptor protein implicated in the integrin-mediated signaling pathways of cell motility. A major component of paxillin immunoprecipitates was poly(A)-binding protein 1, a 70-kDa mRNA-binding protein. Poly(A)-binding protein 1 associated with both the alpha and beta isoforms of paxillin, and this was unaffected by RNase treatment consistent with a protein-protein interaction. The NH(2)-terminal region of paxillin (residues 54-313) associated directly with poly(A)-binding protein 1 in cell lysates, and with His-poly(A)-binding protein 1 immobilized in microtiter wells. Binding was specific, saturable and of high affinity (K(d) of approximately 10 nm). Cell fractionation studies showed that at steady state, the bulk of paxillin and poly(A)-binding protein 1 was present in the "dense" polyribosome-associated endoplasmic reticulum. However, inhibition of nuclear export with leptomycin B caused paxillin and poly(A)-binding protein 1 to accumulate in the nucleus, indicating that they shuttle between the nuclear and cytoplasmic compartments. When cells migrate, poly(A)-binding protein 1 colocalized with paxillin-beta at the tips of lamellipodia. Our results suggest a new mechanism whereby a paxillin x poly(A)-binding protein 1 complex facilitates transport of mRNA from the nucleus to sites of protein synthesis at the endoplasmic reticulum and the leading lamella during cell migration.

  1. Yeast cells contain a heterogeneous population of peroxisomes that segregate asymmetrically during cell division

    NARCIS (Netherlands)

    Kumar, Sanjeev; de Boer, Rinse; van der Klei, Ida J

    2018-01-01

    Here we used fluorescence microscopy and a peroxisome-targeted tandem fluorescent protein timer to determine the relative age of peroxisomes in yeast. Our data indicate that yeast cells contain a heterogeneous population of relatively old and younger peroxisomes. During budding the peroxisome

  2. Cyclophilin Inhibitors Remodel the Endoplasmic Reticulum of HCV-Infected Cells in a Unique Pattern Rendering Cells Impervious to a Reinfection.

    Directory of Open Access Journals (Sweden)

    Udayan Chatterji

    Full Text Available The mechanisms of action by which cyclophilin inhibitors (CypI interfere with the HCV life cycle remain poorly understood. We reported that CypI and NS5A inhibitors (NS5Ai, but not other classes of anti-HCV agents, prevent assembly of double membrane vesicles (DMVs, which protect replication complexes. We demonstrated that both NS5A and the isomerase cyclophilin A (CypA are required for DMV formation. Here, we examined whether CypI mediate an additional antiviral effect that could further explain the high efficacy of CypI. We identified a unique action of CypI. CypI remodel the organization of the endoplasmic reticulum (ER of HCV-infected cells, but not of uninfected cells. This effect is specific since it was not observed for other classes of anti-HCV agents including NS5Ai, and has no effect on the viability of CypI-treated cells. Since ER serves as platform for the establishment of HCV replication complexes, we asked whether the ER reorganization by CypI would prevent cells from being newly infected. Remarkably, CypI-treated HCV-pre-infected cells remain totally impervious to a reinfection, suggesting that the CypI-mediated ER reorganization prevents a reinfection. This block is not due to residual CypI since CypI-resistant HCV variants also fail to infect these cells. The ER reorganization by CypI is rapid and reversible. This study provides the first evidence that CypI trigger a unique ER reorganization of infected cells, rendering cells transiently impervious to a reinfection. This study further suggests that the HCV-induced ER rearrangement represents a key target for the development of new therapies.

  3. Waste segregation

    International Nuclear Information System (INIS)

    Clark, D.E.; Colombo, P.

    1982-01-01

    A scoping study has been undertaken to determine the state-of-the-art of waste segregation technology as applied to the management of low-level waste (LLW). Present-day waste segregation practices were surveyed through a review of the recent literature and by means of personal interviews with personnel at selected facilities. Among the nuclear establishments surveyed were Department of Energy (DOE) laboratories and plants, nuclear fuel cycle plants, public and private laboratories, institutions, industrial plants, and DOE and commercially operated shallow land burial sites. These survey data were used to analyze the relationship between waste segregation practices and waste treatment/disposal processes, to assess the developmental needs for improved segregation technology, and to evaluate the costs and benefits associated with the implementation of waste segregation controls. This task was planned for completion in FY 1981. It should be noted that LLW management practices are now undergoing rapid change such that the technology and requirements for waste segregation in the near future may differ significantly from those of the present day. 8 figures

  4. Endoplasmic reticulum stress in lung disease

    Directory of Open Access Journals (Sweden)

    Stefan J. Marciniak

    2017-06-01

    Full Text Available Exposure to inhaled pollutants, including fine particulates and cigarette smoke is a major cause of lung disease in Europe. While it is established that inhaled pollutants have devastating effects on the genome, it is now recognised that additional effects on protein folding also drive the development of lung disease. Protein misfolding in the endoplasmic reticulum affects the pathogenesis of many diseases, ranging from pulmonary fibrosis to cancer. It is therefore important to understand how cells respond to endoplasmic reticulum stress and how this affects pulmonary tissues in disease. These insights may offer opportunities to manipulate such endoplasmic reticulum stress pathways and thereby cure lung disease.

  5. Chaetocin induces endoplasmic reticulum stress response and leads to death receptor 5-dependent apoptosis in human non-small cell lung cancer cells.

    Science.gov (United States)

    Liu, Xianfang; Guo, Sen; Liu, Xiangguo; Su, Ling

    2015-11-01

    Epigenetic abnormalities are associated with non-small cell lung cancer (NSCLC) initiation and progression. Epigenetic drugs are being studied and in clinical trials. However, the molecular mechanism underlying the apoptosis by the epigenetic agents remains unclear. SUV39H1 is an important methyl-transferase for lysine 9 on histone H3 and usually related to gene transcriptional suppression, and chaetocin acts as the inhibitor of SUV39H1. We demonstrated here that chaetocin effectively suppressed the growth of multiple lung cancer cells through inducing apoptosis in a death receptor 5 (DR5)-dependent manner. Chaetocin treatment activated endoplasmic reticulum (ER) stress which gave rise to the up-regulation of ATF3 and CHOP. Furthermore, ATF3 and CHOP contributed to the induction of DR5 and subsequent apoptosis. When SUV39H1 was silenced with siRNA, the expression of ATF3, CHOP and DR5 was elevated. Thereafter, knockdown of SUV39H1 induced apoptosis in NSCLC cells. In summary, chaetocin pharmacologically inhibits the activity of SUV39H1 which provokes ER stress and results in up-regulation of ATF3 and CHOP, leading to DR5-dependent apoptosis eventually. These findings provide a novel interpretation on the anti-neoplastic activity of epigenetic drugs as a new therapeutic approach in NSCLC.

  6. Benzo(a)pyrene induced cell cycle arrest and apoptosis in human choriocarcinoma cancer cells through reactive oxygen species-induced endoplasmic reticulum-stress pathway.

    Science.gov (United States)

    Kim, Soo-Min; Lee, Hae-Miru; Hwang, Kyung-A; Choi, Kyung-Chul

    2017-09-01

    Cigarette smoke (CS) contains over 60 well established carcinogens. In this study, we examined the effects of benzo(a)pyrene (B(a)P), a main CS component, on the viability and apoptosis of JEG-3 and BeWo human choriocarcinoma cancer cell lines. An MTT assay confirmed that B(a)P decreased the cell viability of JEG-3 and BeWo cells in a dose-dependent manner. Additionally, Western blot (WB) assay revealed that protein expression of cyclin D and cyclin E decreased, while protein expression of p21 and p27 was increased in response to B(a)P treatment for 48 h. The changes in reactive oxygen species (ROS) levels in JEG-3 and BeWo cells exposed to B(a)P were also measured by a dichlorofluorescein diacetate (DCF-DA) assay, which revealed that ROS levels increased in response to B(a)P treatment for 48 h. WB assay also confirmed that each B(a)P treatment of JEG-3 and BeWo cells for 4 h promoted the expression of phosphorylated eukaryotic initiation factor 2 alpha protein (p-eIF2α) and C/EBP homologous protein (CHOP), which are known to be involved in ROS-mediated endoplasmic reticulum stress (ER-stress) related apoptosis. Overall, the protein expression of Bax (a pro-apoptosis marker) increased, while the expression of Bcl-xl (an anti-apoptotic marker) decreased and the number of apoptotic cells increased in response to B(a)P treatment for 48 h. Taken together, these results suggest that B(a)P has the potential to induce apoptosis of JEG-3 and BeWo human choriocarcinoma cancer cells by increasing the ROS level and simultaneously activating ER-stress. Copyright © 2017 Elsevier Ltd. All rights reserved.

  7. Cantharidin Induced Oral Squamous Cell Carcinoma Cell Apoptosis via the JNK-Regulated Mitochondria and Endoplasmic Reticulum Stress-Related Signaling Pathways.

    Directory of Open Access Journals (Sweden)

    Chin-Chuan Su

    Full Text Available Oral cancer is a subtype of head and neck cancer which represents 2.65% of all human malignancies. Most of oral cancer is histopathologically diagnosed as oral squamous cell carcinoma (OSCC. OSCC is characterized by a high degree of local invasion and a high rate of metastasis to the cervical lymph nodes. How to prevention and treatment of OSCC is important and imperative. Here, we investigated the therapeutic effect and molecular mechanism of cantharidin, an active compound isolated from blister beetles, on OSCC in vitro. Results showed that cantharidin significantly decreased cell viability in human tongue squamous carcinoma-derived SAS, CAL-27, and SCC-4 cell lines. The further mechanistic studies were carried out in SAS cells. Cantharidin also significantly increased apoptosis-related signals, including caspase-9, caspase-7 and caspase-3 proteins. Besides, cantharidin decreased mitochondrial transmembrane potential (MMP and induced cytochrome c and apoptosis inducing factor (AIF release. Cantharidin also increased Bax, Bid, and Bak protein expressions and decreased Bcl-2 protein expression. Cantharidin could also increase the endoplasmic reticulum (ER stress signals, including the expressions of phosphorylated eIF-2α and CHOP, but not Grp78 and Grp94. Furthermore, cantharidin reduced pro-caspase-12 protein expression. In signals of mitogen-activated protein kinases, cantharidin increased the phosphorylation of JNK, but not ERK and p38. Transfection of shRNA-JNK to OSCC cells effectively reversed the cantharidin-induced cell apoptotic signals, including the mitochondrial and ER stress-related signaling molecules. Taken together, these findings suggest that cantharidin induces apoptosis in OSCC cells via the JNK-regulated mitochondria and ER stress-related signaling pathways.

  8. Triglyceride-rich lipoprotein modulates endothelial vascular cell adhesion molecule (VCAM-1 expression via differential regulation of endoplasmic reticulum stress.

    Directory of Open Access Journals (Sweden)

    Ying I Wang

    Full Text Available Circulating triglyceride-rich lipoproteins (TGRL from hypertriglyceridemic subjects exacerbate endothelial inflammation and promote monocyte infiltration into the arterial wall. We have recently reported that TGRL isolated from human blood after a high-fat meal can elicit a pro- or anti-atherogenic state in human aortic endothelial cells (HAEC, defined as up- or down-regulation of VCAM-1 expression in response to tumor necrosis factor alpha (TNFα stimulation, respectively. A direct correlation was found between subjects categorized at higher risk for cardiovascular disease based upon serum triglycerides and postprandial production of TGRL particles that increased VCAM-1-dependent monocyte adhesion to inflamed endothelium. To establish how TGRL metabolism is linked to VCAM-1 regulation, we examined endoplasmic reticulum (ER stress and the unfolded protein response (UPR pathways. Regardless of its atherogenicity, the rate and extent of TGRL internalization and lipid droplet formation by HAEC were uniform. However, pro-atherogenic TGRL exacerbated ER membrane expansion and stress following TNFα stimulation, whereas anti-atherogenic TGRL ameliorated such effects. Inhibition of ER stress with a chemical chaperone 4-phenylbutyric acid decreased TNFα-induced VCAM-1 expression and abrogated TGRL's atherogenic effect. Activation of ER stress sensors PKR-like ER-regulated kinase (PERK and inositol requiring protein 1α (IRE1α, and downstream effectors including eukaryotic initiation factor-2α (eIF2α, spliced X-box-binding protein 1 (sXBP1 and C/EBP homologous protein (CHOP, directly correlated with the atherogenic activity of an individual's TGRL. Modulation of ER stress sensors also correlated with changes in expression of interferon regulatory factor 1 (IRF-1, a transcription factor of Vcam-1 responsible for regulation of its expression. Moreover, knockdown studies using siRNA defined a causal relationship between the PERK/eIF2α/CHOP pathway and

  9. TRB3 reverses chemotherapy resistance and mediates crosstalk between endoplasmic reticulum stress and AKT signaling pathways in MHCC97H human hepatocellular carcinoma cells.

    Science.gov (United States)

    Li, Yang; Zhu, Danxi; Hou, Lidan; Hu, Bin; Xu, Min; Meng, Xiangjun

    2018-01-01

    Tribbles homolog 3 (TRB3), a type of pseudokinase that contains a consensus serine/threonine kinase catalytic core structure, is upregulated in hepatocellular carcinoma. However, the effect of TRB3 expression in hepatocellular carcinoma and the molecular mechanisms underlying TRB3-mediated effects on tumorigenesis in hepatocellular carcinoma have not been fully elucidated. The present study focused on the effect of TRB3 expression in MHCC97H hepatocellular carcinoma cells and investigated the underlying molecular mechanisms in MHCC97H cells. In the present study, it was revealed that TRB3 was significantly overexpressed in the MHCC97H hepatocellular carcinoma cell compared with L-02 normal hepatic cells. Under endoplasmic reticulum (ER) stress induced by thapsigargin and tunicamycin, the levels of TRB3, CCAAT/enhancer binding protein homologous protein (CHOP), protein kinase B (AKT) and phosphorylated (p)AKT expression were upregulated. Furthermore, when the expression of TRB3 was silenced by short hairpin (sh)RNA, the survival of MHCC97H hepatocellular carcinoma cells was increased. Notably, following transduction with lentiviral containing TRB3-shRNA, cell survival also increased after treatment with chemotherapy drug cisplatin. The present study demonstrated that knockdown of CHOP by shRNA was able to reduce TRB3 expression, and the knockdown of TRB3 markedly increased the level of pAKT. TRB3 was overexpressed in MHCC97H hepatocellular carcinoma cells, particularly under endoplasmic reticulum stress. Knockdown of TRB3 was able to increase cell survival. Therefore, TRB3 expression may induce apoptosis and reverse resistance to chemotherapy in MHCC97H hepatic carcinoma cells. The present study suggests that TRB3 is a key molecule that mediates the crosstalk between ER stress and AKT signal pathways. Furthermore, the present study may provide further insight into the cancer biology of hepatocellular carcinoma and the development of anticancer drugs targeting the ER

  10. Cytoskeleton, endoplasmic reticulum and nucleus alterations in CHO-K1 cell line after Crotalus durissus terrificus (South American rattlesnake venom treatment

    Directory of Open Access Journals (Sweden)

    B. P. Tamieti

    2007-01-01

    Full Text Available Snake venoms are toxic to a variety of cell types. However, the intracellular damages and the cell death fate induced by venom are unclear. In the present work, the action of the South American rattlesnake Crotalus durissus terrificus venom on CHO-K1 cell line was analyzed. The cells CHO-K1 were incubated with C. d. terrificus venom (10, 50 and 100g/ml for 1 and 24 hours, and structural alterations of actin filaments, endoplasmic reticulum and nucleus were assessed using specific fluorescent probes and agarose gel electrophoresis for DNA fragmentation. Significant structural changes were observed in all analyzed structures. DNA fragmentation was detected suggesting that, at the concentrations used, the venom induced apoptosis.

  11. ParA and ParB coordinate chromosome segregation with cell elongation and division during Streptomyces sporulation

    Science.gov (United States)

    Donczew, Magdalena; Mackiewicz, Paweł; Wróbel, Agnieszka; Flärdh, Klas; Zakrzewska-Czerwińska, Jolanta

    2016-01-01

    In unicellular bacteria, the ParA and ParB proteins segregate chromosomes and coordinate this process with cell division and chromosome replication. During sporulation of mycelial Streptomyces, ParA and ParB uniformly distribute multiple chromosomes along the filamentous sporogenic hyphal compartment, which then differentiates into a chain of unigenomic spores. However, chromosome segregation must be coordinated with cell elongation and multiple divisions. Here, we addressed the question of whether ParA and ParB are involved in the synchronization of cell-cycle processes during sporulation in Streptomyces. To answer this question, we used time-lapse microscopy, which allows the monitoring of growth and division of single sporogenic hyphae. We showed that sporogenic hyphae stop extending at the time of ParA accumulation and Z-ring formation. We demonstrated that both ParA and ParB affect the rate of hyphal extension. Additionally, we showed that ParA promotes the formation of massive nucleoprotein complexes by ParB. We also showed that FtsZ ring assembly is affected by the ParB protein and/or unsegregated DNA. Our results indicate the existence of a checkpoint between the extension and septation of sporogenic hyphae that involves the ParA and ParB proteins. PMID:27248800

  12. Phosphorylated intermediate of (Ca2+ + K+)-stimulated Mg2+-dependent transport ATPase in endoplasmic reticulum from rat pancreatic acinar cells

    International Nuclear Information System (INIS)

    Imamura, K.; Schulz, I.

    1985-01-01

    Formation and decomposition of the phosphorylated intermediate of endoplasmic reticulum (Ca 2+ + Mg 2+ )-ATPase from pancreatic acinar cells have been studied using lithium dodecyl sulfate- and tetradecyltrimethylammonium bromide-polyacrylamide gel electrophoresis. Incorporation of 32 P from [gamma- 32 P]ATP is Ca 2+ -dependent (approximate Km for free [Ca 2+ ] = 2-3 x 10(-8) mol/liter). Formation of the 100-kDa phosphoprotein is rapid, reaching maximal 32 P incorporation within 1 s at room temperature. At 4 degrees C, phosphorylation is slower and dephosphorylation is drastically decreased. For dephosphorylation, Mg 2+ and monovalent cations such as K + or Na + are necessary. Vanadate inhibits both 32 P incorporation and 32 P liberation dose dependently (Km = 3 x 10(-6) mol/liter), whereas mitochondrial inhibitors and ouabain have no effect. The phosphoprotein is stable at pH 2 and destabilizes with increasing pH being completely decomposed at pH 9. Reduction of 32 P incorporation in the presence of high concentrations of cold ATP and hydroxylamine suggests formation of acylphosphate present in the ATPase intermediate. The characteristics of Ca 2+ , cation, and pH dependencies of the ATPase activity are similar to those previously described for MgATP-dependent Ca 2+ transport into rough endoplasmic reticulum from pancreatic acinar cells. The data suggest that the 100-kDa phosphoprotein as described in this study is the intermediate of this Ca2+ transport ATPase

  13. Three-dimensional organization of a transcellular tubulocisternal endoplasmic reticulum in epithelial cells of Reissner's membrane in the guinea-pig

    DEFF Research Database (Denmark)

    Qvortrup, K; Rostgaard, Jørgen

    1990-01-01

    The ultrastructure of the epithelial cells of Reissner's membrane (membrana vestibularis) in the guinea-pig is described following vascular perfusion with glutaraldehyde of live, anaesthetised and artificially respirated animals. Postfixation in a solution containing OsO4 and potassium ferricyanide...... revealed a well-developed tubulocisternal endoplasmic reticulum, not previously described, the continuity of which has been mapped by serial sectioning and reconstruction. Large disc-shaped subsurface cisternae lining the cell membrane, but separated from it by a space approximately 10 nm wide...... is compared to other techniques used for preservation of Reissner's membrane. Each epithelial cell of Reissner's membrane is endowed with one kinocilium, one to four multivesicular bodies, and a number of intercalated bodies. The functional significance of the canalicular pathway is discussed....

  14. Curcumin enhances the effects of irinotecan on colorectal cancer cells through the generation of reactive oxygen species and activation of the endoplasmic reticulum stress pathway.

    Science.gov (United States)

    Huang, Yan-Feng; Zhu, Da-Jian; Chen, Xiao-Wu; Chen, Qi-Kang; Luo, Zhen-Tao; Liu, Chang-Chun; Wang, Guo-Xin; Zhang, Wei-Jie; Liao, Nv-Zhu

    2017-06-20

    Although initially effective against metastatic colorectal cancer (CRC), irinotecan-based chemotherapy leads to resistance and adverse toxicity. Curcumin is well known for its anti-cancer effects in many cancers, including CRC. Here, we describe reactive oxygen species (ROS) generation and endoplasmic reticulum (ER) stress as important mechanisms by which curcumin enhances irinotecan's effects on CRC cells. CRC cell lines were treated with curcumin and/or irinotecan for 24 h, and then evaluated using cell proliferation assays, cell apoptosis assays, cell cycle analysis, intracellular Ca2+ measurements, ROS measurements and immunoblotting for key ER stress-related proteins. We found that cell viability was inhibited and apoptosis was increased, accompanied by ROS generation and ER stress activation in CRC cells treated with curcumin alone or in combination with irinotecan. Blocking ROS production attenuated the expression of two markers of ER stress: binding of immunoglobulin protein (BIP) and CCAAT/enhancer-binding protein homologous protein (CHOP). Blocking CHOP expression using RNA interference also inhibited ROS generation. These results demonstrated that curcumin could enhance the effects of irinotecan on CRC cells by inhibiting cell viability and inducing cell cycle arrest and apoptosis, and that these effects may be mediated, in part, by ROS generation and activation of the ER stress pathway.

  15. Cetuximab enhances cisplatin-induced endoplasmic reticulum stress-associated apoptosis in laryngeal squamous cell carcinoma cells by inhibiting expression of TXNDC5.

    Science.gov (United States)

    Peng, Fusen; Zhang, Hailin; Du, Youhong; Tan, Pingqing

    2018-03-01

    Cisplatin and cetuximab, an anti‑epidermal growth factor receptor (EGFR) monoclonal humanized antibody, have been used for treatment of laryngeal squamous cell carcinoma (LSCC). It has been demonstrated that cisplatin and inhibition of EGFR signaling may induce endoplasmic reticulum (ER) stress‑associated apoptosis. However, ER protein thioredoxin domain‑containing protein 5 (TXNDC5) reportedly protects cells from ER stress‑associated apoptosis. The present study investigated the interaction between cisplatin, cetuximab and TXNDC5 on ER stress‑associated apoptosis in LSCC cells. AMC‑HN‑8 human LSCC cells with or without TXNDC5 overexpression or knockdown were treated with cisplatin (5, 10, 20 and 40 µM) and/or cetuximab (10, 50, 100 and 150 µg/ml), for 12, 24, 36 and 48 h. Cisplatin and cetuximab concentration‑ and time‑dependently increased and decreased the expression of TXNDC5 in AMC‑HN‑8 cells, respectively. Knockdown of TXNDC5 markedly augmented cisplatin‑induced levels of CCAAT/enhancer‑binding protein homologous protein (CHOP), caspase‑3 activity and apoptosis; while overexpression of TXNDC5 largely eliminated cetuximab‑induced levels of CHOP, caspase‑3 activity and apoptosis. Cisplatin and cetuximab demonstrated a combinatorial effect on increasing the levels of CHOP, caspase‑3 activity and apoptosis, which was largely eliminated by overexpression of TXNDC5 or a reactive oxygen species (ROS) scavenger/antagonist. In addition, promoter/luciferase reporter assays revealed that cisplatin and cetuximab regulated the expression of TXNDC5 at the gene transcription/promoter level. In conclusion, the findings suggested that ER stress‑associated apoptosis is a major mechanism underlying the apoptotic effect of cisplatin and cetuximab on LSCC cells; cetuximab enhanced cisplatin‑induced ER stress‑associated apoptosis in LSCC cells largely by inhibiting the expression of TXNDC5 and thereby increasing ROS production

  16. Enhanced heterologous expression of biologically active human granulocyte colony stimulating factor in transgenic tobacco BY-2 cells by localization to endoplasmic reticulum.

    Science.gov (United States)

    Nair, Nisha R; Chidambareswaren, M; Manjula, S

    2014-09-01

    Tobacco Bright Yellow-2 (BY-2) cells, one of the best characterized cell lines is an attractive expression system for heterologous protein expression. However, the expression of foreign proteins is currently hampered by their low yield, which is partially the result of proteolytic degradation. Human granulocyte colony stimulating factor (hG-CSF) is a hematopoietic cytokine. Recombinant hG-CSF is successfully being used for the treatment of chemotherapy-induced neutropenia in cancer patients. Here, we describe a simple strategy for producing biologically active hG-CSF in tobacco BY-2 cells, localized in the apoplast of BY-2 cells, as well as targeted to the endoplasmic reticulum (ER). ER targeting significantly enhanced recombinant production which scaled to 17.89 mg/l from 4.19 mg/l when expressed in the apoplasts. Southern blotting confirmed the stable integration of hG-CSF in the BY-2 nuclear genome, and the expression of hG-CSF was analysed by Western blotting. Total soluble protein containing hG-CSF isolated from positive calli showed proliferative potential when tested on HL-60 cell lines by MTT assay. We also report the potential of a Fluorescence-activated cell sorting approach for an efficient sorting of the hG-CSF-expressing cell lines, which will enable the generation of homogenous high-producing cell lines.

  17. Monensin, a polyether ionophore antibiotic, overcomes TRAIL resistance in glioma cells via endoplasmic reticulum stress, DR5 upregulation and c-FLIP downregulation.

    Science.gov (United States)

    Yoon, Mi Jin; Kang, You Jung; Kim, In Young; Kim, Eun Hee; Lee, Ju Ahn; Lim, Jun Hee; Kwon, Taeg Kyu; Choi, Kyeong Sook

    2013-08-01

    Tumor necrosis factor-related apoptosis-induced ligand (TRAIL) is preferentially cytotoxic to cancer cells over normal cells. However, many cancer cells, including malignant glioma cells, tend to be resistant to TRAIL. Monensin (a polyether ionophore antibiotic that is widely used in veterinary medicine) and salinomycin (a compound that is structurally related to monensin and shows cancer stem cell-inhibiting activity) are currently recognized as anticancer drug candidates. In this study, we show that monensin effectively sensitizes various glioma cells, but not normal astrocytes, to TRAIL-mediated apoptosis; this occurs at least partly via monensin-induced endoplasmic reticulum (ER) stress, CHOP-mediated DR5 upregulation and proteasome-mediated downregulation of c-FLIP. Interestingly, other polyether antibiotics, such as salinomycin, nigericin, narasin and lasalocid A, also stimulated TRAIL-mediated apoptosis in glioma cells via ER stress, CHOP-mediated DR5 upregulation and c-FLIP downregulation. Taken together, these results suggest that combined treatment of glioma cells with TRAIL and polyether ionophore antibiotics may offer an effective therapeutic strategy.

  18. Dll1 maintains quiescence of adult neural stem cells and segregates asymmetrically during mitosis

    OpenAIRE

    Kawaguchi, Daichi; Furutachi, Shohei; Kawai, Hiroki; Hozumi, Katsuto; Gotoh, Yukiko

    2013-01-01

    Stem cells often divide asymmetrically to produce one stem cell and one differentiating cell, thus maintaining the stem cell pool. Although neural stem cells (NSCs) in the adult mouse subventricular zone have been suggested to divide asymmetrically, intrinsic cell fate determinants for asymmetric NSC division are largely unknown. Stem cell niches are important for stem cell maintenance, but the niche for the maintenance of adult quiescent NSCs has remained obscure. Here we show that the Notch...

  19. Acrolein Disrupts Tight Junction Proteins and Causes Endoplasmic Reticulum Stress-Mediated Epithelial Cell Death Leading to Intestinal Barrier Dysfunction and Permeability.

    Science.gov (United States)

    Chen, Wei-Yang; Wang, Min; Zhang, Jingwen; Barve, Shirish S; McClain, Craig J; Joshi-Barve, Swati

    2017-12-01

    Increasing evidence suggests that environmental and dietary factors can affect intestinal epithelial integrity leading to gut permeability and bacterial translocation. Intestinal barrier dysfunction is a pathogenic process associated with many chronic disorders. Acrolein is an environmental and dietary pollutant and a lipid-derived endogenous metabolite. The impact of acrolein on the intestine has not been investigated before and is evaluated in this study, both in vitro and in vivo. Our data demonstrate that oral acrolein exposure in mice caused damage to the intestinal epithelial barrier, resulting in increased permeability and subsequently translocation of bacterial endotoxin-lipopolysaccharide into the blood. Similar results were seen in vitro using established Caco-2 cell monolayers wherein acrolein decreased barrier function and increased permeability. Acrolein also caused the down-regulation and/or redistribution of three representative tight junction proteins (ie, zonula occludens-1, Occludin, Claudin-1) that critically regulate epithelial paracellular permeability. In addition, acrolein induced endoplasmic reticulum stress-mediated death of epithelial cells, which is an important mechanism contributing to intestinal barrier damage/dysfunction, and gut permeability. Overall, we demonstrate that exposure to acrolein affects the intestinal epithelium by decrease/redistribution of tight junction proteins and endoplasmic reticulum stress-mediated epithelial cell death, thereby resulting in loss of barrier integrity and function. Our findings highlight the adverse consequences of environmental and dietary pollutants on intestinal barrier integrity/function with relevance to gut permeability and the development of disease. Copyright © 2017 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

  20. Ebselen impairs cellular oxidative state and induces endoplasmic reticulum stress and activation of crucial mitogen-activated protein kinases in pancreatic tumour AR42J cells.

    Science.gov (United States)

    Santofimia-Castaño, Patricia; Izquierdo-Alvarez, Alicia; Plaza-Davila, María; Martinez-Ruiz, Antonio; Fernandez-Bermejo, Miguel; Mateos-Rodriguez, Jose M; Salido, Gines M; Gonzalez, Antonio

    2018-01-01

    Ebselen (2-phenyl-1,2-benzisoselenazol-3(2H)-one) is an organoselenium radical scavenger compound, which has strong antioxidant and anti-inflammatory effects. However, evidence suggests that this compound could exert deleterious actions on cell physiology. In this study, we have analyzed the effect of ebselen on rat pancreatic AR42J cells. Cytosolic free-Ca 2+ concentration ([Ca 2+ ] c ), cellular oxidative status, setting of endoplasmic reticulum stress, and phosphorylation of major mitogen-activated protein kinases were analyzed. Our results show that ebselen evoked a concentration-dependent increase in [Ca 2+ ] c . The compound induced an increase in the generation of reactive oxygen species in the mitochondria. We also observed an increase in global cysteine oxidation in the presence of ebselen. In the presence of ebselen an impairment of cholecystokinin-evoked amylase release was noted. Moreover, involvement of the unfolded protein response markers, ER chaperone and signaling regulator GRP78/BiP, eukaryotic translation initiation factor 2α and X-box binding protein 1 was detected. Finally, increases in the phosphorylation of SAPK/JNK, p38 MAPK, and p44/42 MAPK in the presence of ebselen were also observed. Our results provide evidences for an impairment of cellular oxidative state and enzyme secretion, the induction of endoplasmic reticulum stress and the activation of crucial mitogen-activated protein kinases in the presence of ebselen. As a consequence ebselen exerts a potential toxic effect on AR42J cells. © 2017 Wiley Periodicals, Inc.

  1. Location matters: the endoplasmic reticulum and protein trafficking in dendrites

    Directory of Open Access Journals (Sweden)

    Omar A Ramírez

    2011-01-01

    Full Text Available Neurons are highly polarized, but the trafficking mechanisms that operate in these cells and the topological organization of their secretory organelles are still poorly understood. Particularly incipient is our knowledge of the role of the neuronal endoplasmic reticulum. Here we review the current understanding of the endoplasmic reticulum in neurons, its structure, composition, dendritic distribution and dynamics. We also focus on the trafficking of proteins through the dendritic endoplasmic reticulum, emphasizing the relevance of transport, retention, assembly of multi-subunit protein complexes and export. We additionally discuss the roles of the dendritic endoplasmic reticulum in synaptic plasticity.

  2. Tang-Luo-Ning, a Traditional Chinese Medicine, Inhibits Endoplasmic Reticulum Stress-Induced Apoptosis of Schwann Cells under High Glucose Environment

    Directory of Open Access Journals (Sweden)

    Weijie Yao

    2017-01-01

    Full Text Available Tang-Luo-Ning (TLN has a definite effect in the clinical treatment of diabetic peripheral neuropathy (DPN. Schwann cells (SCs apoptosis induced by endoplasmic reticulum stress (ER stress is one of the main pathogeneses of DPN. This study investigates whether TLN can inhibit SCs apoptosis by inhibiting ER stress-induced apoptosis. Our previous researches have demonstrated that TLN could increase the expression of ER stress marker protein GRP78 and inhibited the expression of apoptosis marker protein CHOP in ER stress. In this study, the results showed that TLN attenuated apoptosis by decreasing Ca2+ level in SCs and maintaining ER morphology. TLN could decrease downstream proteins of CHOP including GADD34 and Ero1α, while it increased P-eIF2α and decreased the upstream proteins of CHOP including P-IRE1α/IRE1α and XBP-1, thereby reducing ER stress-induced apoptosis.

  3. Quercetin suppresses DNA double-strand break repair and enhances the radiosensitivity of human ovarian cancer cells via p53-dependent endoplasmic reticulum stress pathway

    Directory of Open Access Journals (Sweden)

    Gong C

    2017-12-01

    Full Text Available Cheng Gong,1 Zongyuan Yang,1 Lingyun Zhang,2 Yuehua Wang,2 Wei Gong,2 Yi Liu3 1Department of Obstetrics and Gynecology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 2Department of Oncology, XiangYang Central Hospital, Hubei University of Arts and Science, XiangYang, 3Department of Medicinal Chemistry, School of Pharmacy, Hubei University of Chinese Medicine, Wuhan, China Abstract: Quercetin is proven to have anticancer effects for many cancers. However, the role of tumor suppressor p53 on quercetin’s radiosensitization and regulation of endoplasmic reticulum (ER stress response in this process remains obscure. Here, quercetin exposure resulted in ER stress, prolonged DNA repair, and the expression of p53 protein; phosphorylation on serine 15 and 20 increased in combination with X-irradiation. Quercetin pretreatment could potentiate radiation-induced cell death. The combination of irradiation and quercetin treatment aggravated DNA damages and caused typical apoptotic cell death; as well the expression of Bax and p21 elevated and the expression of Bcl-2 decreased. Knocking down of p53 could reverse all the above effects under quercetin in combination with radiation. In addition, quercetin-induced radiosensitization was through stimulation of ATM phosphorylation. In human ovarian cancer xenograft model, combined treatment of quercetin and radiation significantly restrained the growth of tumors, accompanied with the activation of p53, CCAAT/enhancer-binding protein homologous protein, and γ-H2AX. Overall, these results indicated that quercetin acted as a promising radiosensitizer through p53-dependent ER stress signals. Keywords: quercetin, p53, endoplasmic reticulum stress, DNA double-strand breaks, eIF-2α (eukaryotic initiation factor 2α, ATM kinase

  4. Lgr5 intestinal stem cells have high telomerase activity and randomly segregate their chromosomes

    NARCIS (Netherlands)

    Schepers, A.G.; Vries, R.G.J.; van den Born, M.M.W.; van de Wetering, M.L.; Clevers, H.

    2011-01-01

    Somatic cells have been proposed to be limited in the number of cell divisions they can undergo. This is thought to be a mechanism by which stem cells retain their integrity preventing disease. However, we have recently discovered intestinal crypt stem cells that persist for the lifetime of a mouse,

  5. Endoplasmic Reticulum-Associated rht-PA Processing in CHO Cells: Influence of Mild Hypothermia and Specific Growth Rates in Batch and Chemostat Cultures.

    Directory of Open Access Journals (Sweden)

    Mauricio Vergara

    Full Text Available Chinese hamster ovary (CHO cells are the main host for producing recombinant proteins with human therapeutic applications mainly because of their capability to perform proper folding and glycosylation processes. In addition, mild hypothermia is one of the main strategies for maximising the productivity of these systems. However, little information is available on the effect of culture temperature on the folding and degradation processes of recombinant proteins that takes place in the endoplasmic reticulum.In order to evaluate the effect of the mild hypothermia on processing/endoplasmatic reticulum-associated degradation (ERAD processes, batch cultures of CHO cells producing recombinant human tissue plasminogen activator (rht-PA were carried out at two temperatures (37°C and 33°C and treated with specific inhibitors of glycosylation and ERAD I (Ubiquitin/Proteasome system or ERAD II (Autophagosoma/Lisosomal system pathways. The effect of mild hypothermia was analysed separately from its indirect effect on specific cell growth rate. To do this, chemostat cultures were carried out at the same incubation conditions as the batch cultures, controlling cell growth at high (0.017 h-1 and low (0.012 h-1 dilution rates. For a better understanding of the investigated phenomenon, cell behaviour was also analysed using principal component analysis (PCA.Results suggest that rht-PA is susceptible to degradation by both ERAD pathways studied, revealing that processing and/or ERAD processes are sensitive to temperature cultivation in batch culture. Moreover, by isolating the effect of culture temperature from the effect of cell growth rate verifyed by using chemostat cultures, we have found that processing and/or ERAD processes are more sensitive to reduction in specific growth rate than low temperature, and that temperature reduction may have a positive effect on protein processing. Interestingly, PCA indicated that the integrated performance displayed by CHO

  6. 7-ketocholesterol induces apoptosis of MC3T3-E1 cells associated with reactive oxygen species generation, endoplasmic reticulum stress and caspase-3/7 dependent pathway

    Directory of Open Access Journals (Sweden)

    Yuta Sato

    2017-03-01

    Full Text Available Type 2 diabetes mellitus (T2DM is associated with an increased risk of bone fractures without reduction of bone mineral density. The cholesterol oxide 7-ketocholesterol (7KCHO has been implicated in numerous diseases such as atherosclerosis, Alzheimer's disease, Parkinson's disease, cancer, age-related macular degeneration and T2DM. In the present study, 7KCHO decreased the viability of MC3T3-E1 cells, increased reactive oxygen species (ROS production and apoptotic rate, and upregulated the caspase-3/7 pathway. Furthermore, these effects of 7KCHO were abolished by pre-incubation of the cells with N-acetylcysteine (NAC, an ROS inhibitor. Also, 7KCHO enhanced the mRNA expression of two endoplasmic reticulum (ER stress markers; CHOP and GRP78, in MC3T3-E1 cells. Pre-incubation of the cells with NAC suppressed the 7KCHO-induced upregulation of CHOP, but not GRP78. In conclusion, we demonstrated that 7KCHO induced apoptosis of MC3T3-E1 cells associated with ROS generation, ER stress, and caspase-3/7 activity, and the effects of 7KCHO were abolished by the ROS inhibitor NAC. These findings may provide new insight into the relationship between oxysterol and pathophysiology of osteoporosis seen in T2DM.

  7. Polysaccharide from Angelica sinensis protects H9c2 cells against oxidative injury and endoplasmic reticulum stress by activating the ATF6 pathway.

    Science.gov (United States)

    Niu, Xiaowei; Zhang, Jingjing; Ling, Chun; Bai, Ming; Peng, Yu; Sun, Shaobo; Li, Yingdong; Zhang, Zheng

    2018-01-01

    Objectives Angelica sinensis exerts various pharmacological effects, such as antioxidant and anti-apoptotic activity. This study aimed to investigate the active ingredients in A. sinensis with antioxidant properties and whether A. sinensis polysaccharide (ASP) protects H9c2 cells against oxidative and endoplasmic reticulum (ER) stress. Methods The ingredients of A. sinensis and their targets and related pathways were determined using web-based databases. Markers of oxidative stress, cell viability, apoptosis, and ER stress-related signalling pathways were measured in H9c2 cells treated with hydrogen peroxide (H 2 O 2 ) and ASP. Results The ingredient-pathway-disease network showed that A. sinensis exerted protective effects against oxidative injury through its various active ingredients on regulation of multiple pathways. Subsequent experiments showed that ASP pretreatment significantly decreased H 2 O 2 -induced cytotoxicity and apoptosis in H9c2 cells. ASP pretreatment inhibited H 2 O 2 -induced reactive oxygen species generation, lactic dehydrogenase release, and malondialdehyde production. ASP exerted beneficial effects by inducing activating transcription factor 6 (ATF6) and increasing ATF6 target protein levels, which in turn attenuated ER stress and increased antioxidant activity. Conclusions Our findings indicate that ASP, a major water-soluble component of A. sinensis, exerts protective effects against H 2 O 2 -induced injury in H9c2 cells by activating the ATF6 pathway, thus ameliorating ER and oxidative stress.

  8. Cell adhesive affinity does not dictate primitive endoderm segregation and positioning during murine embryoid body formation.

    Science.gov (United States)

    Moore, Robert; Cai, Kathy Q; Escudero, Diogo O; Xu, Xiang-Xi

    2009-09-01

    The classical cell sorting experiments undertaken by Townes and Holtfreter described the intrinsic propensity of dissociated embryonic cells to self-organize and reconcile into their original embryonic germ layers with characteristic histotypic positioning. Steinberg presented the differential adhesion hypothesis to explain these patterning phenomena. Here, we have reappraised these issues by implementing embryoid bodies to model the patterning of epiblast and primitive endoderm layers. We have used combinations of embryonic stem (ES) cells and their derivatives differentiated by retinoic acid treatment to model epiblast and endoderm cells, and wild-type or E-cadherin null cells to represent strongly or weakly adherent cells, respectively. One cell type was fluorescently labeled and reconstituted with another heterotypically to generate chimeric embryoid bodies, and cell sorting was tracked by time-lapse video microscopy and confirmed by immunostaining. When undifferentiated wild-type and E-cadherin null ES cells were mixed, the resulting cell aggregates consisted of a core of wild-type cells surrounded by loosely associated E-cadherin null cells, consistent with the differential adhesion hypothesis. However, when mixed with undifferentiated ES cells, the differentiated primitive endoderm-like cells sorted to the surface to form a primitive endoderm layer irrespective of cell-adhesive strength, contradicting the differential adhesion hypothesis. We propose that the primitive endoderm cells reach the surface by random movement, and subsequently the cells generate an apical/basal polarity that prevents reentry. Thus, the ability to generate epithelial polarity, rather than adhesive affinity, determines the surface positioning of the primitive endoderm cells. (c) 2009 Wiley-Liss, Inc.

  9. Regulation of Pancreatic β Cell Mass by Cross-Interaction between CCAAT Enhancer Binding Protein β Induced by Endoplasmic Reticulum Stress and AMP-Activated Protein Kinase Activity.

    Directory of Open Access Journals (Sweden)

    Tomokazu Matsuda

    Full Text Available During the development of type 2 diabetes, endoplasmic reticulum (ER stress leads to not only insulin resistance but also to pancreatic beta cell failure. Conversely, cell function under various stressed conditions can be restored by reducing ER stress by activating AMP-activated protein kinase (AMPK. However, the details of this mechanism are still obscure. Therefore, the current study aims to elucidate the role of AMPK activity during ER stress-associated pancreatic beta cell failure. MIN6 cells were loaded with 5-amino-1-β-D-ribofuranosyl-imidazole-4-carboxamide (AICAR and metformin to assess the relationship between AMPK activity and CCAAT enhancer binding protein β (C/EBPβ expression levels. The effect of C/EBPβ phosphorylation on expression levels was also investigated. Vildagliptin and metformin were administered to pancreatic beta cell-specific C/EBPβ transgenic mice to investigate the relationship between C/EBPβ expression levels and AMPK activity in the pancreatic islets. When pancreatic beta cells are exposed to ER stress, the accumulation of the transcription factor C/EBPβ lowers the AMP/ATP ratio, thereby decreasing AMPK activity. In an opposite manner, incubation of MIN6 cells with AICAR or metformin activated AMPK, which suppressed C/EBPβ expression. In addition, administration of the dipeptidyl peptidase-4 inhibitor vildagliptin and metformin to pancreatic beta cell-specific C/EBPβ transgenic mice decreased C/EBPβ expression levels and enhanced pancreatic beta cell mass in proportion to the recovery of AMPK activity. Enhanced C/EBPβ expression and decreased AMPK activity act synergistically to induce ER stress-associated pancreatic beta cell failure.

  10. ERK1/2 signalling protects against apoptosis following endoplasmic reticulum stress but cannot provide long-term protection against BAX/BAK-independent cell death.

    Science.gov (United States)

    Darling, Nicola J; Balmanno, Kathryn; Cook, Simon J

    2017-01-01

    Disruption of protein folding in the endoplasmic reticulum (ER) causes ER stress. Activation of the unfolded protein response (UPR) acts to restore protein homeostasis or, if ER stress is severe or persistent, drive apoptosis, which is thought to proceed through the cell intrinsic, mitochondrial pathway. Indeed, cells that lack the key executioner proteins BAX and BAK are protected from ER stress-induced apoptosis. Here we show that chronic ER stress causes the progressive inhibition of the extracellular signal-regulated kinase (ERK1/2) signalling pathway. This is causally related to ER stress since reactivation of ERK1/2 can protect cells from ER stress-induced apoptosis whilst ERK1/2 pathway inhibition sensitises cells to ER stress. Furthermore, cancer cell lines harbouring constitutively active BRAFV600E are addicted to ERK1/2 signalling for protection against ER stress-induced cell death. ERK1/2 signalling normally represses the pro-death proteins BIM, BMF and PUMA and it has been proposed that ER stress induces BIM-dependent cell death. We found no evidence that ER stress increased the expression of these proteins; furthermore, BIM was not required for ER stress-induced death. Rather, ER stress caused the PERK-dependent inhibition of cap-dependent mRNA translation and the progressive loss of pro-survival proteins including BCL2, BCLXL and MCL1. Despite these observations, neither ERK1/2 activation nor loss of BAX/BAK could confer long-term clonogenic survival to cells exposed to ER stress. Thus, ER stress induces cell death by at least two biochemically and genetically distinct pathways: a classical BAX/BAK-dependent apoptotic response that can be inhibited by ERK1/2 signalling and an alternative ERK1/2- and BAX/BAK-independent cell death pathway.

  11. ERK1/2 signalling protects against apoptosis following endoplasmic reticulum stress but cannot provide long-term protection against BAX/BAK-independent cell death.

    Directory of Open Access Journals (Sweden)

    Nicola J Darling

    Full Text Available Disruption of protein folding in the endoplasmic reticulum (ER causes ER stress. Activation of the unfolded protein response (UPR acts to restore protein homeostasis or, if ER stress is severe or persistent, drive apoptosis, which is thought to proceed through the cell intrinsic, mitochondrial pathway. Indeed, cells that lack the key executioner proteins BAX and BAK are protected from ER stress-induced apoptosis. Here we show that chronic ER stress causes the progressive inhibition of the extracellular signal-regulated kinase (ERK1/2 signalling pathway. This is causally related to ER stress since reactivation of ERK1/2 can protect cells from ER stress-induced apoptosis whilst ERK1/2 pathway inhibition sensitises cells to ER stress. Furthermore, cancer cell lines harbouring constitutively active BRAFV600E are addicted to ERK1/2 signalling for protection against ER stress-induced cell death. ERK1/2 signalling normally represses the pro-death proteins BIM, BMF and PUMA and it has been proposed that ER stress induces BIM-dependent cell death. We found no evidence that ER stress increased the expression of these proteins; furthermore, BIM was not required for ER stress-induced death. Rather, ER stress caused the PERK-dependent inhibition of cap-dependent mRNA translation and the progressive loss of pro-survival proteins including BCL2, BCLXL and MCL1. Despite these observations, neither ERK1/2 activation nor loss of BAX/BAK could confer long-term clonogenic survival to cells exposed to ER stress. Thus, ER stress induces cell death by at least two biochemically and genetically distinct pathways: a classical BAX/BAK-dependent apoptotic response that can be inhibited by ERK1/2 signalling and an alternative ERK1/2- and BAX/BAK-independent cell death pathway.

  12. Chromosome segregation in plant meiosis

    Directory of Open Access Journals (Sweden)

    Linda eZamariola

    2014-06-01

    Full Text Available Faithful chromosome segregation in meiosis is essential for ploidy stability over sexual life cycles. In plants, defective chromosome segregation caused by gene mutations or other factors leads to the formation of unbalanced or unreduced gametes creating aneuploid or polyploid progeny, respectively. Accurate segregation requires the coordinated execution of conserved processes occurring throughout the two meiotic cell divisions. Synapsis and recombination ensure the establishment of chiasmata that hold homologous chromosomes together allowing their correct segregation in the first meiotic division, which is also tightly regulated by cell-cycle dependent release of cohesin and monopolar attachment of sister kinetochores to microtubules. In meiosis II, bi-orientation of sister kinetochores and proper spindle orientation correctly segregate chromosomes in four haploid cells. Checkpoint mechanisms acting at kinetochores control the accuracy of kinetochore-microtubule attachment, thus ensuring the completion of segregation. Here we review the current knowledge on the processes taking place during chromosome segregation in plant meiosis, focusing on the characterization of the molecular factors involved.

  13. Coordination between chromosome replication, segregation, and cell division in Caulobacter crescentus

    DEFF Research Database (Denmark)

    Jensen, Rasmus Bugge

    2006-01-01

    Progression through the Caulobacter crescentus cell cycle is coupled to a cellular differentiation program. The swarmer cell is replicationally quiescent, and DNA replication initiates at the swarmer-to-stalked cell transition. There is a very short delay between initiation of DNA replication...

  14. Tetrandrine Induces Apoptosis in Human Nasopharyngeal Carcinoma NPC-TW 039 Cells by Endoplasmic Reticulum Stress and Ca2+/Calpain Pathways.

    Science.gov (United States)

    Liu, Kuo-Ching; Lin, Ya-Jing; Hsiao, Yung-Ting; Lin, Meng-Liang; Yang, Jiun-Long; Huang, Yi-Ping; Chu, Yung-Lin; Chung, Jing-Gung

    2017-11-01

    Tetrandrine is an alkaloid extracted from a traditional China medicine plant, and is considered part of food therapy as well. In addition, it has been widely reported to induce apoptotic cell death in many human cancer cells. However, the mechanism of Tetrandrine on human nasopharyngeal carcinoma cells (NPC) is still questioned. In our study, we examined whether Tetrandrine can induce apoptosis of NPC-TW 039 cells. We found that cell morphology was changed after treatment with different concentrations of Tetrandrine. Further, we indicated that the NPC-TW 039 cells viability decreased in a Tetrandrine dose-dependent manner. We also found that tetrandrine induced cell cycle arrest in G 0 /G 1 phase. Tetrandrine induced DNA condensation by DAPI staining as well. In addition, we found that Tetrandrine induced Ca 2+ release in the cytosol. At the same time, endoplasmic reticulum (ER) stress occurred. Then we used western blotting to examine the protein expression which is associated with mitochondria-mediated apoptotic pathways and caspase-dependent pathways. To further examine whether Ca 2+ was released or not with Tetrandrine induced-apoptosis, we used the chelator of Ca 2+ and showed that cell viability increased. At the same time, caspase-3 expression was decreased. Furthermore, confocal microscopy examination revealed that Tetrandrine induced expression of ER stress-related proteins GADD153 and GRP78. Our results indicate that Tetrandrine induces apoptosis through calcium-mediated ER stress and caspase pathway in NPC-TW 039 cells. In conclusion, Tetrandrine may could be used for treatment of human nasopharyngeal carcinoma in future. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

  15. Continuous in vitro exposure of intestinal epithelial cells to E171 food additive causes oxidative stress, inducing oxidation of DNA bases but no endoplasmic reticulum stress.

    Science.gov (United States)

    Dorier, Marie; Béal, David; Marie-Desvergne, Caroline; Dubosson, Muriel; Barreau, Frédérick; Houdeau, Eric; Herlin-Boime, Nathalie; Carriere, Marie

    2017-08-01

    The whitening and opacifying properties of titanium dioxide (TiO 2 ) are commonly exploited when it is used as a food additive (E171). However, the safety of this additive can be questioned as TiO 2 nanoparticles (TiO 2 -NPs) have been classed at potentially toxic. This study aimed to shed some light on the mechanisms behind the potential toxicity of E171 on epithelial intestinal cells, using two in vitro models: (i) a monoculture of differentiated Caco-2 cells and (ii) a coculture of Caco-2 with HT29-MTX mucus-secreting cells. Cells were exposed to E171 and two different types of TiO 2 -NPs, either acutely (6-48 h) or repeatedly (three times a week for 3 weeks). Our results confirm that E171 damaged these cells, and that the main mechanism of toxicity was oxidation effects. Responses of the two models to E171 were similar, with a moderate, but significant, accumulation of reactive oxygen species, and concomitant downregulation of the expression of the antioxidant enzymes catalase, superoxide dismutase and glutathione reductase. Oxidative damage to DNA was detected in exposed cells, proving that E171 effectively induces oxidative stress; however, no endoplasmic reticulum stress was detected. E171 effects were less intense after acute exposure compared to repeated exposure, which correlated with higher Ti accumulation. The effects were also more intense in cells exposed to E171 than in cells exposed to TiO 2 -NPs. Taken together, these data show that E171 induces only moderate toxicity in epithelial intestinal cells, via oxidation.

  16. Fisetin Induces Apoptosis of HSC3 Human Oral Cancer Cells Through Endoplasmic Reticulum Stress and Dysfunction of Mitochondria-mediated Signaling Pathways.

    Science.gov (United States)

    Shih, Yung-Luen; Hung, Fang-Ming; Lee, Ching-Hsiao; Yeh, Ming-Yang; Lee, Mei-Hui; Lu, Hsu-Feng; Chen, Yung-Liang; Liu, Jia-You; Chung, Jing-Gung

    2017-01-01

    Oral cancer has been reported to be one of the major cancer-related diseases in human populations and the treatment of oral cancer is still unsatisfied. Fisetin, is a flavonoid from plants and has several biological activities such as antioxidant, anti-inflammatory and anticancer function, but its cytotoxicity in human oral cancer cells is unknown. In the present study, we investigated fisetin-induced cytotoxic effects on HSC3 human oral cancer cells in vitro. Materials and Methods/Results: We used flow cytometric assay to show fisetin induced apoptotic cell death through increased reactive oxygen species and Ca 2+ , but reduced the mitochondrial membrane potential and increased caspase-8, -9 and -3 activities in HSC3 cells. Furthermore, we also used 4' 6-diamidino-2-phenylindole staining to show that fisetin induced chromatin condensation (apoptotic cell death), and Comet assay to show that fisetin induced DNA damage in HSC3 cells. Western blotting was used to examine the levels of apoptotic-associated protein and results indicated that fisetin increased expression of pro-apoptotic proteins such as B-cell lymphoma 2 (BCL2) antagonist/killer (BAK) and BCL2-associated X (BAX) but reduced that of anti-apoptotic protein such as BCL2 and BCL-x, and increased the cleaved forms of caspase-3, -8 and -9, and cytochrome c, apoptosis-inducing factor (AIF) and endonuclease G (ENDO G) in HSC3 cells. Confocal microscopy showed that fisetin increased the release of cytochrome c, AIF and ENDO G from mitochondria into the cytoplasm. Based on these observations, we suggest that fisetin induces apoptotic cell death through endoplasmic reticulum stress- and mitochondria-dependent pathways. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

  17. Sodium phenylbutyrate, a drug with known capacity to reduce endoplasmic reticulum stress, partially alleviates lipid-induced insulin resistance and beta-cell dysfunction in humans.

    Science.gov (United States)

    Xiao, Changting; Giacca, Adria; Lewis, Gary F

    2011-03-01

    Chronically elevated free fatty acids contribute to insulin resistance and pancreatic β-cell failure. Among numerous potential factors, the involvement of endoplasmic reticulum (ER) stress has been postulated to play a mechanistic role. Here we examined the efficacy of the chemical chaperone, sodium phenylbutyrate (PBA), a drug with known capacity to reduce ER stress in animal models and in vitro, on lipid-induced insulin resistance and β-cell dysfunction in humans. Eight overweight or obese nondiabetic men underwent four studies each, in random order, 4 to 6 weeks apart. Two studies were preceded by 2 weeks of oral PBA (7.5 g/day), followed by a 48-h i.v. infusion of intralipid/heparin or saline, and two studies were preceded by placebo treatment, followed by similar infusions. Insulin secretion rates (ISRs) and sensitivity (S(I)) were assessed after the 48-h infusions by hyperglycemic and hyperinsulinemic-euglycemic clamps, respectively. Lipid infusion reduced S(I), which was significantly ameliorated by pretreatment with PBA. Absolute ISR was not affected by any treatment; however, PBA partially ameliorated the lipid-induced reduction in the disposition index (DI = ISR × S(I)), indicating that PBA prevented lipid-induced β-cell dysfunction. These results suggest that PBA may provide benefits in humans by ameliorating the insulin resistance and β-cell dysfunction induced by prolonged elevation of free fatty acids.

  18. Statolith Sedimentation Kinetics and Force Transduction to the Cortical Endoplasmic Reticulum in Gravity-Sensing Arabidopsis Columella Cells[W][OA

    Science.gov (United States)

    Leitz, Guenther; Kang, Byung-Ho; Schoenwaelder, Monica E.A.; Staehelin, L. Andrew

    2009-01-01

    The starch statolith hypothesis of gravity sensing in plants postulates that the sedimentation of statoliths in specialized statocytes (columella cells) provides the means for converting the gravitational potential energy into a biochemical signal. We have analyzed the sedimentation kinetics of statoliths in the central S2 columella cells of Arabidopsis thaliana. The statoliths can form compact aggregates with gap sizes between statoliths approaching sedimentation phase, the statoliths tend to move at a distance to the cortical endoplasmic reticulum (ER) boundary and interact only transiently with the ER. Statoliths moved by laser tweezers against the ER boundary experience an elastic lift force upon release from the optical trap. High-resolution electron tomography analysis of statolith-to-ER contact sites indicate that the weight of statoliths is sufficient to locally deform the ER membranes that can potentially activate mechanosensitive ion channels. We suggest that in root columella cells, the transduction of the kinetic energy of sedimenting statoliths into a biochemical signal involves a combination of statolith-driven motion of the cytosol, statolith-induced deformation of the ER membranes, and a rapid release of kinetic energy from the ER during reorientation to activate mechanosensitive sites within the central columella cells. PMID:19276442

  19. Inhibitory Effects of Simvastatin on Oxidized Low-Density Lipoprotein-Induced Endoplasmic Reticulum Stress and Apoptosis in Vascular Endothelial Cells.

    Science.gov (United States)

    Zhang, Guo-Qiang; Tao, Yong-Kang; Bai, Yong-Ping; Yan, Sheng-Tao; Zhao, Shui-Ping

    2018-04-20

    Oxidized low-density lipoprotein (ox-LDL)-induced oxidative stress and endothelial apoptosis are essential for atherosclerosis. Our previous study has shown that ox-LDL-induced apoptosis is mediated by the protein kinase RNA-like endoplasmic reticulum kinase (PERK)/eukaryotic translation initiation factor 2α-subunit (eIF2α)/CCAAT/enhancer-binding protein homologous protein (CHOP) endoplasmic reticulum (ER) stress pathway in endothelial cells. Statins are cholesterol-lowering drugs that exert pleiotropic effects including suppression of oxidative stress. This study aimed to explore the roles of simvastatin on ox-LDL-induced ER stress and apoptosis in endothelial cells. Human umbilical vein endothelial cells (HUVECs) were treated with simvastatin (0.1, 0.5, or 2.5 μmol/L) or DEVD-CHO (selective inhibitor of caspase-3, 100 μmol/L) for 1 h before the addition of ox-LDL (100 μg/ml) and then incubated for 24 h, and untreated cells were used as a control group. Apoptosis, expression of PERK, phosphorylation of eIF2α, CHOP mRNA level, and caspase-3 activity were measured. Comparisons among multiple groups were performed with one-way analysis of variance (ANOVA) followed by post hoc pairwise comparisons using Tukey's tests. A value of P LDL resulted in a significant increase in apoptosis (31.9% vs. 4.9%, P LDL-induced apoptosis (28.0%, 24.7%, and 13.8%, F = 15.039, all P LDL significantly increased the expression of PERK (499.5%, P LDL-induced expression of PERK (407.8%, 339.1%, and 187.5%, F = 10.121, all P LDL-induced expression of PERK (486.4%) and phosphorylation of eIF2α (418.8%). Exposure of HUVECs to ox-LDL also markedly induced caspase-3 activity together with increased CHOP mRNA level; these effects were inhibited by simvastatin treatment. This study suggested that simvastatin could inhibit ox-LDL-induced ER stress and apoptosis in vascular endothelial cells.

  20. Rhein triggers apoptosis via induction of endoplasmic reticulum stress, caspase-4 and intracellular calcium in primary human hepatic HL-7702 cells

    Energy Technology Data Exchange (ETDEWEB)

    KoraMagazi, Arouna [Department of Clinical Pharmacy, China Pharmaceutical University, Nanjing, Jiangsu (China); Wang, Dandan [Department of Pharmacology, China Pharmaceutical University, Nanjing, Jiangsu (China); Yousef, Bashir; Guerram, Mounia [Jiangsu Key Laboratory of Drug Screening, China Pharmaceutical University, Nanjing, Jiangsu (China); Yu, Feng, E-mail: yufengcpu14@yahoo.com [Department of Clinical Pharmacy, China Pharmaceutical University, Nanjing, Jiangsu (China); Department of Pharmacology, China Pharmaceutical University, Nanjing, Jiangsu (China); Key Laboratory of Drug Quality Control and Pharmacovigilance, China Pharmaceutical University, Nanjing, Jiangsu (China)

    2016-04-22

    Rhein is an active component of rhubarb; a traditional Chinese medicine reported to induce apoptosis and cause liver toxicity. However, rhein's apoptotic-inducing effects, as well as its molecular mechanisms of action on hepatic cells need to be further explored. In the present study, rhein was found to trigger apoptosis in primary human hepatic HL-7702 cells as showed by annexin V/PI double staining assay and nuclear morphological changes demonstrated by Hoechst 33258 staining. Moreover, it was observed that the mechanism implicated in rhein-induced apoptosis was caspase-dependent, presumably via ER-stress associated pathways, as illustrated by up-regulation of glucose-regulated protein 78 (GRP 78), PKR-like ER kinase (PERK), C-Jun N-terminal kinase (JNK) and CCAAT/enhancer-binding protein homologous protein (CHOP). Meanwhile, caspase-4 as a hallmark of ER-stress, was also showed to be activated following by caspase-3 activation. Furthermore, rhein also promoted intracellular elevation of calcium that contributed in apoptosis induction. Interestingly, pre-treatment with calpain inhibitor I reduced the effects of rhein on apoptosis induction and JNK activation. These data suggested that rhein-induced apoptosis through ER-stress and elevated intracellular calcium level in HL-7702 cells. - Highlights: • Rhein triggers apoptotic cell death on primary human hepatic HL-7702 cells. • Rhein leads to caspase-4 activation in HL-7702 cells. • Rhein induces endoplasmic reticulum stress pathways in HL-7702 cells. • Rhein causes elevation of intracellular calcium concentrations in HL-7702 cells.

  1. 14-Deoxy-11,12-didehydroandrographolide induces DDIT3-dependent endoplasmic reticulum stress-mediated autophagy in T-47D breast carcinoma cells

    International Nuclear Information System (INIS)

    Tan, Heng Kean; Muhammad, Tengku Sifzizul Tengku; Tan, Mei Lan

    2016-01-01

    14-Deoxy-11,12-didehydroandrographolide (14-DDA), a major diterpenoid isolated from Andrographis paniculata (Burm.f.) Nees, is known to be cytotoxic and elicits a non-apoptotic cell death in T-47D breast carcinoma cells. In this study, the mechanistic toxicology properties of 14-DDA in T-47D cells were further investigated. 14-DDA is found to induce the formation of endoplasmic reticulum (ER) vacuoles and autophagosomes, with concurrent upregulation of LC3-II in the breast carcinoma cells. It stimulated an increase in cytosolic calcium concentration and caused a collapse in mitochondrial membrane potential in these cells. In addition, both DDIT3 and GADD45A, molecules implicated in ER stress pathway, were significantly upregulated. DDIT3 knockdown suppressed the formation of both ER vacuoles and autophagosomes, indicating that 14-DDA-induced ER stress and autophagy is dependent on this transcription factor. Collectively, it is possible that GADD45A/p38 MAPK/DDIT3 pathway is involved in the 14-DDA-induced ER-stress-mediated autophagy in T-47D cells. - Highlights: • The mechanistic toxicology properties of 14-DDA in T-47D breast carcinoma cells were investigated. • 14-DDA induces the formation of ER vacuoles and autophagosomes, with concurrent upregulation of LC3-II. • It stimulates an increase in cytosolic calcium concentration and causing collapse in the mitochondrial membrane potential. • Both DDIT3 and GADD45A, molecules implicated in ER stress pathway, were significantly upregulated. • 4-DDA induces ER stress-mediated autophagy in T-47D cells possibly via GADD45A/p38 MAPK/DDIT3 pathway.

  2. Endoplasmic Reticulum-Targeted Subunit Toxins Provide a New Approach to Rescue Misfolded Mutant Proteins and Revert Cell Models of Genetic Diseases.

    Directory of Open Access Journals (Sweden)

    Humaira Adnan

    Full Text Available Many germ line diseases stem from a relatively minor disturbance in mutant protein endoplasmic reticulum (ER 3D assembly. Chaperones are recruited which, on failure to correct folding, sort the mutant for retrotranslocation and cytosolic proteasomal degradation (ER-associated degradation-ERAD, to initiate/exacerbate deficiency-disease symptoms. Several bacterial (and plant subunit toxins, retrograde transport to the ER after initial cell surface receptor binding/internalization. The A subunit has evolved to mimic a misfolded protein and hijack the ERAD membrane translocon (dislocon, to effect cytosolic access and cytopathology. We show such toxins compete for ERAD to rescue endogenous misfolded proteins. Cholera toxin or verotoxin (Shiga toxin containing genetically inactivated (± an N-terminal polyleucine tail A subunit can, within 2-4 hrs, temporarily increase F508delCFTR protein, the major cystic fibrosis (CF mutant (5-10x, F508delCFTR Golgi maturation (<10x, cell surface expression (20x and chloride transport (2x in F508del CFTR transfected cells and patient-derived F508delCFTR bronchiolar epithelia, without apparent cytopathology. These toxoids also increase glucocerobrosidase (GCC in N370SGCC Gaucher Disease fibroblasts (3x, another ERAD-exacerbated misfiling disease. We identify a new, potentially benign approach to the treatment of certain genetic protein misfolding diseases.

  3. SIRT1 attenuates palmitate-induced endoplasmic reticulum stress and insulin resistance in HepG2 cells via induction of oxygen-regulated protein 150

    Science.gov (United States)

    Jung, T.W.; Lee, K.T.; Lee, M.W.; Ka, K.H.

    2012-01-01

    Endoplasmic reticulum (ER) stress has been implicated in the pathology of type 2 diabetes mellitus (T2DM). Although SIRT1 has a therapeutic effect on T2DM, the mechanisms by which SIRT1 ameliorates insulin resistance (IR) remain unclear. In this study, we investigated the impact of SIRT1 on palmitate-induced ER stress in HepG2 cells and its underlying signal pathway. Treatment with resveratrol, a SIRT1 activator significantly inhibited palmitate-induced ER stress, leading to the protection against palmitate-induced ER stress and insulin resistance. Resveratrol and SIRT1 overexpression induced the expression of oxygen-regulated protein (ORP) 150 in HepG2 cells. Forkhead box O1 (FOXO1) was involved in the regulation of ORP150 expression because suppression of FOXO1 inhibited the induction of ORP150 by SIRT1. Our results indicate a novel mechanism by which SIRT1 regulates ER stress by overexpression of ORP150, and suggest that SIRT1 ameliorates palmitate-induced insulin resistance in HepG2 cells via regulation of ER stress.

  4. TSA protects H9c2 cells against thapsigargin-induced apoptosis related to endoplasmic reticulum stress-mediated mitochondrial injury.

    Science.gov (United States)

    Li, Zhiping; Liu, Yan; Dai, Xinlun; Zhou, Qiangqiang; Liu, Xueli; Li, Zeyu; Chen, Xia

    2017-05-01

    Endoplasmic reticulum stress (ERS) activates an adaptive unfolded protein response (UPR) that facilitates cellular repair, however, under prolonged ER stress, the UPR can ultimately trigger apoptosis thereby terminating damaged cells. Recently, TSA has shown protective effects on ERS and its mechanisms related to ER pathway has been previously characterized. However, whether TSA exerts its protective role via metabolic events remain largely undefined. Objectives : To explore the possible involvement of the metabolic changes during ERS and to better understand how TSA influence mitochondrial function to facilitate cellular adaptation. Results : TSA is an inhibitor of histone deacetylase which could significantly inhibit H9c2 cell apoptosis induced by Thapsigargin (TG). It also intervene the decrease of mitochondrial membrane potential. By immunofluorescence staining, we have shown that GRP78 was concentrated in the perinuclear region and co-localized with ER. However, treatments with TG and TSA could let it overlap with the mitochondrial marker MitoTracker. Cellular fractionation also confirmed the location of GRP78 in mitochondrion. TSA decreases ERS-induced cell apoptosis and mitochondrial injury may related to enhance the location of GRP78 in mitochondrion.

  5. Disease-causing mitochondrial heteroplasmy segregated within induced pluripotent stem cell clones derived from a patient with MELAS.

    Science.gov (United States)

    Folmes, Clifford D L; Martinez-Fernandez, Almudena; Perales-Clemente, Ester; Li, Xing; McDonald, Amber; Oglesbee, Devin; Hrstka, Sybil C; Perez-Terzic, Carmen; Terzic, Andre; Nelson, Timothy J

    2013-07-01

    Mitochondrial diseases display pathological phenotypes according to the mixture of mutant versus wild-type mitochondrial DNA (mtDNA), known as heteroplasmy. We herein examined the impact of nuclear reprogramming and clonal isolation of induced pluripotent stem cells (iPSC) on mitochondrial heteroplasmy. Patient-derived dermal fibroblasts with a prototypical mitochondrial deficiency diagnosed as mitochondrial encephalomyopathy with lactic acidosis and stroke-like episodes (MELAS) demonstrated mitochondrial dysfunction with reduced oxidative reserve due to heteroplasmy at position G13513A in the ND5 subunit of complex I. Bioengineered iPSC clones acquired pluripotency with multilineage differentiation capacity and demonstrated reduction in mitochondrial density and oxygen consumption distinguishing them from the somatic source. Consistent with the cellular mosaicism of the original patient-derived fibroblasts, the MELAS-iPSC clones contained a similar range of mtDNA heteroplasmy of the disease-causing mutation with identical profiles in the remaining mtDNA. High-heteroplasmy iPSC clones were used to demonstrate that extended stem cell passaging was sufficient to purge mutant mtDNA, resulting in isogenic iPSC subclones with various degrees of disease-causing genotypes. On comparative differentiation of iPSC clones, improved cardiogenic yield was associated with iPSC clones containing lower heteroplasmy compared with isogenic clones with high heteroplasmy. Thus, mtDNA heteroplasmic segregation within patient-derived stem cell lines enables direct comparison of genotype/phenotype relationships in progenitor cells and lineage-restricted progeny, and indicates that cell fate decisions are regulated as a function of mtDNA mutation load. The novel nuclear reprogramming-based model system introduces a disease-in-a-dish tool to examine the impact of mutant genotypes for MELAS patients in bioengineered tissues and a cellular probe for molecular features of individual

  6. 6-Shogaol induces apoptosis in human hepatocellular carcinoma cells and exhibits anti-tumor activity in vivo through endoplasmic reticulum stress.

    Directory of Open Access Journals (Sweden)

    Rong Hu

    Full Text Available 6-Shogaol is an active compound isolated from Ginger (Zingiber officinale Rosc. In this work, we demonstrated that 6-shogaol induces apoptosis in human hepatocellular carcinoma cells in relation to caspase activation and endoplasmic reticulum (ER stress signaling. Proteomic analysis revealed that ER stress was accompanied by 6-shogaol-induced apoptosis in hepatocellular carcinoma cells. 6-shogaol affected the ER stress signaling by regulating unfolded protein response (UPR sensor PERK and its downstream target eIF2α. However, the effect on the other two UPR sensors IRE1 and ATF6 was not obvious. In prolonged ER stress, 6-shogaol inhibited the phosphorylation of eIF2α and triggered apoptosis in SMMC-7721 cells. Salubrinal, an activator of the PERK/eIF2α pathway, strikingly enhanced the phosphorylation of eIF2α in SMMC-7721 cells with no toxicity. However, combined treatment with 6-shogaol and salubrinal resulted in significantly increase of apoptosis and dephosphorylation of eIF2α. Overexpression of eIF2α prevented 6-shogaol-mediated apoptosis in SMMC-7721 cells, whereas inhibition of eIF2α by small interfering RNA markedly enhanced 6-shogaol-mediated cell death. Furthermore, 6-shogaol-mediated inhibition of tumor growth of mouse SMMC-7721 xenograft was associated with induction of apoptosis, activation of caspase-3, and inactivation of eIF2α. Altogether our results indicate that the PERK/eIF2α pathway plays an important role in 6-shogaol-mediated ER stress and apoptosis in SMMC-7721 cells in vitro and in vivo.

  7. Ubiquitin-proteasomal degradation of COX-2 in TGF-β stimulated human endometrial cells is mediated through endoplasmic reticulum mannosidase I.

    Science.gov (United States)

    Singh, Mohan; Chaudhry, Parvesh; Parent, Sophie; Asselin, Eric

    2012-01-01

    Cyclooxygenase (COX)-2 is a key regulatory enzyme in the production of prostaglandins (PG) during various physiological processes. Mechanisms of COX-2 regulation in human endometrial stromal cells (human endometrial stromal cells) are not fully understood. In this study, we investigate the role of TGF-β in the regulation of COX-2 in human uterine stromal cells. Each TGF-β isoform decreases COX-2 protein level in human uterine stromal cells in Smad2/3-dependent manner. The decrease in COX-2 is accompanied by a decrease in PG synthesis. Knockdown of Smad4 using specific small interfering RNA prevents the decrease in COX-2 protein, confirming that Smad pathway is implicated in the regulation of COX-2 expression in human endometrial stromal cells. Pretreatment with 26S proteasome inhibitor, MG132, significantly restores COX-2 protein and PG synthesis, indicating that COX-2 undergoes proteasomal degradation in the presence of TGF-β. In addition, each TGF-β isoform up-regulates endoplasmic reticulum (ER)-mannosidase I (ERManI) implying that COX-2 degradation is mediated through ER-associated degradation pathway in these cells. Furthermore, inhibition of ERManI activity using the mannosidase inhibitor (kifunensine), or small interfering RNA-mediated knockdown of ERManI, prevents TGF-β-induced COX-2 degradation. Taken together, these studies suggest that TGF-β promotes COX-2 degradation in a Smad-dependent manner by up-regulating the expression of ERManI and thereby enhancing ER-associated degradation and proteasomal degradation pathways.

  8. PICH promotes mitotic chromosome segregation

    DEFF Research Database (Denmark)

    Nielsen, Christian Thomas Friberg; Hickson, Ian D

    2016-01-01

    PICH is an SNF2-family DNA translocase that appears to play a role specifically in mitosis. Characterization of PICH in human cells led to the initial discovery of "ultra-fine DNA bridges" (UFBs) that connect the 2 segregating DNA masses in the anaphase of mitosis. These bridge structures, which...... further the role of PICH in the timely segregation of the rDNA locus....

  9. Effect of Cudrania tricuspidata and Kaempferol in Endoplasmic Reticulum Stress-Induced Inflammation and Hepatic Insulin Resistance in HepG2 Cells.

    Science.gov (United States)

    Kim, Ok-Kyung; Jun, Woojin; Lee, Jeongmin

    2016-01-21

    In this study, we quantitated kaempferol in water extract from Cudrania tricuspidata leaves (CTL) and investigated its effects on endoplasmic reticulum (ER) stress-induced inflammation and insulin resistance in HepG2 cells. The concentration of kaempferol in the CTL was 5.07 ± 0.08 mg/g. The HepG2 cells were treated with 300 µg/mL of CTL, 500 µg/mL of CTL, 1.5 µg/mL of kaempferol or 2.5 µg/mL of kaempferol, followed immediately by stimulation with 100 nM of thapsigargin for ER stress induction for 24 h. There was a marked increase in the activation of the ER stress and inflammation response in the thapsigargin-stimulated control group. The CTL treatment interrupted the ER stress response and ER stress-induced inflammation. Kaempferol partially inhibited the ER stress response and inflammation. There was a significant increase in serine phosphorylation of insulin receptor substrate (IRS)-1 and the expression of C/EBPα and gluconeogenic genes in the thapsigargin-stimulated control group compared to the normal control. Both CTL and kaempferol suppressed serine phosphorylation of IRS-1, and the treatments did not interrupt the C/EBPα/gluconeogenic gene pathway. These results suggest that kaempferol might be the active compound of CTL and that it might protect against ER stress-induced inflammation and hyperglycemia.

  10. A hydrophobic organelle probe based on aggregation-induced emission: Nanosuspension preparation and direct use for endoplasmic reticulum imaging in living cells

    Science.gov (United States)

    Zheng, Sichao; Huang, Cuihong; Zhao, Xuyan; Zhang, Yong; Liu, Shuwen; Zhu, Qiuhua

    2018-01-01

    Organic fluorophores have a wide range of biological uses and are usually needed to be prepared as water-soluble compounds or nanoparticles for applications in aqueous biosystems owing to their hydrophobic properties, which often is a complex, time-consuming and high-cost process. Here, the nanoparticle preparation of hydrophobic fluorophores and their application in cell imaging have been investigated. It was found: a) fetal bovine serum (FBS) shows an excellent dispersion effect on hydrophobic small-molecule organic compounds; b) a hydrophobic C6-unsubstituted tetrahydropyrimidine (Me-THP-Naph) can be prepared as nanosuspensions utilizing cell culture medium with 10% FBS and directly be used as a specific real-time imaging probe for the endoplasmic reticulum (ER), a dynamic organelle playing a crucial role in many cellular processes. Compared with existing ER-targeted organic fluorescent probes, Me-THP-Naph, a product of an efficient five-component reaction that we developed, has unconventional aggregation-induced emission characteristics and shows advantages of low cost, long-term staining, good photostability, high signal-to-noise ratio and excellent biocompatibility, which make it a potential specific probe for real-time ER imaging. More importantly, this work affords a simple strategy for direct application of hydrophobic organic compounds in aqueous biological systems.

  11. Dynamin-Related Protein 1 Inhibitors Protect against Ischemic Toxicity through Attenuating Mitochondrial Ca2+ Uptake from Endoplasmic Reticulum Store in PC12 Cells

    Directory of Open Access Journals (Sweden)

    Ye Tian

    2014-02-01

    Full Text Available Intracellular calcium homeostasis disorder and mitochondrial dysfunction are involved in many acute and chronic brain diseases, including ischemic brain injury. An imbalance in mitochondrial fission and fusion is one of the most important structural abnormalities found in a large number of mitochondrial dysfunction related diseases. Here, we investigated the effects of mitochondrial division inhibitor A (mdivi A and mdivi B, two small molecule inhibitors of mitochondrial fission protein dunamin-related protein 1 (Drp-1, in neuronal injury induced by oxygen-glucose deprivation (OGD in PC12 cells. We found that mdivi A and mdivi B inhibited OGD-induced neuronal injury through attenuating apoptotic cell death. These two inhibitors also preserved mitochondrial function, as evidenced by reduced reactive oxygen species (ROS generation and cytochrome c release, as well as prevented loss of mitochondrial membrane potential (MMP. Moreover, mdivi A and mdivi B significantly suppressed mitochondrial Ca2+ uptake, but had no effect on cytoplasmic Ca2+ after OGD injury. The results of calcium imaging and immunofluorescence staining showed that Drp-1 inhibitors attenuated endoplasmic reticulum (ER Ca2+ release and prevented ER morphological changes induced by OGD. These results demonstrate that Drp-1 inhibitors protect against ischemic neuronal injury through inhibiting mitochondrial Ca2+ uptake from the ER store and attenuating mitochondrial dysfunction.

  12. Segregation of glutamatergic and cholinergic transmission at the mixed motoneuron Renshaw cell synapse.

    OpenAIRE

    Lamotte d Incamps, B.; Bhumbra, G. S.; Foster, J. D.; Beato, M.; Ascher, P.

    2017-01-01

    In neonatal mice motoneurons excite Renshaw cells by releasing both acetylcholine (ACh) and glutamate. These two neurotransmitters activate two types of nicotinic receptors (nAChRs) (the homomeric α7 receptors and the heteromeric α*ß* receptors) as well as the two types of glutamate receptors (GluRs) (AMPARs and NMDARs). Using paired recordings, we confirm that a single motoneuron can release both transmitters on a single post-synaptic Renshaw cell. We then show that co-transmission is prese...

  13. Angiotensin II receptor one (AT1) mediates dextrose induced endoplasmic reticulum stress and superoxide production in human coronary artery endothelial cells.

    Science.gov (United States)

    Haas, Michael J; Onstead-Haas, Luisa; Lee, Tracey; Torfah, Maisoon; Mooradian, Arshag D

    2016-10-01

    Renin-angiotensin-aldosterone system (RAAS) has been implicated in diabetes-related vascular complications partly through oxidative stress. To determine the role of angiotensin II receptor subtype one (AT1) in dextrose induced endoplasmic reticulum (ER) stress, another cellular stress implicated in vascular disease. Human coronary artery endothelial cells with or without AT1 receptor knock down were treated with 27.5mM dextrose for 24h in the presence of various pharmacologic blockers of RAAS and ER stress and superoxide (SO) production were measured. Transfection of cells with AT1 antisense RNA knocked down cellular AT1 by approximately 80%. The ER stress was measured using the placental alkaline phosphatase (ES-TRAP) assay and western blot analysis of glucose regulated protein 78 (GRP78), c-jun-N-terminal kinase 1 (JNK1), phospho-JNK1, eukaryotic translation initiation factor 2α (eIF2α) and phospho-eIF2α measurements. Superoxide (SO) generation was measured using the superoxide-reactive probe 2-methyl-6-(4-methoxyphenyl)-3,7-dihydroimidazo[1,2-A]pyrazin-3-one hydrochloride (MCLA) chemiluminescence. In cells with AT1 knock down, dextrose induced ER stress was significantly blunted and treatment with 27.5mM dextrose resulted in significantly smaller increase in SO production compared to 27.5mM dextrose treated and sham transfected cells. Dextrose induced ER stress was reduced with pharmacologic blockers of AT1 (losartan and candesartan) and mineralocorticoid receptor blocker (spironolactone) but not with angiotensin converting enzyme inhibitors (captopril and lisinopril). The dextrose induced SO generation was inhibited by all pharmacologic blockers of RAAS tested. The results indicate that dextrose induced ER stress and SO production in endothelial cells are mediated at least partly through AT1 receptor activation. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  14. Morphology versus vertical phase segregation in solvent annealed small molecule bulk heterojunction organic solar cells

    Czech Academy of Sciences Publication Activity Database

    Kovalenko, A.; Stoyanova, V.; Pospisil, J.; Zhivkov, I.; Fekete, Ladislav; Karashanova, D.; Kratochvílová, Irena; Vala, M.; Weiter, M.

    2015-01-01

    Roč. 2015, Oct (2015), s. 238981 ISSN 1110-662X R&D Projects: GA ČR(CZ) GA15-05095S; GA TA ČR TA04020156; GA MŠk LO1409 Institutional support: RVO:68378271 Keywords : organic solar cells Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 1.226, year: 2015

  15. Endoplasmic reticulum stress and IRE-1 signaling cause apoptosis in colon cancer cells in response to andrographolide treatment

    OpenAIRE

    Banerjee, Aditi; Ahmed, Hafiz; Yang, Peixin; Czinn, Steven J.; Blanchard, Thomas G.

    2016-01-01

    The plant metabolite andrographolide induces cell cycle arrest and apoptosis in cancer cells. The mechanism(s) by which andrographolide induces apoptosis however, have not been elucidated. The present study was performed to determine the molecular events that promote apoptosis in andrographolide treated cells using T84, HCT116 and COLO 205 colon cancer cell lines. Andrographolide was determined to limit colony formation and Ki67 expression, alter nuclear morphology, increase cytoplasmic histo...

  16. Growth of SJL/J-derived transplantable reticulum cell sarcoma as related to its ability to induce T-cell proliferation in the host- III. Studies on thymectomized and congenitally athymic SJL mice

    International Nuclear Information System (INIS)

    Katz, I.R.; Chapman-Alexander, J.; Jacobson, E.B.; Lerman, S.P.; Thorbecke, G.J.

    1981-01-01

    When SJL mice are irradiated and reconstituted with syngeneic bone marrow (XBM) they support growth of transplantable reticulum cell sarcoma to approximately 60% of that in normal mice. The ability to support RCS growth gradually improves with time after irradiation and reaches 90% of normal by 8-12 weeks. However, if the mice are thymectomized 4 weeks prior to treatment (Tx-XBM) they initially show 50% which increases to only 65% of growth in normal mice after 12 weeks. The ability of lymphoid cells from these mice to proliferate in vitro in response to irradiated RCS cells is normal 4 weeks after treatment in XBM, but remains <10% of normal in Tx-XBM mice. Nude mice of SJL background also show greatly diminished RCS growth. It is concluded that T cells promote RCS growth in vivo possibly via their tendency to proliferate upon exposure to RCS

  17. Ampelopsin-induced reactive oxygen species enhance the apoptosis of colon cancer cells by activating endoplasmic reticulum stress-mediated AMPK/MAPK/XAF1 signaling

    Science.gov (United States)

    Park, Ga Bin; Jeong, Jee-Yeong; Kim, Daejin

    2017-01-01

    Ampelopsin (Amp) is bioactive natural product and exerts anti-cancer effects against several cancer types. The present study investigated the anti-colon cancer activity of Amp and explored its mechanism of action. The treatment of colon cancer cells with Amp resulted in the dose- and time-dependent induction of apoptosis via the activation of endoplasmic reticulum (ER) stress, 5′ adenosine monophosphate-activated protein kinase (AMPK), and c-Jun N-terminal protein kinase (JNK)/p38 mitogen-activated protein kinases (MAPKs). Salubrinal, an ER stress inhibitor, prevented the upregulation of ER stress-associated proteins, including phosphorylated protein kinase RNA-like ER kinase, phosphorylated eukaryotic translation initiation factor 2α, glucose-regulated protein 78, and CCAAT/enhancer-binding protein homologous protein, as well as suppressing AMPK activation and the MAPK signaling pathway. Knockdown of AMPK by RNA interference failed to block ER stress. Additionally, SP600125 (a JNK inhibitor) and SB203580 (a p38-MAPK inhibitor) effectively inhibited apoptosis and attenuated the expression of X-linked IAP-associated factor 1 (XAF1) and apoptotic Bcl-2 family proteins (BCL2 antagonist/killer 1 and BCL2-associated X protein) in Amp-treated colon cancer cells. Furthermore, reactive oxygen species (ROS)-mediated ER stress/AMPK apoptotic signaling pathway in Amp-treated colon cancer cells were markedly inhibited by treatment with N-acetyl-L-cysteine, a ROS scavenger. These results demonstrate that treatment with Amp induces the apoptotic death of colon cancer cells through ER stress-initiated AMPK/MAPK/XAF1 signaling. These results also provide experimental information for developing Amp as therapeutic drug against colon cancer. PMID:29250183

  18. Calix[6]arene bypasses human pancreatic cancer aggressiveness: downregulation of receptor tyrosine kinases and induction of cell death by reticulum stress and autophagy.

    Science.gov (United States)

    Pelizzaro-Rocha, Karin Juliane; de Jesus, Marcelo Bispo; Ruela-de-Sousa, Roberta Regina; Nakamura, Celso Vataru; Reis, Fabiano Souza; de Fátima, Angelo; Ferreira-Halder, Carmen Veríssima

    2013-12-01

    Pancreatic cancer ranks fourth among cancer-related causes of death in North America. Minimal progress has been made in the diagnosis and treatment of patients with late-stage tumors. Moreover, pancreatic cancer aggressiveness is closely related to high levels of pro-survival mediators, which can ultimately lead to rapid disease progression, resistance and metastasis. The main goal of this study was to define the mechanisms by which calix[6]arene, but not other calixarenes, efficiently decreases the aggressiveness of a drug resistant human pancreas carcinoma cell line (Panc-1). Calix[6]arene was more potent in reducing Panc-1 cell viability than gemcitabine and 5-fluorouracil. In relation to the underlying mechanisms of cytotoxic effects, it led to cell cycle arrest in the G0/G1 phase through downregulation of PIM1, CDK2, CDK4 and retinoblastoma proteins. Importantly, calix[6]arene abolished signal transduction of Mer and AXL tyrosine kinase receptors, both of which are usually overexpressed in pancreatic cancer. Accordingly, inhibition of PI3K and mTOR was also observed, and these proteins are positively modulated by Mer and AXL. Despite decreasing the phosphorylation of AKT at Thr308, calix[6]arene caused an increase in phosphorylation at Ser473. These findings in conjunction with increased BiP and IRE1-α provide a molecular basis explaining the capacity of calix[6]arene to trigger endoplasmic reticulum stress and autophagic cell death. Our findings highlight calix[6]arene as a potential candidate for overcoming pancreatic cancer aggressiveness. Importantly, we provide evidence that calix[6]arene affects a broad array of key targets that are usually dysfunctional in pancreatic cancer, a highly desirable characteristic for chemotherapeutics. © 2013.

  19. Orientia tsutsugamushi ankyrin repeat-containing protein family members are Type 1 secretion system substrates that traffic to the host cell endoplasmic reticulum.

    Science.gov (United States)

    VieBrock, Lauren; Evans, Sean M; Beyer, Andrea R; Larson, Charles L; Beare, Paul A; Ge, Hong; Singh, Smita; Rodino, Kyle G; Heinzen, Robert A; Richards, Allen L; Carlyon, Jason A

    2014-01-01

    Scrub typhus is an understudied, potentially fatal infection that threatens one billion persons in the Asia-Pacific region. How the causative obligate intracellular bacterium, Orientia tsutsugamushi, facilitates its intracellular survival and pathogenesis is poorly understood. Many intracellular bacterial pathogens utilize the Type 1 (T1SS) or Type 4 secretion system (T4SS) to translocate ankyrin repeat-containing proteins (Anks) that traffic to distinct subcellular locations and modulate host cell processes. The O. tsutsugamushi genome encodes one of the largest known bacterial Ank repertoires plus T1SS and T4SS components. Whether these potential virulence factors are expressed during infection, how the Anks are potentially secreted, and to where they localize in the host cell are not known. We determined that O. tsutsugamushi transcriptionally expresses 20 unique ank genes as well as genes for both T1SS and T4SS during infection of mammalian host cells. Examination of the Anks' C-termini revealed that the majority of them resemble T1SS substrates. Escherichia coli expressing a functional T1SS was able to secrete chimeric hemolysin proteins bearing the C-termini of 19 of 20 O. tsutsugamushi Anks in an HlyBD-dependent manner. Thus, O. tsutsugamushi Anks C-termini are T1SS-compatible. Conversely, Coxiella burnetii could not secrete heterologously expressed Anks in a T4SS-dependent manner. Analysis of the subcellular distribution patterns of 20 ectopically expressed Anks revealed that, while 6 remained cytosolic or trafficked to the nucleus, 14 localized to, and in some cases, altered the morphology of the endoplasmic reticulum. This study identifies O. tsutsugamushi Anks as T1SS substrates and indicates that many display a tropism for the host cell secretory pathway.

  20. Radioresistant Spodoptera frugiperda 9 insect cells display excessive resistance to 'endoplasmic reticulum' stress and calcium disturbances via pre-emptive activation of unfolded protein response pathway

    International Nuclear Information System (INIS)

    Guleria, Ayushi; Chandna, Sudhir

    2016-01-01

    Endoplasmic Reticulum (ER) performs multiple cellular functions such as proper folding of newly synthesized proteins and calcium homeostasis. ER stress triggers unfolded protein response (UPR) that attempts to restore normal ER function and resists damage-induced cell death. Lepidopteran Sf9 insect cells (derived from Spodoptera frugiperda) display 100-200 times higher radioresistance than mammalian cells. We have earlier reported that gamma-radiation doses <1000 Gy fail to trigger increase in cytosolic calcium in Sf9 cells, indicating resilience to calcium/ ER disturbances

  1. Ins(1,4,5)P{sub 3} facilitates ATP accumulation via phosphocreatine/creatine kinase in the endoplasmic reticulum extracted from MDCK cells

    Energy Technology Data Exchange (ETDEWEB)

    Sun, Jing [Medical Research Center, School of Medicine, Fukuoka University, Fukuoka 814-0180 (Japan); Department of Dental Implantology, School of Stomatology, Tongji University, Shanghai 200072 (China); Ogata, Shigenori [Joint Laboratory for Frontier Medical Science, School of Medicine, Fukuoka University, Fukuoka 814-0180 (Japan); Segawa, Masaru [Central Laboratory for Pathology and Morphology, School of Medicine, Fukuoka University, Fukuoka 814-0180 (Japan); Usune, Sadaharu [Research Laboratory of Biodynamics, School of Medicine, Fukuoka University, Fukuoka 814-0180 (Japan); Zhao, Yumei [Department of Pediatric Dentistry, School of Dentistry of Shanghai Tongji University, Shanghai 200072 (China); Katsuragi, Takeshi, E-mail: katsurag@fukuoka-u.ac.jp [Medical Research Center, School of Medicine, Fukuoka University, Fukuoka 814-0180 (Japan)

    2010-07-02

    So far, the content and accumulation of ATP in isolated endoplasmic reticulum (ER) are little understood. First, we confirmed using electron microscopic and Western blotting techniques that the samples extracted from MDCK cells are endoplasmic reticulum (ER). The amounts of ATP in the extracted ER were measured from the filtrate after a spinning down of ultrafiltration spin column packed with ER. When the ER sample (5 {mu}g) after 3 days freezing was suspended in intracellular medium (ICM), 0.1% Triton X and ultrapure water (UPW), ATP amounts from the ER with UPW were the highest and over 10 times compared with that from the control with ICM, indicating that UPW is the most effective tool in destroying the ER membrane. After a 10-min-incubation with ICM containing phosphocreatine (PCr)/creatine kinase (CK) of the fresh ER. ATP amounts in the filtrate obtained by spinning down were not changed from that in the control (no PCr/CK). However, ATP amounts in the filtrate from the second spinning down of the ER (treated with PCr/CK) suspended in UPW became over 10-fold compared with the control. When 1 {mu}M inositol(1,4,5)trisphosphate (Ins(1,4,5)P{sub 3}) was added in the incubation medium (ICM with PCr/CK), ATP amounts from the filtrate after the second spinning down were further enhanced around three times. This enhancement was almost canceled by Ca{sup 2+}-removal from ICM and by adding thapsigargin, a Ca{sup 2+}-ATPase inhibitor, but not by 2-APB and heparin, Ins(1,4,5)P{sub 3} receptor antagonists. Administration of 500 {mu}M adenosine to the incubation medium (with PCr/CK) failed to enhance the accumulation of ATP in the ER. These findings suggest that the ER originally contains ATP and ATP accumulation in the ER is promoted by PCr/CK and Ins(1,4,5)P{sub 3}.

  2. Endoplasmic Reticulum-Targeted Subunit Toxins Provide a New Approach to Rescue Misfolded Mutant Proteins and Revert Cell Models of Genetic Diseases.

    Science.gov (United States)

    Adnan, Humaira; Zhang, Zhenbo; Park, Hyun-Joo; Tailor, Chetankumar; Che, Clare; Kamani, Mustafa; Spitalny, George; Binnington, Beth; Lingwood, Clifford

    2016-01-01

    Many germ line diseases stem from a relatively minor disturbance in mutant protein endoplasmic reticulum (ER) 3D assembly. Chaperones are recruited which, on failure to correct folding, sort the mutant for retrotranslocation and cytosolic proteasomal degradation (ER-associated degradation-ERAD), to initiate/exacerbate deficiency-disease symptoms. Several bacterial (and plant) subunit toxins, retrograde transport to the ER after initial cell surface receptor binding/internalization. The A subunit has evolved to mimic a misfolded protein and hijack the ERAD membrane translocon (dislocon), to effect cytosolic access and cytopathology. We show such toxins compete for ERAD to rescue endogenous misfolded proteins. Cholera toxin or verotoxin (Shiga toxin) containing genetically inactivated (± an N-terminal polyleucine tail) A subunit can, within 2-4 hrs, temporarily increase F508delCFTR protein, the major cystic fibrosis (CF) mutant (5-10x), F508delCFTR Golgi maturation (glucocerobrosidase (GCC) in N370SGCC Gaucher Disease fibroblasts (3x), another ERAD-exacerbated misfiling disease. We identify a new, potentially benign approach to the treatment of certain genetic protein misfolding diseases.

  3. Surface Segregation in YSZ

    DEFF Research Database (Denmark)

    Bay, Lasse; Zachau-Christiansen, Birgit; Jacobsen, Torben

    1998-01-01

    The space charge layer formed due to segregation of yttria and oxygen ion vacancies in YSZ is described by a simple model. Effects of impurities segregation are omitted.......The space charge layer formed due to segregation of yttria and oxygen ion vacancies in YSZ is described by a simple model. Effects of impurities segregation are omitted....

  4. Brain-derived neurotrophic factor improves proliferation of endometrial epithelial cells by inhibition of endoplasmic reticulum stress during early pregnancy.

    Science.gov (United States)

    Lim, Whasun; Bae, Hyocheol; Bazer, Fuller W; Song, Gwonhwa

    2017-12-01

    Brain-derived neurotrophic factor (BDNF) is a member of the neurotrophin family binds to two transmembrane receptors; neurotrophic receptor tyrosine kinase 2 (NTRK2) with high affinity and p75 with low affinity. Although BDNF-NTRK2 signaling in the central nervous system is known, signaling in the female reproductive system is unknown. Therefore, we determined effects of BDNF on porcine endometrial luminal epithelial (pLE) cells isolated from Day 12 of pregnancy, as well as expression of BDNF and NTRK2 in endometria of cyclic and pregnant pigs. BDNF-NTRK2 genes were expressed in uterine glandular (GE) and luminal (LE) epithelia during early pregnancy. In addition, their expression in uterine GE and LE decreased with increasing parity of sows. Recombinant BDNF increased proliferation in pLE cells in a dose-dependent, as well as expression of PCNA and Cyclin D1 in nuclei of pLE cells. BDNF also activated phosphorylation of AKT, P70S6K, S6, ERK1/2, JNK, P38 proteins in pLE cells. In addition, cell death resulting from tunicamycin-induced ER stress was prevented when pLE cells were treated with the combination of tunicamycin and BDNF which also decreased cells in the Sub-G 1 phase of the cell cycle. Furthermore, tunicamycin-induced unfolded protein response genes were mostly down-regulated to the basal levels as compared to non-treated pLE cells. Our finding suggests that BDNF acts via NTRK2 to induce development of pLE cells for maintenance of implantation and pregnancy by activating cell signaling via the PI3K and MAPK pathways and by inhibiting ER stress. © 2017 Wiley Periodicals, Inc.

  5. Calcium plays a key role in paraoxon-induced apoptosis in EL4 cells by regulating both endoplasmic reticulum- and mitochondria-associated pathways.

    Science.gov (United States)

    Li, Lan; Du, Yi; Ju, Furong; Ma, Shunxiang; Zhang, Shengxiang

    2016-01-01

    Paraoxon (POX) is one of the most toxic organophosphorus pesticides, but its toxic mechanisms associated with apoptosis remain unclear. The aim of this study was to investigate calcium-associated mechanisms in POX-induced apoptosis in EL4 cells. EL4 cells were exposed to POX for 0-16 h. EGTA was used to chelate Ca(2+ ) in extracellular medium, and heparin and procaine were used to inhibit Ca(2+ )efflux from the endoplasmic reticulum (ER). Z-ATAD-FMK was used to inhibit caspase-12 activity. The apoptotic rate assay, western blotting and immunocytochemistry (ICC) were used to reveal the mechanisms of POX-induced apoptosis. POX significantly increased the expression and activation of caspase-12 and caspase-3, enhanced expression of calpain 1 and calpain 2, and induced the release of cyt c, but did not change the expression of Grp 78. Inhibiting caspase-12 activity alleviated POX-induced upregulation of calpain 1 and caspase-3, promoted POX-induced upregulation of calpain 2, and reduced POX-induced cyt c release, suggesting that there was a cross-talk between the ER-associated pathway and mitochondria-associated apoptotic signals. Attenuating intracellular calcium concentration with EGTA, heparin or procaine decreased POX-induced upregulation of calpain 1, calpain 2, caspase-12 and caspase-3, and reduced POX-induced cyt c release. After pretreatment with EGTA or procaine, POX significantly promoted expression of Grp 78. Calcium played a key role in POX-induced apoptosis in EL4 cells by regulating both ER- and mitochondria-associated pathways. The cross-talk of ER- and mitochondria-associated pathways was accomplished through calcium signal.

  6. Equol Attenuates Atherosclerosis in Apolipoprotein E-Deficient Mice by Inhibiting Endoplasmic Reticulum Stress via Activation of Nrf2 in Endothelial Cells.

    Directory of Open Access Journals (Sweden)

    Ting Zhang

    Full Text Available The development of atherosclerosis is closely related to excessive endoplasmic reticulum stress (ERs. Equol reportedly protects against cardiovascular disease; however, the underlying mechanism for this protection remains unknown. Herein, the mechanisms contributing to the atheroprotective effect of equol were addressed using apolipoprotein E knockout (apoE-/- mice fed a high-fat diet (HFD with or without equol. Equol intervention reduced atherosclerotic lesions in the aorta in HFD-fed apoE-/- mice. Plasma lipid analysis showed that equol intervention reduced triglycerides, total cholesterol and LDL-cholesterol and increased HDL-cholesterol. Additionally, equol administration decreased lipid accumulation in the liver. Simultaneously, equol treatment inhibited cell apoptosis induced by t-BHP and thapsigargin in human umbilical vein endothelial cells (HUVECs. Furthermore, equol treatment attenuated palmitate, t-BHP or thapsigargin-induced upregulation of ER stress markers, including p-PERK, p-eIF2α, GRP78, ATF6 and CHOP proteins expression. The same tendency was also observed in aortic lysates in apoE-/- mice fed with equol plus HFD compared with HFD alone. Moreover, equol treatment dose dependently activated the Nrf2 signaling pathway under oxidative stress. Additionally, elevation of Nrf2 induction was found in aortic lysates in apoE-/- mice fed with a HFD diet containing equol compared with a HFD diet without equol. Importantly, Nrf2 siRNA interference induced CHOP and attenuated the effect of equol to inhibit t-BHP mediated CHOP induction, furthermore, abrogated cell apoptosis induced by t-BHP, suggesting a role for Nrf2 in the protective effect of equol in HUVECs. Collectively, these findings implicate that the improvement of atherosclerosis by equol through attenuation of ER stress is mediated, at least in part, by activating the Nrf2 signaling pathway.

  7. Nitric oxide stress and activation of AMP-activated protein kinase impair β-cell sarcoendoplasmic reticulum calcium ATPase 2b activity and protein stability.

    Science.gov (United States)

    Tong, X; Kono, T; Evans-Molina, C

    2015-06-18

    The sarcoendoplasmic reticulum Ca(2+) ATPase 2b (SERCA2b) pump maintains a steep Ca(2+) concentration gradient between the cytosol and ER lumen in the pancreatic β-cell, and the integrity of this gradient has a central role in regulated insulin production and secretion, maintenance of ER function and β-cell survival. We have previously demonstrated loss of β-cell SERCA2b expression under diabetic conditions. To define the mechanisms underlying this, INS-1 cells and rat islets were treated with the proinflammatory cytokine interleukin-1β (IL-1β) combined with or without cycloheximide or actinomycin D. IL-1β treatment led to increased inducible nitric oxide synthase (iNOS) gene and protein expression, which occurred concurrently with the activation of AMP-activated protein kinase (AMPK). IL-1β led to decreased SERCA2b mRNA and protein expression, whereas time-course experiments revealed a reduction in protein half-life with no change in mRNA stability. Moreover, SERCA2b protein but not mRNA levels were rescued by treatment with the NOS inhibitor l-NMMA (NG-monomethyl L-arginine), whereas the NO donor SNAP (S-nitroso-N-acetyl-D,L-penicillamine) and the AMPK activator AICAR (5-aminoimidazole-4-carboxamide ribonucleotide) recapitulated the effects of IL-1β on SERCA2b protein stability. Similarly, IL-1β-induced reductions in SERCA2b expression were rescued by pharmacological inhibition of AMPK with compound C or by transduction of a dominant-negative form of AMPK, whereas β-cell death was prevented in parallel. Finally, to determine a functional relationship between NO and AMPK signaling and SERCA2b activity, fura-2/AM (fura-2-acetoxymethylester) Ca(2+) imaging experiments were performed in INS-1 cells. Consistent with observed changes in SERCA2b expression, IL-1β, SNAP and AICAR increased cytosolic Ca(2+) and decreased ER Ca(2+) levels, suggesting congruent modulation of SERCA activity under these conditions. In aggregate, these results show that SERCA2b

  8. Endoplasmic reticulum stress causes EBV lytic replication

    OpenAIRE

    Taylor, Gwen Marie; Raghuwanshi, Sandeep K.; Rowe, David T.; Wadowsky, Robert M.; Rosendorff, Adam

    2011-01-01

    Endoplasmic reticulum (ER) stress triggers a homeostatic cellular response in mammalian cells to ensure efficient folding, sorting, and processing of client proteins. In lytic-permissive lymphoblastoid cell lines (LCLs), pulse exposure to the chemical ER-stress inducer thapsigargin (TG) followed by recovery resulted in the activation of the EBV immediate-early (BRLF1, BZLF1), early (BMRF1), and late (gp350) genes, gp350 surface expression, and virus release. The protein phosphatase 1 a (PP1a)...

  9. A sexy spin on nonrandom chromosome segregation.

    Science.gov (United States)

    Charville, Gregory W; Rando, Thomas A

    2013-06-06

    Nonrandom chromosome segregation is an intriguing phenomenon linked to certain asymmetric stem cell divisions. In a recent report in Nature, Yadlapalli and Yamashita (2013) observe nonrandom segregation of X and Y chromosomes in Drosophila germline stem cells and shed light on the complex mechanisms of this fascinating process. Copyright © 2013 Elsevier Inc. All rights reserved.

  10. Pulsed electromagnetic field affects intrinsic and endoplasmatic reticulum apoptosis induction pathways in MonoMac6 cell line culture.

    Science.gov (United States)

    Kaszuba-Zwoinska, J; Chorobik, P; Juszczak, K; Zaraska, W; Thor, P J

    2012-10-01

    Current studies were aimed to elucidate influence of pulsed electromagnetic field stimulation on cell viability and apoptosis induction pathways. For the experimental model we have chosen monocytic cell line MonoMac6 and several apoptosis inducers with different mechanism of death induction like puromycin, colchicine, cyclophosphamide, minocycline and hydrogen peroxide. MonoMac6 cell line was grown at density 1x10(5) cells/well in 96-well culture plates. To induce cell death cell cultures were treated with different apoptosis inducers like puromycin, colchicine, cyclophosphamide, minocycline, hydrogen peroxide and at the same time with pulsed electromagnetic field 50 Hz, 45±5 mT (PEMF) for 4 hour per each stimulation, three times, in 24 hours intervals. Afterwards, cells were harvested for flow cytometry analysis of cell viability measured by annexin V-APC labeled and propidium iodide staining. Expression of apoptosis related genes was evaluated by semi quantitative reverse transcription (RT)-PCR assay. NuPAGE Novex Western blot analysis was carried out for apoptosis inducing factor (AIF) abundance in cytosolic and nuclear extracts of MonoMac6 cells. Puromycin, colchicine and minocycline activated cells and simultaneously treated with PEMF have shown out diminished percentage of annexinV positive (AnV+) cells comparing to controls without PEMF stimulation. MonaMac6 cells puromycin/colchicyne and PEMF treated were to a higher extent double stained (AnV+,PI+), which means increased late apoptotic as well as necrotic (PI+) cells, than non-stimulated controls. On the other hand, minocycline activated cells prior to PEMF treatment showed diminished amount of apoptotic and necrotic (annexin V, annexin V and propidium iodide, propidium iodide positive staining) cells. The opposite effect of PEMF on the percentage of annexin V positively stained cells has been achieved after treatment of MonoMac6 culture with cyclophoshamide and hydrogen peroxide. PEMF enhanced early

  11. Progressive segregation of the Escherichia coli chromosome

    DEFF Research Database (Denmark)

    Nielsen, Henrik Jørck; Youngren, Brenda; Hansen, Flemming G.

    2006-01-01

    We have followed the fate of 14 different loci around the Escherichia coli chromosome in living cells at slow growth rate using a highly efficient labelling system and automated measurements. Loci are segregated as they are replicated, but with a marked delay. Most markers segregate in a smooth...... temporal progression from origin to terminus. Thus, the overall pattern is one of continuous segregation during replication and is not consistent with recently published models invoking extensive sister chromosome cohesion followed by simultaneous segregation of the bulk of the chromosome. The terminus......, and a region immediately clockwise from the origin, are exceptions to the overall pattern and are subjected to a more extensive delay prior to segregation. The origin region and nearby loci are replicated and segregated from the cell centre, later markers from the various positions where they lie...

  12. Myosin XI-Dependent Formation of Tubular Structures from Endoplasmic Reticulum Isolated from Tobacco Cultured BY-2 Cells1[W][OA

    Science.gov (United States)

    Yokota, Etsuo; Ueda, Haruko; Hashimoto, Kohsuke; Orii, Hidefumi; Shimada, Tomoo; Hara-Nishimura, Ikuko; Shimmen, Teruo

    2011-01-01

    The reticular network of the endoplasmic reticulum (ER) consists of tubular and lamellar elements and is arranged in the cortical region of plant cells. This network constantly shows shape change and remodeling motion. Tubular ER structures were formed when GTP was added to the ER vesicles isolated from tobacco (Nicotiana tabacum) cultured BY-2 cells expressing ER-localized green fluorescent protein. The hydrolysis of GTP during ER tubule formation was higher than that under conditions in which ER tubule formation was not induced. Furthermore, a shearing force, such as the flow of liquid, was needed for the elongation/extension of the ER tubule. The shearing force was assumed to correspond to the force generated by the actomyosin system in vivo. To confirm this hypothesis, the S12 fraction was prepared, which contained both cytosol and microsome fractions, including two classes of myosins, XI (175-kD myosin) and VIII (BY-2 myosin VIII-1), and ER-localized green fluorescent protein vesicles. The ER tubules and their mesh-like structures were arranged in the S12 fraction efficiently by the addition of ATP, GTP, and exogenous filamentous actin. The tubule formation was significantly inhibited by the depletion of 175-kD myosin from the S12 fraction but not BY-2 myosin VIII-1. Furthermore, a recombinant carboxyl-terminal tail region of 175-kD myosin also suppressed ER tubule formation. The tips of tubules moved along filamentous actin during tubule elongation. These results indicated that the motive force generated by the actomyosin system contributes to the formation of ER tubules, suggesting that myosin XI is responsible not only for the transport of ER in cytoplasm but also for the reticular organization of cortical ER. PMID:21427277

  13. Acid-Sensing Ion Channel 1a Regulates Fate of Rat Nucleus Pulposus Cells in Acid Stimulus Through Endoplasmic Reticulum Stress

    Directory of Open Access Journals (Sweden)

    Xie Zhi-Yang

    2018-02-01

    Full Text Available Acid-sensing ion channel 1a (ASIC1a participates in human intervertebral disc degeneration (IVDD and regulates the destiny of nucleus pulposus cells (NPCs in acid stimulus. However, the mechanism of ASIC1a activation and its downstream pathway remain unclear. Endoplasmic reticulum (ER stress also participates in the acid-induced apoptosis of NPCs. The main purpose of this study was to investigate whether there is any connection between ASIC1a and ER stress in an acid-induced nucleus pulposus degeneration model. The IVDs of Sprague-Dawley rats were stained by immunohistochemical staining to evaluate the expression of ASIC1a in normal and degenerated rat nucleus pulposus. ASIC1a expression was also quantified by quantitative real-time-polymerase chain reaction and Western blotting analysis. NPCs were exposed to the culture media with acidity at pH 7.2 and 6.5 for 24 h, with or without 4-phenylbutyrate (4-PBA, a blocker of the ER stress pathway. Cell apoptosis was examined by Annexin V/Propidium Iodide (PI staining and was quantified using flow cytometry analysis. ASIC1a-mediated intracellular calcium was determined by Ca2+ imaging using Fura-2-AM. Acidity-induced changes in ER stress markers were studied using Western blotting analysis. In vivo, ASIC1a expression was upregulated in natural degeneration. In vitro, acid stimulus increased intracellular calcium levels, but this effect was blocked by knockdown of ASIC1a, and this reversal was partly inhibited by 4-PBA. In addition, blockade of ASIC1a reduced expression of ER stress markers, especially the proapoptotic markers. ASIC1a partly regulates ER stress and promotes apoptosis of NPCs under acid stimulus and may be a novel therapeutic target in IVDD.

  14. Acid-Sensing Ion Channel 1a Regulates Fate of Rat Nucleus Pulposus Cells in Acid Stimulus Through Endoplasmic Reticulum Stress.

    Science.gov (United States)

    Xie, Zhi-Yang; Chen, Lu; Zhang, Cong; Liu, Lei; Wang, Feng; Cai, Feng; Wang, Xiao-Hu; Shi, Rui; Sinkemani, Arjun; Yu, Hao-Min; Hong, Xin; Wu, Xiao-Tao

    2018-01-01

    Acid-sensing ion channel 1a (ASIC1a) participates in human intervertebral disc degeneration (IVDD) and regulates the destiny of nucleus pulposus cells (NPCs) in acid stimulus. However, the mechanism of ASIC1a activation and its downstream pathway remain unclear. Endoplasmic reticulum (ER) stress also participates in the acid-induced apoptosis of NPCs. The main purpose of this study was to investigate whether there is any connection between ASIC1a and ER stress in an acid-induced nucleus pulposus degeneration model. The IVDs of Sprague-Dawley rats were stained by immunohistochemical staining to evaluate the expression of ASIC1a in normal and degenerated rat nucleus pulposus. ASIC1a expression was also quantified by quantitative real-time-polymerase chain reaction and Western blotting analysis. NPCs were exposed to the culture media with acidity at pH 7.2 and 6.5 for 24 h, with or without 4-phenylbutyrate (4-PBA, a blocker of the ER stress pathway). Cell apoptosis was examined by Annexin V/Propidium Iodide (PI) staining and was quantified using flow cytometry analysis. ASIC1a-mediated intracellular calcium was determined by Ca 2+ imaging using Fura-2-AM. Acidity-induced changes in ER stress markers were studied using Western blotting analysis. In vivo , ASIC1a expression was upregulated in natural degeneration. In vitro , acid stimulus increased intracellular calcium levels, but this effect was blocked by knockdown of ASIC1a, and this reversal was partly inhibited by 4-PBA. In addition, blockade of ASIC1a reduced expression of ER stress markers, especially the proapoptotic markers. ASIC1a partly regulates ER stress and promotes apoptosis of NPCs under acid stimulus and may be a novel therapeutic target in IVDD.

  15. Caffeic acid attenuates the inflammatory stress induced by glycated LDL in human endothelial cells by mechanisms involving inhibition of AGE-receptor, oxidative, and endoplasmic reticulum stress.

    Science.gov (United States)

    Toma, Laura; Sanda, Gabriela M; Niculescu, Loredan S; Deleanu, Mariana; Stancu, Camelia S; Sima, Anca V

    2017-09-10

    Type 2 diabetes mellitus is a worldwide epidemic and its atherosclerotic complications determine the high morbidity and mortality of diabetic patients. Caffeic acid (CAF), a phenolic acid present in normal diets, is known for its antioxidant properties. The aim of this study was to investigate CAF's anti-inflammatory properties and its mechanism of action, using cultured human endothelial cells (HEC) incubated with glycated low-density lipoproteins (gLDL). Levels of the receptor for advanced glycation end-products (RAGE), inflammatory stress markers (C reactive protein, CRP; vascular cell adhesion molecule-1, VCAM-1; monocyte chemoattractant protein-1, MCP-1), and oxidative stress and endoplasmic reticulum stress (ERS) markers were evaluated in gLDL-exposed HEC, in the presence/absence of CAF. RAGE silencing or blocking, specific inhibitors for oxidative stress (apocynin, N-acetyl-cysteine), and ERS (salubrinal) were used. The results showed that: (i) gLDL induced CRP synthesis and secretion through mechanisms involving NADPH oxidase-dependent oxidative stress and ERS in HEC; (ii) gLDL-RAGE interaction, oxidative stress, and ERS stimulated the secretion of VCAM-1 and MCP-1 in HEC; and (iii) CAF reduced the secretion of CRP, VCAM-1, and MCP-1 in gLDL-exposed HEC by inhibiting RAGE expression, oxidative stress, and ERS. In conclusion, CAF might be a promising alternative to ameliorate a wide spectrum of disorders due to its complex mechanisms of action resulting in anti-inflammatory and antioxidative properties. © 2017 BioFactors, 43(5):685-697, 2017. © 2017 International Union of Biochemistry and Molecular Biology.

  16. Dengue Virus Selectively Annexes Endoplasmic Reticulum-Associated Translation Machinery as a Strategy for Co-opting Host Cell Protein Synthesis.

    Science.gov (United States)

    Reid, David W; Campos, Rafael K; Child, Jessica R; Zheng, Tianli; Chan, Kitti Wing Ki; Bradrick, Shelton S; Vasudevan, Subhash G; Garcia-Blanco, Mariano A; Nicchitta, Christopher V

    2018-04-01

    A primary question in dengue virus (DENV) biology is the molecular strategy for recruitment of host cell protein synthesis machinery. Here, we combined cell fractionation, ribosome profiling, and transcriptome sequencing (RNA-seq) to investigate the subcellular organization of viral genome translation and replication as well as host cell translation and its response to DENV infection. We report that throughout the viral life cycle, DENV plus- and minus-strand RNAs were highly partitioned to the endoplasmic reticulum (ER), identifying the ER as the primary site of DENV translation. DENV infection was accompanied by an ER compartment-specific remodeling of translation, where ER translation capacity was subverted from host transcripts to DENV plus-strand RNA, particularly at late stages of infection. Remarkably, translation levels and patterns in the cytosol compartment were only modestly affected throughout the experimental time course of infection. Comparisons of ribosome footprinting densities of the DENV plus-strand RNA and host mRNAs indicated that DENV plus-strand RNA was only sparsely loaded with ribosomes. Combined, these observations suggest a mechanism where ER-localized translation and translational control mechanisms, likely cis encoded, are used to repurpose the ER for DENV virion production. Consistent with this view, we found ER-linked cellular stress response pathways commonly associated with viral infection, namely, the interferon response and unfolded protein response, to be only modestly activated during DENV infection. These data support a model where DENV reprograms the ER protein synthesis and processing environment to promote viral survival and replication while minimizing the activation of antiviral and proteostatic stress response pathways. IMPORTANCE DENV, a prominent human health threat with no broadly effective or specific treatment, depends on host cell translation machinery for viral replication, immune evasion, and virion biogenesis. The

  17. OASIS/CREB3L1 is induced by endoplasmic reticulum stress in human glioma cell lines and contributes to the unfolded protein response, extracellular matrix production and cell migration.

    Directory of Open Access Journals (Sweden)

    Ravi N Vellanki

    Full Text Available OASIS is a transcription factor similar to ATF6 that is activated by endoplasmic reticulum stress. In this study we investigated the expression of OASIS in human glioma cell lines and the effect of OASIS knock-down on the ER stress response and cell migration. OASIS mRNA was detected in three distinct glioma cell lines (U373, A172 and U87 and expression levels were increased upon treatment with ER stress-inducing compounds in the U373 and U87 lines. OASIS protein, which is glycosylated on Asn-513, was detected in the U373 and U87 glioma lines at low levels in control cells and protein expression was induced by ER stress. Knock-down of OASIS in human glioma cell lines resulted in an attenuated unfolded protein response to ER stress (reduced GRP78/BiP and GRP94 induction and decreased expression of chondroitin sulfate proteoglycan extracellular matrix proteins, but induction of the collagen gene Col1a1 was unaffected. Cells in which OASIS was knocked-down exhibited altered cell morphology and reduced cell migration. These results suggest that OASIS is important for the ER stress response and maintenance of some extracellular matrix proteins in human glioma cells.

  18. A mechanism for ParB-dependent waves of ParA, a protein related to DNA segregation during cell division in prokaryotes

    DEFF Research Database (Denmark)

    Hunding, Axel; Gerdes, Kenn; Charbon, Gitte Ebersbach

    2003-01-01

    in an autocatalytic process. We discuss this mechanism in relation to recent models for MinDE oscillations in E.coli and to microtubule degradation in mitosis. The study points to an ancestral role for the presented pattern types in generating bipolarity in prokaryotes and eukaryotes.......Prokaryotic plasmids encode partitioning (par) loci involved in segregation of DNA to daughter cells at cell division. A functional fusion protein consisting of Walker-type ParA ATPase and green fluorescent protein (Gfp) oscillates back and forth within nucleoid regions with a wave period of about...

  19. Inhibition of mitochondria- and endoplasmic reticulum stress-mediated autophagy augments temozolomide-induced apoptosis in glioma cells.

    Directory of Open Access Journals (Sweden)

    Chien-Ju Lin

    Full Text Available Autophagy is a crucial process for cells to maintain homeostasis and survival through degradation of cellular proteins and organelles, including mitochondria and endoplasmic reticula (ER. We previously demonstrated that temozolomide (TMZ, an alkylating agent for brain tumor chemotherapy, induced reactive oxygen species (ROS/extracellular signal-regulated kinase (ERK-mediated autophagy to protect glioma cells from apoptosis. In this study, we investigated the role of mitochondrial damage and ER stress in TMZ-induced cytotoxicity. Mitochondrial depolarization and mitochondrial permeability transition pore (MPTP opening were observed as a prelude to TMZ-induced autophagy, and these were followed by the loss of mitochondrial mass. Electron transport chain (ETC inhibitors, such as rotenone (a complex I inhibitor, sodium azide (a complex IV inhibitor, and oligomycin (a complex V inhibitor, or the MPTP inhibitor, cyclosporine A, decreased mitochondrial damage-mediated autophagy, and therefore increased TMZ-induced apoptosis. TMZ treatment triggered ER stress with increased expression of GADD153 and GRP78 proteins, and deceased pro-caspase 12 protein. ER stress consequently induced autophagy through c-Jun N-terminal kinases (JNK and Ca(2+ signaling pathways. Combination of TMZ with 4-phenylbutyrate (4-PBA, an ER stress inhibitor, augmented TMZ-induced cytotoxicity by inhibiting autophagy. Taken together, our data indicate that TMZ induced autophagy through mitochondrial damage- and ER stress-dependent mechanisms to protect glioma cells. This study provides evidence that agents targeting mitochondria or ER may be potential anticancer strategies.

  20. Cardiac sarcoplasmic reticulum

    International Nuclear Information System (INIS)

    Jacobson, M.S.; Ambudkar, I.S.; Young, E.P.; Naseem, S.M.; Heald, F.P.; Shamoo, A.E.

    1985-01-01

    The effect on the cardiac sarcoplasmic reticulum of an atherogenic (1% cholesterol) diet fed during the neonatal vs the juvenile period of life was studied in Yorkshire swine. Male piglets were randomly assigned at birth to 1 of 4 groups: group I (control), group II (lactation feeding), group III (juvenile period feeding) and group IV (lactation and juvenile feeding). All animals were killed at 55 weeks of age and cardiac sarcoplasmic reticulum (SR) isolated for assay of calcium uptake, Ca 2+ -Mg 2+ ATPase activity, and lipid analysis by thin-layer chromatography and gas chromatography. The amount of cholesterol/mg SR protein and the cholesterol/phospholipid ratio were higher in the animals fed during lactation (groups II and IV) and lower in those fed only during the juvenile period (group III). Phospholipid fatty acid patterns as measured by gas chromatography were unaltered in any group. Calcium uptake was markedly diminished in all experimental conditions: group II 47%, group III 65% and group IV 96%. Compared to the observed changes in calcium transport, the ATP hydrolytic activity was relatively less affected. Only in group IV a significant decrease (41%) was seen. Groups II and III show no change in ATP hydrolytic activity. The decrease in calcium uptake and altered cholesterol/phospholipid ratio without effect on ATP hydrolytic activity is consistent with an uncoupling of calcium transport related to the atherogenic diet in early life. (author)

  1. Disturbed mitotic progression and genome segregation are involved in cell transformation mediated by nano-TiO2 long-term exposure

    International Nuclear Information System (INIS)

    Huang Shing; Chueh Pinju; Lin Yunwei; Shih Tungsheng; Chuang Showmei

    2009-01-01

    Titanium dioxide (TiO2) nano-particles (< 100 nm in diameter) have been of interest in a wide range of applications, such as in cosmetics and pharmaceuticals because of their low toxicity. However, recent studies have shown that TiO2 nano-particles (nano-TiO2) induce cytotoxicity and genotoxicity in various lines of cultured cells as well as tumorigenesis in animal models. The biological roles of nano-TiO2 are shown to be controversial and no comprehensive study paradigm has been developed to investigate their molecular mechanisms. In this study, we showed that short-term exposure to nano-TiO2 enhanced cell proliferation, survival, ERK signaling activation and ROS production in cultured fibroblast cells. Moreover, long-term exposure to nano-TiO2 not only increased cell survival and growth on soft agar but also the numbers of multinucleated cells and micronucleus (MN) as suggested in confocal microscopy analysis. Cell cycle phase analysis showed G2/M delay and slower cell division in long-term exposed cells. Most importantly, long-term TiO2 exposure remarkably affected mitotic progression at anaphase and telophase leading to aberrant multipolar spindles and chromatin alignment/segregation. Moreover, PLK1 was demonstrated as the target for nano-TiO2 in the regulation of mitotic progression and exit. Notably, a higher fraction of sub-G1 phase population appeared in TiO2-exposed cells after releasing from G2/M synchronization. Our results demonstrate that long-term exposure to nano-TiO2 disturbs cell cycle progression and duplicated genome segregation, leading to chromosomal instability and cell transformation.

  2. Phenylbutyric acid rescues endoplasmic reticulum stress-induced suppression of APP proteolysis and prevents apoptosis in neuronal cells.

    Directory of Open Access Journals (Sweden)

    Jesse C Wiley

    Full Text Available BACKGROUND: The familial and sporadic forms of Alzheimer's disease (AD have an identical pathology with a severe disparity in the time of onset [1]. The pathological similarity suggests that epigenetic processes may phenocopy the Familial Alzheimer's disease (FAD mutations within sporadic AD. Numerous groups have demonstrated that FAD mutations in presenilin result in 'loss of function' of gamma-secretase mediated APP cleavage [2], [3], [4], [5]. Accordingly, ER stress is prominent within the pathologically impacted brain regions in AD patients [6] and is reported to inhibit APP trafficking through the secretory pathway [7], [8]. As the maturation of APP and the cleaving secretases requires trafficking through the secretory pathway [9], [10], [11], we hypothesized that ER stress may block trafficking requisite for normal levels of APP cleavage and that the small molecular chaperone 4-phenylbutyrate (PBA may rescue the proteolytic deficit. METHODOLOGY/PRINCIPAL FINDINGS: The APP-Gal4VP16/Gal4-reporter screen was stably incorporated into neuroblastoma cells in order to assay gamma-secretase mediated APP proteolysis under normal and pharmacologically induced ER stress conditions. Three unrelated pharmacological agents (tunicamycin, thapsigargin and brefeldin A all repressed APP proteolysis in parallel with activation of unfolded protein response (UPR signaling-a biochemical marker of ER stress. Co-treatment of the gamma-secretase reporter cells with PBA blocked the repressive effects of tunicamycin and thapsigargin upon APP proteolysis, UPR activation, and apoptosis. In unstressed cells, PBA stimulated gamma-secretase mediated cleavage of APP by 8-10 fold, in the absence of any significant effects upon amyloid production, by promoting APP trafficking through the secretory pathway and the stimulation of the non-pathogenic alpha/gamma-cleavage. CONCLUSIONS/SIGNIFICANCE: ER stress represses gamma-secretase mediated APP proteolysis, which replicates

  3. Intracellular Ca(2+) release from endoplasmic reticulum regulates slow wave currents and pacemaker activity of interstitial cells of Cajal.

    Science.gov (United States)

    Zhu, Mei Hong; Sung, Tae Sik; O'Driscoll, Kate; Koh, Sang Don; Sanders, Kenton M

    2015-04-15

    Interstitial cells of Cajal (ICC) provide pacemaker activity in gastrointestinal muscles that underlies segmental and peristaltic contractions. ICC generate electrical slow waves that are due to large-amplitude inward currents resulting from anoctamin 1 (ANO1) channels, which are Ca(2+)-activated Cl(-) channels. We investigated the hypothesis that the Ca(2+) responsible for the stochastic activation of ANO1 channels during spontaneous transient inward currents (STICs) and synchronized activation of ANO1 channels during slow wave currents comes from intracellular Ca(2+) stores. ICC, obtained from the small intestine of Kit(+/copGFP) mice, were studied under voltage and current clamp to determine the effects of blocking Ca(2+) uptake into stores and release of Ca(2+) via inositol 1,4,5-trisphosphate (IP3)-dependent and ryanodine-sensitive channels. Cyclocpiazonic acid, thapsigargin, 2-APB, and xestospongin C inhibited STICs and slow wave currents. Ryanodine and tetracaine also inhibited STICs and slow wave currents. Store-active compounds had no direct effects on ANO1 channels expressed in human embryonic kidney-293 cells. Under current clamp, store-active drugs caused significant depolarization of ICC and reduced spontaneous transient depolarizations (STDs). After block of ryanodine receptors with ryanodine and tetracaine, repolarization did not restore STDs. ANO1 expressed in ICC has limited access to cytoplasmic Ca(2+) concentration, suggesting that pacemaker activity depends on Ca(2+) dynamics in restricted microdomains. Our data from studies of isolated ICC differ somewhat from studies on intact muscles and suggest that release of Ca(2+) from both IP3 and ryanodine receptors is important in generating pacemaker activity in ICC. Copyright © 2015 the American Physiological Society.

  4. Intracellular Ca2+ release from endoplasmic reticulum regulates slow wave currents and pacemaker activity of interstitial cells of Cajal

    Science.gov (United States)

    Zhu, Mei Hong; Sung, Tae Sik; O'Driscoll, Kate; Koh, Sang Don

    2015-01-01

    Interstitial cells of Cajal (ICC) provide pacemaker activity in gastrointestinal muscles that underlies segmental and peristaltic contractions. ICC generate electrical slow waves that are due to large-amplitude inward currents resulting from anoctamin 1 (ANO1) channels, which are Ca2+-activated Cl− channels. We investigated the hypothesis that the Ca2+ responsible for the stochastic activation of ANO1 channels during spontaneous transient inward currents (STICs) and synchronized activation of ANO1 channels during slow wave currents comes from intracellular Ca2+ stores. ICC, obtained from the small intestine of Kit+/copGFP mice, were studied under voltage and current clamp to determine the effects of blocking Ca2+ uptake into stores and release of Ca2+ via inositol 1,4,5-trisphosphate (IP3)-dependent and ryanodine-sensitive channels. Cyclocpiazonic acid, thapsigargin, 2-APB, and xestospongin C inhibited STICs and slow wave currents. Ryanodine and tetracaine also inhibited STICs and slow wave currents. Store-active compounds had no direct effects on ANO1 channels expressed in human embryonic kidney-293 cells. Under current clamp, store-active drugs caused significant depolarization of ICC and reduced spontaneous transient depolarizations (STDs). After block of ryanodine receptors with ryanodine and tetracaine, repolarization did not restore STDs. ANO1 expressed in ICC has limited access to cytoplasmic Ca2+ concentration, suggesting that pacemaker activity depends on Ca2+ dynamics in restricted microdomains. Our data from studies of isolated ICC differ somewhat from studies on intact muscles and suggest that release of Ca2+ from both IP3 and ryanodine receptors is important in generating pacemaker activity in ICC. PMID:25631870

  5. When the genome plays dice: circumvention of the spindle assembly checkpoint and near-random chromosome segregation in multipolar cancer cell mitoses.

    Science.gov (United States)

    Gisselsson, David; Håkanson, Ulf; Stoller, Patrick; Marti, Dominik; Jin, Yuesheng; Rosengren, Anders H; Stewénius, Ylva; Kahl, Fredrik; Panagopoulos, Ioannis

    2008-04-02

    Normal cell division is coordinated by a bipolar mitotic spindle, ensuring symmetrical segregation of chromosomes. Cancer cells, however, occasionally divide into three or more directions. Such multipolar mitoses have been proposed to generate genetic diversity and thereby contribute to clonal evolution. However, this notion has been little validated experimentally. Chromosome segregation and DNA content in daughter cells from multipolar mitoses were assessed by multiphoton cross sectioning and fluorescence in situ hybridization in cancer cells and non-neoplastic transformed cells. The DNA distribution resulting from multipolar cell division was found to be highly variable, with frequent nullisomies in the daughter cells. Time-lapse imaging of H2B/GFP-labelled multipolar mitoses revealed that the time from the initiation of metaphase to the beginning of anaphase was prolonged and that the metaphase plates often switched polarity several times before metaphase-anaphase transition. The multipolar metaphase-anaphase transition was accompanied by a normal reduction of cellular cyclin B levels, but typically occurred before completion of the normal separase activity cycle. Centromeric AURKB and MAD2 foci were observed frequently to remain on the centromeres of multipolar ana-telophase chromosomes, indicating that multipolar mitoses were able to circumvent the spindle assembly checkpoint with some sister chromatids remaining unseparated after anaphase. Accordingly, scoring the distribution of individual chromosomes in multipolar daughter nuclei revealed a high frequency of nondisjunction events, resulting in a near-binomial allotment of sister chromatids to the daughter cells. The capability of multipolar mitoses to circumvent the spindle assembly checkpoint system typically results in a near-random distribution of chromosomes to daughter cells. Spindle multipolarity could thus be a highly efficient generator of genetically diverse minority clones in transformed cell

  6. When the genome plays dice: circumvention of the spindle assembly checkpoint and near-random chromosome segregation in multipolar cancer cell mitoses.

    Directory of Open Access Journals (Sweden)

    David Gisselsson

    Full Text Available BACKGROUND: Normal cell division is coordinated by a bipolar mitotic spindle, ensuring symmetrical segregation of chromosomes. Cancer cells, however, occasionally divide into three or more directions. Such multipolar mitoses have been proposed to generate genetic diversity and thereby contribute to clonal evolution. However, this notion has been little validated experimentally. PRINCIPAL FINDINGS: Chromosome segregation and DNA content in daughter cells from multipolar mitoses were assessed by multiphoton cross sectioning and fluorescence in situ hybridization in cancer cells and non-neoplastic transformed cells. The DNA distribution resulting from multipolar cell division was found to be highly variable, with frequent nullisomies in the daughter cells. Time-lapse imaging of H2B/GFP-labelled multipolar mitoses revealed that the time from the initiation of metaphase to the beginning of anaphase was prolonged and that the metaphase plates often switched polarity several times before metaphase-anaphase transition. The multipolar metaphase-anaphase transition was accompanied by a normal reduction of cellular cyclin B levels, but typically occurred before completion of the normal separase activity cycle. Centromeric AURKB and MAD2 foci were observed frequently to remain on the centromeres of multipolar ana-telophase chromosomes, indicating that multipolar mitoses were able to circumvent the spindle assembly checkpoint with some sister chromatids remaining unseparated after anaphase. Accordingly, scoring the distribution of individual chromosomes in multipolar daughter nuclei revealed a high frequency of nondisjunction events, resulting in a near-binomial allotment of sister chromatids to the daughter cells. CONCLUSION: The capability of multipolar mitoses to circumvent the spindle assembly checkpoint system typically results in a near-random distribution of chromosomes to daughter cells. Spindle multipolarity could thus be a highly efficient

  7. Gender Segregation Small Firms

    OpenAIRE

    Kenneth R Troske; William J Carrington

    1992-01-01

    This paper studies interfirm gender segregation in a unique sample of small employers. We focus on small firms because previous research on interfirm segregation has studied only large firms and because it is easier to link the demographic characteristics of employers and employees in small firms. This latter feature permits an assessment of the role of employer discrimination in creating gender segregation. Our first finding is that interfirm segregation is prevalent among small employers. I...

  8. Differential expression of conserved germ line markers and delayed segregation of male and female primordial germ cells in a hermaphrodite, the leech helobdella.

    Science.gov (United States)

    Cho, Sung-Jin; Vallès, Yvonne; Weisblat, David A

    2014-02-01

    In sexually reproducing animals, primordial germ cells (PGCs) are often set aside early in embryogenesis, a strategy that minimizes the risk of genomic damage associated with replication and mitosis during the cell cycle. Here, we have used germ line markers (piwi, vasa, and nanos) and microinjected cell lineage tracers to show that PGC specification in the leech genus Helobdella follows a different scenario: in this hermaphrodite, the male and female PGCs segregate from somatic lineages only after more than 20 rounds of zygotic mitosis; the male and female PGCs share the same (mesodermal) cell lineage for 19 rounds of zygotic mitosis. Moreover, while all three markers are expressed in both male and female reproductive tissues of the adult, they are expressed differentially between the male and female PGCs of the developing embryo: piwi and vasa are expressed preferentially in female PGCs at a time when nanos is expressed preferentially in male PGCs. A priori, the delayed segregation of male and female PGCs from somatic tissues and from one another increases the probability of mutations affecting both male and female PGCs of a given individual. We speculate that this suite of features, combined with a capacity for self-fertilization, may contribute to the dramatically rearranged genome of Helobdella robusta relative to other animals.

  9. Multipolar spindle pole coalescence is a major source of kinetochore mis-attachment and chromosome mis-segregation in cancer cells.

    Directory of Open Access Journals (Sweden)

    William T Silkworth

    Full Text Available Many cancer cells display a CIN (Chromosome Instability phenotype, by which they exhibit high rates of chromosome loss or gain at each cell cycle. Over the years, a number of different mechanisms, including mitotic spindle multipolarity, cytokinesis failure, and merotelic kinetochore orientation, have been proposed as causes of CIN. However, a comprehensive theory of how CIN is perpetuated is still lacking. We used CIN colorectal cancer cells as a model system to investigate the possible cellular mechanism(s underlying CIN. We found that CIN cells frequently assembled multipolar spindles in early mitosis. However, multipolar anaphase cells were very rare, and live-cell experiments showed that almost all CIN cells divided in a bipolar fashion. Moreover, fixed-cell analysis showed high frequencies of merotelically attached lagging chromosomes in bipolar anaphase CIN cells, and higher frequencies of merotelic attachments in multipolar vs. bipolar prometaphases. Finally, we found that multipolar CIN prometaphases typically possessed gamma-tubulin at all spindle poles, and that a significant fraction of bipolar metaphase/early anaphase CIN cells possessed more than one centrosome at a single spindle pole. Taken together, our data suggest a model by which merotelic kinetochore attachments can easily be established in multipolar prometaphases. Most of these multipolar prometaphase cells would then bi-polarize before anaphase onset, and the residual merotelic attachments would produce chromosome mis-segregation due to anaphase lagging chromosomes. We propose this spindle pole coalescence mechanism as a major contributor to chromosome instability in cancer cells.

  10. Deubiquitinase inhibitor b-AP15 activates endoplasmic reticulum (ER) stress and inhibits Wnt/Notch1 signaling pathway leading to the reduction of cell survival in hepatocellular carcinoma cells.

    Science.gov (United States)

    Ding, Youming; Chen, Xiaoyan; Wang, Bin; Yu, Bin; Ge, Jianhui

    2018-04-15

    b-AP15, a potent and selective inhibitor of the ubiquitin-specific peptidase 14 (USP14), displays in vitro and in vivo antitumor abilities on some types of cancer cells. However, the mechanism underlying its action is not well elucidated. The purposes of the present study are to observe the potential impacts of b-AP15 on cell survival of hepatocellular carcinoma cells and to investigate whether and how this compound inhibits some survival-promoting signaling pathways. We found that b-AP15 significantly decreased cell viability and increased cell apoptosis in a dose-dependent manner in hepatocellular carcinoma cells, along with the perturbation of cell cycle and the decreased expressions of cell cycle-related proteins. We also demonstrated that the endoplasmic reticulum (ER) stress and the unfolded protein response (UPR) were enhanced by b-AP15 supplementation. The inhibition of ER stress/UPR only partly attenuated the cytotoxicity of b-AP15 on hepatocellular carcinoma cells. In addition, b-AP15 treatment inhibited Wnt/β-catenin and Notch1 signaling pathways, and suppressed phosphorylation of STAT3, Akt, and Erk1/2, which were not restored by the inhibition of ER stress/UPR. Furthermore, the expression levels of signaling molecules in Notch1 were reduced by specific inhibitor of Wnt/β-catenin pathway. Notably, either Wnt or Notch1 signaling inhibitor mitigated phosphorylation of STAT3, Akt, and Erk1/2, and mimicked the cytotoxicity of b-AP15 on hepatocellular carcinoma cells. These results clearly indicate that b-AP15 induced cytotoxic response to hepatocellular carcinoma cells by augmenting ER stress/UPR and inhibiting Wnt/Notch1 signaling pathways. This new finding provides a novel mechanism by which b-AP15 produces its antitumor therapeutic effects. Copyright © 2018 Elsevier B.V. All rights reserved.

  11. Cucurbita ficifolia Bouché increases insulin secretion in RINm5F cells through an influx of Ca(2+) from the endoplasmic reticulum.

    Science.gov (United States)

    Miranda-Perez, Maria Elizabeth; Ortega-Camarillo, Clara; Del Carmen Escobar-Villanueva, Maria; Blancas-Flores, Gerardo; Alarcon-Aguilar, Francisco Javier

    2016-07-21

    Cucurbita ficifolia Bouché(C. ficifolia) is a plant used in Mexican traditional medicine to control type 2 diabetes (T2D). The hypoglycemic effect of the fruit of C. ficifolia has been demonstrated in different experimental models and in T2D patients. It has been proposed that D-chiro-inositol (DCI) is the active compound of the fruit. Additionally, it has been reported that C. ficifolia increases the mRNA expression of insulin and Kir 6.2 (a component of the ATP-sensitive potassium (K(+)ATP) channel, which is activated by sulphonylurea) in RINm5F cells. However, it remains unclear whether C. ficifolia and DCI causes the secretion of insulin by increasing the concentration of intracellular calcium ([Ca(2+)]i) through K(+)ATP channel blockage or from the reservoir in the endoplasmic reticulum (ER). The aqueous extract of C. ficifolia was obtained and standardized with regard to its DCI content. RINm5F pancreatic β-cells were incubated with different concentrations (50, 100, 200 and 400μM) of DCI alone or C. ficifolia (9, 18, 36 and 72µg of extract/mL), and the [Ca(2+)]i of the cells was quantified. The cells were preloaded with the Ca(2+) fluorescent dye fluo4-acetoxymethyl ester (AM) and visualized by confocal microscopy. Insulin secretion was measured by an ELISA method. Subsequently, the effect of C. ficifolia on the K(+)ATP channel was evaluated. In this case, the blocker activator diazoxide was used to inhibit the C. ficifolia-induced calcium influx. In addition, the inositol 1,4,5-trisphosphate (IP3)-receptor-selective inhibitor 2-amino-thoxydiphenylborate (2-APB) was used to inhibit the influx of calcium from the ER that was induced by C. ficifolia. It was found that DCI alone did not increase [Ca(2+)]i or insulin secretion. In contrast, treatment with C. ficifolia increased [Ca(2+)]i 10-fold compared with the control group. Insulin secretion increased by 46.9%. In the presence of diazoxide, C. ficifolia decreased [Ca(2+)]i by 50%, while insulin secretion

  12. Nonstructural NSs protein of rift valley fever virus interacts with pericentromeric DNA sequences of the host cell, inducing chromosome cohesion and segregation defects.

    Science.gov (United States)

    Mansuroglu, Z; Josse, T; Gilleron, J; Billecocq, A; Leger, P; Bouloy, M; Bonnefoy, E

    2010-01-01

    Rift Valley fever virus (RVFV) is an emerging, highly pathogenic virus; RVFV infection can lead to encephalitis, retinitis, or fatal hepatitis associated with hemorrhagic fever in humans, as well as death, abortions, and fetal deformities in animals. RVFV nonstructural NSs protein, a major factor of the virulence, forms filamentous structures in the nuclei of infected cells. In order to further understand RVFV pathology, we investigated, by chromatin immunoprecipitation, immunofluorescence, fluorescence in situ hybridization, and confocal microscopy, the capacity of NSs to interact with the host genome. Our results demonstrate that even though cellular DNA is predominantly excluded from NSs filaments, NSs interacts with some specific DNA regions of the host genome such as clusters of pericentromeric gamma-satellite sequence. Targeting of these sequences by NSs was correlated with the induction of chromosome cohesion and segregation defects in RVFV-infected murine, as well as sheep cells. Using recombinant nonpathogenic virus rZHDeltaNSs210-230, expressing a NSs protein deleted of its region of interaction with cellular factor SAP30, we showed that the NSs-SAP30 interaction was essential for NSs to target pericentromeric sequences, as well as for induction of chromosome segregation defects. The effect of RVFV upon the inheritance of genetic information is discussed with respect to the pathology associated with fetal deformities and abortions, highlighting the main role played by cellular cofactor SAP30 on the establishment of NSs interactions with host DNA sequences and RVFV pathogenesis.

  13. Cytotoxicity of CdTe quantum dots in human umbilical vein endothelial cells: the involvement of cellular uptake and induction of pro-apoptotic endoplasmic reticulum stress

    Directory of Open Access Journals (Sweden)

    Yan M

    2016-02-01

    Full Text Available Ming Yan,1,* Yun Zhang,2,* Haiyan Qin,3 Kezhou Liu,1 Miao Guo,1 Yakun Ge,1 Mingen Xu,1 Yonghong Sun,4 Xiaoxiang Zheng4 1Department of Biomedical Engineering, College of Life Information Science and Instrument Engineering, Hangzhou Dianzi University, Hangzhou, 2Basic Medical Sciences, College of Medicine, Shaoxing University, Shaoxing, 3Department of Chemistry, Zhejiang University, 4Zhejiang Provincial Key Laboratory of Cardio-Cerebral Vascular Detection Technology and Medicinal Effectiveness Appraisal, Department of Biomedical Engineering, Zhejiang University, Hangzhou, People’s Republic of China *These authors contributed equally to this work Abstract: Cadmium telluride quantum dots (CdTe QDs have been proposed to induce oxidative stress, which plays a crucial role in CdTe QDs-mediated mitochondrial-dependent apoptosis in human umbilical vein endothelial cells (HUVECs. However, the direct interactions of CdTe QDs with HUVECs and their potential impairment of other organelles like endoplasmic reticulum (ER in HUVECs are poorly understood. In this study, we reported that the negatively charged CdTe QDs (–21.63±0.91 mV, with good dispersity and fluorescence stability, were rapidly internalized via endocytosis by HUVECs, as the notable internalization could be inhibited up to 95.52% by energy depletion (NaN3/deoxyglucose or low temperature. The endocytosis inhibitors (methyl-β-cyclodextrin, genistein, sucrose, chlorpromazine, and colchicine dramatically decreased the uptake of CdTe QDs by HUVECs, suggesting that both caveolae/raft- and clathrin-mediated endocytosis were involved in the endothelial uptake of CdTe QDs. Using immunocytochemistry, a striking overlap of the internalized CdTe QDs and ER marker was observed, which indicates that QDs may be transported to ER. The CdTe QDs also caused remarkable ER stress responses in HUVECs, confirmed by significant dilatation of ER cisternae, upregulation of ER stress markers GRP78/GRP94, and

  14. Endoplasmic reticulum stress and diabetic retinopathy

    Directory of Open Access Journals (Sweden)

    Toshiyuki Oshitari

    2008-02-01

    Full Text Available Toshiyuki Oshitari1,2, Natsuyo Hata1, Shuichi Yamamoto11Department of Ophthalmology and Visual Science, Chiba University Graduate School of Medicine, Chiba City, Chiba, Japan; 2Department of Ophthalmology, Kimitsu Central Hospital, Kisarazu City, Chiba, JapanAbstract: Endoplasmic reticulum (ER stress is involved in the pathogenesis of several diseases including Alzheimer disease and Parkinson disease. Many recent studies have shown that ER stress is related to the pathogenesis of diabetes mellitus, and with the death of pancreatic β-cells, insulin resistance, and the death of the vascular cells in the retina. Diabetic retinopathy is a major complication of diabetes and results in death of both neural and vascular cells. Because the death of the neurons directly affects visual function, the precise mechanism causing the death of neurons in early diabetic retinopathy must be determined. The ideal therapy for preventing the onset and the progression of diabetic retinopathy would be to treat the factors involved with both the vascular and neuronal abnormalities in diabetic retinopathy. In this review, we present evidence that ER stress is involved in the death of both retinal neurons and vascular cells in diabetic eyes, and thus reducing or blocking ER stress may be a potential therapy for preventing the onset and the progression of diabetic retinopathy.Keywords: endoplasmic reticulum stress, diabetic retinopathy, vascular cell death, neuronal cell death

  15. Prediction of endoplasmic reticulum resident proteins using fragmented amino acid composition and support vector machine

    Directory of Open Access Journals (Sweden)

    Ravindra Kumar

    2017-09-01

    Full Text Available Background The endoplasmic reticulum plays an important role in many cellular processes, which includes protein synthesis, folding and post-translational processing of newly synthesized proteins. It is also the site for quality control of misfolded proteins and entry point of extracellular proteins to the secretory pathway. Hence at any given point of time, endoplasmic reticulum contains two different cohorts of proteins, (i proteins involved in endoplasmic reticulum-specific function, which reside in the lumen of the endoplasmic reticulum, called as endoplasmic reticulum resident proteins and (ii proteins which are in process of moving to the extracellular space. Thus, endoplasmic reticulum resident proteins must somehow be distinguished from newly synthesized secretory proteins, which pass through the endoplasmic reticulum on their way out of the cell. Approximately only 50% of the proteins used in this study as training data had endoplasmic reticulum retention signal, which shows that these signals are not essentially present in all endoplasmic reticulum resident proteins. This also strongly indicates the role of additional factors in retention of endoplasmic reticulum-specific proteins inside the endoplasmic reticulum. Methods This is a support vector machine based method, where we had used different forms of protein features as inputs for support vector machine to develop the prediction models. During training leave-one-out approach of cross-validation was used. Maximum performance was obtained with a combination of amino acid compositions of different part of proteins. Results In this study, we have reported a novel support vector machine based method for predicting endoplasmic reticulum resident proteins, named as ERPred. During training we achieved a maximum accuracy of 81.42% with leave-one-out approach of cross-validation. When evaluated on independent dataset, ERPred did prediction with sensitivity of 72.31% and specificity of 83

  16. Exposure to submicron particles (PM1.0) from diesel exhaust and pollen allergens of human lung epithelial cells induces morphological changes of mitochondria tonifilaments and rough endoplasmic reticulum.

    Science.gov (United States)

    Mazzarella, Gennaro; Lucariello, Angela; Bianco, Andrea; Calabrese, Cecilia; Thanassoulas, Theodoros; Savarese, Leonilde; Fiumarella, Angelamaria; Esposito, Vincenzo; DE Luca, Antonio

    2014-01-01

    In recent literature, little has been said regarding the morphological changes that occur in lung cells after treatment with particles and nanoparticles. Using an in vitro model of type-II lung epithelium (A549), we studied the effects of submicron particles (PM1.0), Parietaria officinalis (ALL), and PM1.0 + ALL together. To date several biochemical effects have been described, instead few data exist in literature regarding morphological events following these treatments, in particular we focused on the morphological changes and distribution of mitochondria, tonifilaments and rough endoplasmic reticulum, using a transmission electron microscopic (TEM) approach. After exposure to PM1.0 particles (PM1.0), Parietaria officinalis as allergen, and PM1.0 with P. officinalis, changes in the cytoplasmic area were observed, such as damage to mitochondria and morphological alterations of the tonifilaments and rough endoplasmic reticulum. The data obtained strongly support the hypothesis that cells in contact with submicron particles (PM1.0), or P. officinalis, undergo alteration of their metabolism. Copyright © 2014 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  17. Dysfunctional MreB inhibits chromosome segregation in Escherichia coli

    DEFF Research Database (Denmark)

    Kruse, Thomas; Møller-Jensen, Jakob; Løbner-Olesen, Anders

    2003-01-01

    The mechanism of prokaryotic chromosome segregation is not known. MreB, an actin homolog, is a shape-determining factor in rod-shaped prokaryotic cells. Using immunofluorescence microscopy we found that MreB of Escherichia coli formed helical filaments located beneath the cell surface. Flow...... cytometric and cytological analyses indicated that MreB-depleted cells segregated their chromosomes in pairs, consistent with chromosome cohesion. Overexpression of wild-type MreB inhibited cell division but did not perturb chromosome segregation. Overexpression of mutant forms of MreB inhibited cell...... that MreB filaments participate in directional chromosome movement and segregation....

  18. The role of the endoplasmic reticulum stress response following cerebral ischemia.

    Science.gov (United States)

    Hadley, Gina; Neuhaus, Ain A; Couch, Yvonne; Beard, Daniel J; Adriaanse, Bryan A; Vekrellis, Kostas; DeLuca, Gabriele C; Papadakis, Michalis; Sutherland, Brad A; Buchan, Alastair M

    2018-06-01

    Background Cornu ammonis 3 (CA3) hippocampal neurons are resistant to global ischemia, whereas cornu ammonis (CA1) 1 neurons are vulnerable. Hamartin expression in CA3 neurons mediates this endogenous resistance via productive autophagy. Neurons lacking hamartin demonstrate exacerbated endoplasmic reticulum stress and increased cell death. We investigated endoplasmic reticulum stress responses in CA1 and CA3 regions following global cerebral ischemia, and whether pharmacological modulation of endoplasmic reticulum stress or autophagy altered neuronal viability . Methods In vivo: male Wistar rats underwent sham or 10 min of transient global cerebral ischemia. CA1 and CA3 areas were microdissected and endoplasmic reticulum stress protein expression quantified at 3 h and 12 h of reperfusion. In vitro: primary neuronal cultures (E18 Wistar rat embryos) were exposed to 2 h of oxygen and glucose deprivation or normoxia in the presence of an endoplasmic reticulum stress inducer (thapsigargin or tunicamycin), an endoplasmic reticulum stress inhibitor (salubrinal or 4-phenylbutyric acid), an autophagy inducer ([4'-(N-diethylamino) butyl]-2-chlorophenoxazine (10-NCP)) or autophagy inhibitor (3-methyladenine). Results In vivo, decreased endoplasmic reticulum stress protein expression (phospho-eIF2α and ATF4) was observed at 3 h of reperfusion in CA3 neurons following ischemia, and increased in CA1 neurons at 12 h of reperfusion. In vitro, endoplasmic reticulum stress inducers and high doses of the endoplasmic reticulum stress inhibitors also increased cell death. Both induction and inhibition of autophagy also increased cell death. Conclusion Endoplasmic reticulum stress is associated with neuronal cell death following ischemia. Neither reduction of endoplasmic reticulum stress nor induction of autophagy demonstrated neuroprotection in vitro, highlighting their complex role in neuronal biology following ischemia.

  19. Segregation and civic virtue

    NARCIS (Netherlands)

    Merry, M.S.

    2012-01-01

    In this essay Michael Merry defends the following prima facie argument: that civic virtue is not dependent on integration and in fact may be best fostered under conditions of segregation. He demonstrates that civic virtue can and does take place under conditions of involuntary segregation, but that

  20. The nairovirus nairobi sheep disease virus/ganjam virus induces the translocation of protein disulphide isomerase-like oxidoreductases from the endoplasmic reticulum to the cell surface and the extracellular space.

    Science.gov (United States)

    Lasecka, Lidia; Baron, Michael D

    2014-01-01

    Nairobi sheep disease virus (NSDV) of the genus Nairovirus causes a haemorrhagic gastroenteritis in sheep and goats with mortality up to 90%; the virus is found in East and Central Africa, and in India, where the virus is called Ganjam virus. NSDV is closely related to the human pathogen Crimean-Congo haemorrhagic fever virus, which also causes a haemorrhagic disease. As with other nairoviruses, replication of NSDV takes place in the cytoplasm and the new virus particles bud into the Golgi apparatus; however, the effect of viral replication on cellular compartments has not been studied extensively. We have found that the overall structure of the endoplasmic reticulum (ER), the ER-Golgi intermediate compartment and the Golgi were unaffected by infection with NSDV. However, we observed that NSDV infection led to the loss of protein disulphide isomerase (PDI), an oxidoreductase present in the lumen of the endoplasmic reticulum (ER) and which assists during protein folding, from the ER. Further investigation showed that NSDV-infected cells have high levels of PDI at their surface, and PDI is also secreted into the culture medium of infected cells. Another chaperone from the PDI family, ERp57, was found to be similarly affected. Analysis of infected cells and expression of individual viral glycoproteins indicated that the NSDV PreGn glycoprotein is involved in redistribution of these soluble ER oxidoreductases. It has been suggested that extracellular PDI can activate integrins and tissue factor, which are involved respectively in pro-inflammatory responses and disseminated intravascular coagulation, both of which manifest in many viral haemorrhagic fevers. The discovery of enhanced PDI secretion from NSDV-infected cells may be an important finding for understanding the mechanisms underlying the pathogenicity of haemorrhagic nairoviruses.

  1. The nairovirus nairobi sheep disease virus/ganjam virus induces the translocation of protein disulphide isomerase-like oxidoreductases from the endoplasmic reticulum to the cell surface and the extracellular space.

    Directory of Open Access Journals (Sweden)

    Lidia Lasecka

    Full Text Available Nairobi sheep disease virus (NSDV of the genus Nairovirus causes a haemorrhagic gastroenteritis in sheep and goats with mortality up to 90%; the virus is found in East and Central Africa, and in India, where the virus is called Ganjam virus. NSDV is closely related to the human pathogen Crimean-Congo haemorrhagic fever virus, which also causes a haemorrhagic disease. As with other nairoviruses, replication of NSDV takes place in the cytoplasm and the new virus particles bud into the Golgi apparatus; however, the effect of viral replication on cellular compartments has not been studied extensively. We have found that the overall structure of the endoplasmic reticulum (ER, the ER-Golgi intermediate compartment and the Golgi were unaffected by infection with NSDV. However, we observed that NSDV infection led to the loss of protein disulphide isomerase (PDI, an oxidoreductase present in the lumen of the endoplasmic reticulum (ER and which assists during protein folding, from the ER. Further investigation showed that NSDV-infected cells have high levels of PDI at their surface, and PDI is also secreted into the culture medium of infected cells. Another chaperone from the PDI family, ERp57, was found to be similarly affected. Analysis of infected cells and expression of individual viral glycoproteins indicated that the NSDV PreGn glycoprotein is involved in redistribution of these soluble ER oxidoreductases. It has been suggested that extracellular PDI can activate integrins and tissue factor, which are involved respectively in pro-inflammatory responses and disseminated intravascular coagulation, both of which manifest in many viral haemorrhagic fevers. The discovery of enhanced PDI secretion from NSDV-infected cells may be an important finding for understanding the mechanisms underlying the pathogenicity of haemorrhagic nairoviruses.

  2. Effects of endoplasmic reticulum stress on the autophagy, apoptosis, and chemotherapy resistance of human breast cancer cells by regulating the PI3K/AKT/mTOR signaling pathway.

    Science.gov (United States)

    Zhong, Jia-Teng; Yu, Jian; Wang, Hai-Jun; Shi, Yu; Zhao, Tie-Suo; He, Bao-Xia; Qiao, Bin; Feng, Zhi-Wei

    2017-05-01

    Nowadays, although chemotherapy is an established therapy for breast cancer, the molecular mechanisms of chemotherapy resistance in breast cancer remain poorly understood. This study aims to explore the effects of endoplasmic reticulum stress on autophagy, apoptosis, and chemotherapy resistance in human breast cancer cells by regulating PI3K/AKT/mTOR signaling pathway. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay was performed to detect the cell viability of six human breast cancer cell lines (MCF-7, ZR-75-30, T47D, MDA-MB-435s, MDA-MB-453, and MDA-MB-231) treated with tunicamycin (5 µM), after which MCF-7 cells were selected for further experiment. Then, MCF-7 cells were divided into the control (without any treatment), tunicamycin (8 µ), BEZ235 (5 µ), and tunicamycin + BEZ235 groups. Cell viability of each group was testified by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Western blotting was applied to determine the expressions of endoplasmic reticulum stress and PI3K/AKT/mTOR pathway-related proteins and autophagy- and apoptosis-related proteins. Monodansylcadaverine and Annexin V-fluorescein isothiocyanate/propidium iodide staining were used for determination of cell autophagy and apoptosis. Furthermore, MCF-7 cells were divided into the control (without any treatment), tunicamycin (5 µM), cisplatin (16 µM), cisplatin (16 µM) + BEZ235 (5 µM), tunicamycin (5 µM) + cisplatin (16 µM), and tunicamycin (5 µM) + cisplatin (16 µM) + BEZ235 groups. Cell viability and apoptosis were also evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and Annexin V-fluorescein isothiocyanate/propidium iodide staining. In MCF-7 cells treated with tunicamycin, cell viability decreased significantly, but PEAK, eIF2, and CHOP were upregulated markedly and p-PI3K, p-AKT, and p-MTOR were downregulated in dose- and time-dependent manners. In the tunicamycin

  3. Endoplasmic Reticulum-Plasma Membrane Contact Sites.

    Science.gov (United States)

    Saheki, Yasunori; De Camilli, Pietro

    2017-06-20

    The endoplasmic reticulum (ER) has a broad localization throughout the cell and forms direct physical contacts with all other classes of membranous organelles, including the plasma membrane (PM). A number of protein tethers that mediate these contacts have been identified, and study of these protein tethers has revealed a multiplicity of roles in cell physiology, including regulation of intracellular Ca 2+ dynamics and signaling as well as control of lipid traffic and homeostasis. In this review, we discuss the cross talk between the ER and the PM mediated by direct contacts. We review factors that tether the two membranes, their properties, and their dynamics in response to the functional state of the cell. We focus in particular on the role of ER-PM contacts in nonvesicular lipid transport between the two bilayers mediated by lipid transfer proteins.

  4. Endoplasmic Reticulum (ER Stress and Endocrine Disorders

    Directory of Open Access Journals (Sweden)

    Daisuke Ariyasu

    2017-02-01

    Full Text Available The endoplasmic reticulum (ER is the organelle where secretory and membrane proteins are synthesized and folded. Unfolded proteins that are retained within the ER can cause ER stress. Eukaryotic cells have a defense system called the “unfolded protein response” (UPR, which protects cells from ER stress. Cells undergo apoptosis when ER stress exceeds the capacity of the UPR, which has been revealed to cause human diseases. Although neurodegenerative diseases are well-known ER stress-related diseases, it has been discovered that endocrine diseases are also related to ER stress. In this review, we focus on ER stress-related human endocrine disorders. In addition to diabetes mellitus, which is well characterized, several relatively rare genetic disorders such as familial neurohypophyseal diabetes insipidus (FNDI, Wolfram syndrome, and isolated growth hormone deficiency type II (IGHD2 are discussed in this article.

  5. Endoplasmic Reticulum (ER) Stress and Endocrine Disorders

    Science.gov (United States)

    Ariyasu, Daisuke; Yoshida, Hiderou; Hasegawa, Yukihiro

    2017-01-01

    The endoplasmic reticulum (ER) is the organelle where secretory and membrane proteins are synthesized and folded. Unfolded proteins that are retained within the ER can cause ER stress. Eukaryotic cells have a defense system called the “unfolded protein response” (UPR), which protects cells from ER stress. Cells undergo apoptosis when ER stress exceeds the capacity of the UPR, which has been revealed to cause human diseases. Although neurodegenerative diseases are well-known ER stress-related diseases, it has been discovered that endocrine diseases are also related to ER stress. In this review, we focus on ER stress-related human endocrine disorders. In addition to diabetes mellitus, which is well characterized, several relatively rare genetic disorders such as familial neurohypophyseal diabetes insipidus (FNDI), Wolfram syndrome, and isolated growth hormone deficiency type II (IGHD2) are discussed in this article. PMID:28208663

  6. Segregation in cast products

    Indian Academy of Sciences (India)

    Unknown

    The agreement with experimental data is mostly qualitative. The paper also ... For example, a high degree of positive segregation in the central region .... solute in a cast product, important ones being: size of casting, rate of solidification, mode.

  7. Segregation and Hispanic Homicide

    OpenAIRE

    Michael G. Bisciglia

    2014-01-01

    As the overall population of Hispanics within the United States has eclipsed that of African Americans, a mounting concern has developed regarding the rise in Hispanic lethal violence as a result of social and economic inequality. One means to measure this inequality is in the form of segregation. Research indicates that in many Hispanic communities, their levels of segregation from the White non-Hispanic population ar...

  8. Source Segregation and Collection of Source-Segregated Waste

    DEFF Research Database (Denmark)

    Christensen, Thomas Højlund; Matsufuji, Y.

    2011-01-01

    of optimal handling of the waste. But in a few cases, the waste must also be separated at source, for example removing the protective plastic cover from a commercial advertisement received by mail, prior to putting the advertisement into the waste collection bin for recyclable paper. These issues are often...... in wastes segregation addressing: - Purpose of source segregation. - Segregation criteria and guidance. - Segregation potentials and efficiencies. - Systems for collecting segregated fraction....

  9. Heat Shock Protein HSP27 Secretion by Ovarian Cancer Cells Is Linked to Intracellular Expression Levels, Occurs Independently of the Endoplasmic Reticulum Pathway and HSP27’s Phosphorylation Status, and Is Mediated by Exosome Liberation

    Directory of Open Access Journals (Sweden)

    Matthias B. Stope

    2017-01-01

    Full Text Available The heat shock protein HSP27 has been correlated in ovarian cancer (OC patients with aggressiveness and chemoresistance and, therefore, represents a promising potential biomarker for OC diagnosis, prognosis, and treatment response. Notably, secretion of soluble HSP27 has been described by a few cell types and may take place as well in OC cells. Therefore, we studied HSP27 secretion mechanisms under diverse cellular conditions in an OC cell model system. Secretion of HSP27 was characterized after overexpression of HSP27 by transfected plasmids and after heat shock. Intra- and extracellular HSP27 amounts were assessed by Western blotting and ELISA. Protein secretion was blocked by brefeldin A and the impact of the HSP27 phosphorylation status was analyzed overexpressing HSP27 phosphomutants. The present study demonstrated that HSP27 secretion by OVCAR-3 and SK-OV-3 cells depends on intracellular HSP27 concentrations. Moreover, HSP27 secretion is independent of the endoplasmic reticulum secretory pathway and HSP27 phosphorylation. Notably, analysis of OC cell-born exosomes not only confirmed the concentration-dependent correlation of HSP27 expression and secretion but also demonstrated a concentration-dependent incorporation of HSP27 protein into exosomes. Thus, secreted HSP27 may become more important as an extracellular factor which controls the tumor microenvironment and might be a noninvasive biomarker.

  10. Atorvastatin helps preserve pancreatic β cell function in obese C57BL/6 J mice and the effect is related to increased pancreas proliferation and amelioration of endoplasmic-reticulum stress

    Science.gov (United States)

    2014-01-01

    Background 3-Hydroxy-3-methyl-glutaryl CoA (HMG-CoA) reductase inhibitors or statins are competitive inhibitors of the rate-limiting enzyme in cholesterol biosynthesis. Currently, statins are used as first-line therapy in the treatment of diabetic dyslipidemia. However, effects of statins on β cell function remains unclear. This study aims to examine effects of atorvastatin treatment on pancreatic β cell function in obese C57BL/6 J mice and the possible mechanisms. Methods Diet-induced obesity (DIO) C57BL/6 J mice were treated with atorvastatin (30 mg/kg/day) for 58 days. β cell function was assessed by hyperglycemic clamp and the area of insulin-positive β cells was examined by immunofluorescence. Gene expression was assessed by RT-PCR, and endoplasmic reticulum (ER) stress related proteins were examined by Western blot. Additionally, cell viability and apoptosis of the cholesterol-loaded NIT-1 cells were investigated after atorvastatin treatment. Results Hyperglycemic clamp study revealed that glucose infusion rate (GIR) and insulin stimulation ratio in atorvastatin-treated DIO mice were markedly higher than control mice (P atorvastatin treatment (P atorvastatin-treated mice had significantly larger insulin-positive β cell area (P atorvastatin-treated mice (P atorvastatin protected the pancreatic β cell line of NIT-1 from cholesterol-induced apoptosis. Western blot showed increased expression of anti-apoptotic protein of B-cell lymphoma 2 (Bcl-2). Conclusion Pancreatic β cell function of obese C57BL/6 J mice was preserved after atorvastatin treatment, and this improvement may be attributed to enhanced pancreas proliferation and amelioration of pancreatic ER stress. PMID:24950764

  11. Cell wall metabolism and hexose allocation contribute to biomass accumulation in high yielding extreme segregants of a Saccharum interspecific F2 population.

    Science.gov (United States)

    Wai, Ching Man; Zhang, Jisen; Jones, Tyler C; Nagai, Chifumi; Ming, Ray

    2017-10-11

    Sugarcane is an emerging dual-purpose biofuel crop for energy and sugar production, owing to its rapid growth rate, high sucrose storage in the stems, and high lignocellulosic yield. It has the highest biomass production reaching 1.9 billion tonnes in 2014 worldwide. To improve sugarcane biomass accumulation, we developed an interspecific cross between Saccharum officinarum 'LA Purple' and Saccharum robustum 'MOL5829'. Selected F1 individuals were self-pollinated to generate a transgressive F2 population with a wide range of biomass yield. Leaf and stem internodes of fourteen high biomass and eight low biomass F2 extreme segregants were used for RNA-seq to decipher the molecular mechanism of rapid plant growth and dry weight accumulation. Gene Ontology terms involved in cell wall metabolism and carbohydrate catabolism were enriched among 3274 differentially expressed genes between high and low biomass groups. Up-regulation of cellulose metabolism, pectin degradation and lignin biosynthesis genes were observed in the high biomass group, in conjunction with higher transcript levels of callose metabolic genes and the cell wall loosening enzyme expansin. Furthermore, UDP-glucose biosynthesis and sucrose conversion genes were differentially expressed between the two groups. A positive correlation between stem glucose, but not sucrose, levels and dry weight was detected. We thus postulated that the high biomass sugarcane plants rapidly convert sucrose to UDP-glucose, which is the building block of cell wall polymers and callose, in order to maintain the rapid plant growth. The gene interaction of cell wall metabolism, hexose allocation and cell division contributes to biomass yield.

  12. Rapid phase segregation of P3HT:PCBM composites by thermal annealing for high-performance bulk-heterojunction solar cells

    Energy Technology Data Exchange (ETDEWEB)

    Fan, X.; Fang, G.J.; Qin, P.L.; Cheng, F.; Zhao, X.Z. [Wuhan University, Key Laboratory of Artificial Micro- and Nano-Structures of Ministry of Education, Department of Electronic Science and Technology, School of Physics and Technology, Wuhan (China)

    2011-12-15

    The performances of bulk-heterojunction (BHJ) solar cells are investigated for time-dependent thermal annealing with different morphology evolution scales, having special consideration for the diffusion and aggregation of fullerene derivative molecules based on blends of poly(3-hexylthiophene):[6,6]-phenyl-C{sub 61}-butyric acid methyl ester (P3HT:PCBM). Meaningfully, rapid formation of dot-like and needle-like crystalline PCBM structures of a few micrometers up to 60 {mu}m in size is obtained with thermal annealing treatment from 2 to 15 min, which dynamically reflects a fast process of PCBM molecule and cluster aggregation. Upon ultrasonic-assisted processing and annealing treatment, the scale of P3HT crystals is drastically increased in view of X-ray diffraction (XRD) patterns, leading to a high hole mobility. And, the P3HT domains can be gradually converted into larger P3HT crystals approved by the decreased full width at half-maximum in the XRD patterns. Corresponding current-voltage curves are measured in quantity and we propose a model to explain the effect of the crystalline degree of P3HT domains and aggregation of PCBM molecules and clusters on the phase segregation, expressing a viewpoint towards high performance of BHJ solar cells. (orig.)

  13. Endoplasmic reticulum: ER stress regulates mitochondrial bioenergetics

    Science.gov (United States)

    Bravo, Roberto; Gutierrez, Tomás; Paredes, Felipe; Gatica, Damián; Rodriguez, Andrea E.; Pedrozo, Zully; Chiong, Mario; Parra, Valentina; Quest, Andrew F.G.; Rothermel, Beverly A.; Lavandero, Sergio

    2014-01-01

    Endoplasmic reticulum (ER) stress activates an adaptive unfolded protein response (UPR) that facilitates cellular repair, however, under prolonged ER stress, the UPR can ultimately trigger apoptosis thereby terminating damaged cells. The molecular mechanisms responsible for execution of the cell death program are relatively well characterized, but the metabolic events taking place during the adaptive phase of ER stress remain largely undefined. Here we discuss emerging evidence regarding the metabolic changes that occur during the onset of ER stress and how ER influences mitochondrial function through mechanisms involving calcium transfer, thereby facilitating cellular adaptation. Finally, we highlight how dysregulation of ER–mitochondrial calcium homeostasis during prolonged ER stress is emerging as a novel mechanism implicated in the onset of metabolic disorders. PMID:22064245

  14. Segregation and Hispanic Homicide

    Directory of Open Access Journals (Sweden)

    Michael G. Bisciglia

    2014-01-01

    Full Text Available As the overall population of Hispanics within the United States has eclipsed that of African Americans, a mounting concern has developed regarding the rise in Hispanic lethal violence as a result of social and economic inequality. One means to measure this inequality is in the form of segregation. Research indicates that in many Hispanic communities, their levels of segregation from the White non-Hispanic population are similar to that of African Americans. Although a multitude of previous studies have looked at the impact of segregation among African Americans, the literature remains under-represented in terms of multi-city macro-level analyses among Hispanics. This current study extends the analysis of segregation’s effects on lethal violence to this population. To this end, two measures of segregation were used, the index of dissimilarity and exposure. Using data from the census and the Centers for Disease Control (CDC mortality files, negative binominal regression models were created using a sample of 236 U.S. cities. The results indicated that both measures of segregation show a strong positive influence on rates of Hispanic homicides.

  15. Understanding Segregation Processes

    Science.gov (United States)

    Bruch, Elizabeth

    There is growing consensus that living in neighborhoods of concentrated poverty increases the likelihood of social problems such as teenage parenthood, drug and alcohol use, crime victimization, and chronic unemployment. Neighborhood inequality is also implicated in studies of enduring race/ethnic health disparities, and there are recent moves to broaden the definition of health care policy to policies targeting social inequality (Mechanic 2007). Residential segregation affects health outcomes in several different ways. First, income, education, and occupation are all strongly related to health (Adler and Newman 2002). Segregation is a key mechanism through which socioeconomic inequality is perpetuated and reinforced, as it hinders the upward mobility of disadvantaged groups by limiting their educational and employment opportunities. Second, segregation increases minority exposure to unhealthy neighborhood environments. Residential segregation creates areas with concentrated poverty and unemployment, both of which are key factors that predict violence and create racial differences in homicide (Samson and Wilson 1995). Neighborhood characteristics, such as exposure to environmental hazards, fear of violence, and access to grocery stores, affect health risks and health behaviors (Cheadle et al. 1991). Tobacco and alcohol industries also advertise their products disproportionately in poor, minority areas (Moore, Williams, and Qualls 1996). Finally, residential segregation leads to inequalitie in health care resources, which contributes to disparities in quality of treatment (Smedley, Stith, and Nelson 2002).

  16. Salivary mRNA markers having the potential to detect oral squamous cell carcinoma segregated from oral leukoplakia with dysplasia.

    Science.gov (United States)

    Michailidou, Evangelia; Tzimagiorgis, Georgios; Chatzopoulou, Fani; Vahtsevanos, Konstantinos; Antoniadis, Konstantinos; Kouidou, Sofia; Markopoulos, Anastasios; Antoniades, Dimitrios

    2016-08-01

    In the current study the presence of extracellular IL-1B, IL-8, OAZ and SAT mRNAs in the saliva was evaluated as a tool in the early detection of oral squamous cell carcinoma. 34 patients with primary oral squamous cell carcinoma stage T1N0M0/T2N0M0, 20 patients with oral leukoplakia and dysplasia (15 patients with mild dysplasia and 5 with severe dysplasia/in situ carcinoma) and 31 matched healthy-control subjects were included in the study. The presence of IL-1B, IL-8, OAZ and SAT mRNA was evaluated in extracellular RNA isolated from saliva samples using sequence-specific primers and real-time RT-PCR. ROC curve analysis was used to estimate the ability of the biomarkers to detect oral squamous cell carcinoma patients. The data reveal that the combination of these four biomarkers provides a good predictive probability of up to 80% (AUC=0.799, p=0.002) for patients with oral squamous cell carcinoma but not patients suffering from oral leukoplakia with dysplasia. Moreover, the combination of only the two biomarkers (SAT and IL-8) also raises a high predictive ability of 75.5% (AUC=0.755, p=0.007) approximately equal to the four biomarkers suggesting the use of the two biomarkers only in the prediction model for oral squamous cell carcinoma patients limiting the economic and health cost in half. SAT and IL-8 mRNAs are present in the saliva in high quality and quantity, with a good discriminatory ability for oral squamous cell carcinoma patients only but not for patients with oral leukoplakia and dysplasia an oral potentially malignant disorder. Copyright © 2016. Published by Elsevier Ltd.

  17. Sch proteins are localized on endoplasmic reticulum membranes and are redistributed after tyrosine kinase receptor activation

    DEFF Research Database (Denmark)

    Lotti, L V; Lanfrancone, L; Migliaccio, E

    1996-01-01

    area of the cell and mostly associated with the cytosolic side of rough endoplasmic reticulum membranes. Upon epidermal growth factor treatment and receptor tyrosine kinase activation, the immunolabeling became peripheral and was found to be associated with the cytosolic surface of the plasma membrane....... The rough endoplasmic reticulum localization of Shc proteins in unstimulated cells and their massive recruitment to the plasma membrane, endocytic structures, and peripheral cytosol following receptor tyrosine kinase activation could account for multiple putative functions of the adaptor protein....

  18. A role for protein kinase C in the regulation of membrane fluidity and Ca²(+) flux at the endoplasmic reticulum and plasma membranes of HEK293 and Jurkat cells.

    Science.gov (United States)

    Chen, Lihong; Meng, Qingli; Jing, Xian; Xu, Pingxiang; Luo, Dali

    2011-02-01

    Protein kinase C (PKC) plays a prominent role in the regulation of a variety of cellular functions, including Ca²(+) signalling. In HEK293 and Jurkat cells, the Ca²(+) release and Ca²(+) uptake stimulated by several different activators were attenuated by activation of PKC with phorbol myristate acetate (PMA) or 1-oleoyl-2-acetyl-sn-glycerol (OAG) and potentiated by PKC inhibition with Gö6983 or knockdown of PKCα or PKCβ using shRNA. Immunostaining and Western blotting analyses revealed that PKCα and PKCβII accumulated at the plasma membrane (PM) and that these isoforms, along with PKCβI, also translocated to the endoplasmic reticulum (ER) upon activation with PMA. Measurements of membrane fluidity showed that, like the cell membrane stabilizers bovine serum albumin (BSA) and ursodeoxycholate (UDCA), PMA and OAG significantly reduced the fluidity of both the PM and ER membranes; these effects were blocked in PKC-knockdown cells. Interestingly, both BSA and UDCA inhibited the Ca²(+) responses to agonists to the same extent as PMA, whereas Tween 20, which increases membrane fluidity, raised the internal Ca²(+) concentration. Thus, activation of PKC induces both translocation of PKC to the PM and ER membranes and downregulation of membrane fluidity, thereby negatively modulating Ca²(+) flux. Copyright © 2010 Elsevier Inc. All rights reserved.

  19. Evidence that proliferation of golgi apparatus depends on both de novo generation from the endoplasmic reticulum and formation from pre-existing stacks during the growth of tobacco BY-2 cells.

    Science.gov (United States)

    Abiodun, Moses Olabiyi; Matsuoka, Ken

    2013-04-01

    In higher plants, the numbers of cytoplasmic-distributed Golgi stacks differ based on function, age and cell type. It has not been clarified how the numbers are controlled, whether all the Golgi apparatus in a cell function equally and whether the increase in Golgi number is a result of the de novo formation from the endoplasmic reticulum (ER) or fission of pre-existing stacks. A tobacco prolyl 4-hydroxylase (NtP4H1.1), which is a cis-Golgi-localizing type II membrane protein, was tagged with a photoconvertible fluorescent protein, mKikGR (monomeric Kikume green red), and expressed in tobacco bright yellow 2 (BY-2) cells. Transformed cells were exposed to purple light to convert the fluorescence from green to red. A time-course analysis after the conversion revealed a progressive increase in green puncta and a decrease in the red puncta. From 3 to 6 h, we observed red, yellow and green fluorescent puncta corresponding to pre-existing Golgi; Golgi containing both pre-existing and newly synthesized protein; and newly synthesized Golgi. Analysis of the number and fluorescence of Golgi at different phases of the cell cycle suggested that an increase in Golgi number with both division and de novo synthesis occurred concomitantly with DNA replication. Investigation with different inhibitors suggested that the formation of new Golgi and the generation of Golgi containing both pre-existing and newly synthesized protein are mediated by different machineries. These results and modeling based on quantified results indicate that the Golgi apparatuses in tobacco BY-2 cells are not uniform and suggest that both de novo synthesis from the ER and Golgi division contribute almost equally to the increase in proliferating cells.

  20. Nucleolus disassembly and distribution of segregated nucleolar material in prophase of root-tip meristematic cells in Triticum aestivum L.

    Directory of Open Access Journals (Sweden)

    Wang Jianyue

    2015-01-01

    Full Text Available This paper presents details of the process of nucleolar disassembly, studied by conventional transmission electron microscopy (TEM in wheat root cells. In early prophase, chromatin condensation and irregular nucleolar morphology are observed, with many small particles appearing around the nucleolus. In middle prophase, the nucleolus radiates outwards; in late prophase, the fine structure of the nucleolus disappears and nucleolar material diffuses away. Using “en bloc” silver-staining to distinguish between nucleoli and chromatin, we observed that the dispersed nucleolar material aggregates around the chromatin, forming a sheath-like perichromosomal structure that coats the chromosomes in late prophase.

  1. Dynamics of Escherichia coli Chromosome Segregation during Multifork Replication

    DEFF Research Database (Denmark)

    Nielsen, Henrik Jørck; Youngren, Brenda; Hansen, Flemming G.

    2007-01-01

    Slowly growing Escherichia coli cells have a simple cell cycle, with replication and progressive segregation of the chromosome completed before cell division. In rapidly growing cells, initiation of replication occurs before the previous replication rounds are complete. At cell division, the chro......Slowly growing Escherichia coli cells have a simple cell cycle, with replication and progressive segregation of the chromosome completed before cell division. In rapidly growing cells, initiation of replication occurs before the previous replication rounds are complete. At cell division......, the chromosomes contain multiple replication forks and must be segregated while this complex pattern of replication is still ongoing. Here, we show that replication and segregation continue in step, starting at the origin and progressing to the replication terminus. Thus, early-replicated markers on the multiple......-branched chromosomes continue to separate soon after replication to form separate protonucleoids, even though they are not segregated into different daughter cells until later generations. The segregation pattern follows the pattern of chromosome replication and does not follow the cell division cycle. No extensive...

  2. Sulforaphane induces apoptosis in T24 human urinary bladder cancer cells through a reactive oxygen species-mediated mitochondrial pathway: the involvement of endoplasmic reticulum stress and the Nrf2 signaling pathway.

    Science.gov (United States)

    Jo, Guk Heui; Kim, Gi-Young; Kim, Wun-Jae; Park, Kun Young; Choi, Yung Hyun

    2014-10-01

    Sulforaphane, a naturally occurring isothiocyanate found in cruciferous vegetables, has received a great deal of attention because of its ability to inhibit cell proliferation and induce apoptosis in cancer cells. In this study, we investigated the anticancer activity of sulforaphane in the T24 human bladder cancer line, and explored its molecular mechanism of action. Our results showed that treatment with sulforaphane inhibited cell viability and induced apoptosis in T24 cells in a concentration-dependent manner. Sulforaphane-induced apoptosis was associated with mitochondria dysfunction, cytochrome c release and Bcl-2/Bax dysregulation. Furthermore, the increased activity of caspase-9 and -3, but not caspase-8, was accompanied by the cleavage of poly ADP-ribose polymerase, indicating the involvement of the mitochondria-mediated intrinsic apoptotic pathway. Concomitant with these changes, sulforaphane triggered reactive oxygen species (ROS) generation, which, along with the blockage of sulforaphane-induced loss of mitochondrial membrane potential and apoptosis, was strongly attenuated by the ROS scavenger N-acetyl-L-cysteine. Furthermore, sulforaphane was observed to activate endoplasmic reticulum (ER) stress and the nuclear factor-E2-related factor-2 (Nrf2) signaling pathway, as demonstrated by the upregulation of ER stress‑related proteins, including glucose-regulated protein 78 and C/EBP-homologous protein, and the accumulation of phosphorylated Nrf2 proteins in the nucleus and induction of heme oxygenase-1 expression, respectively. Taken together, these results demonstrate that sulforaphane has antitumor effects against bladder cancer cells through an ROS-mediated intrinsic apoptotic pathway, and suggest that ER stress and Nrf2 may represent strategic targets for sulforaphane-induced apoptosis.

  3. Endoplasmic reticulum stress causes EBV lytic replication.

    Science.gov (United States)

    Taylor, Gwen Marie; Raghuwanshi, Sandeep K; Rowe, David T; Wadowsky, Robert M; Rosendorff, Adam

    2011-11-17

    Endoplasmic reticulum (ER) stress triggers a homeostatic cellular response in mammalian cells to ensure efficient folding, sorting, and processing of client proteins. In lytic-permissive lymphoblastoid cell lines (LCLs), pulse exposure to the chemical ER-stress inducer thapsigargin (TG) followed by recovery resulted in the activation of the EBV immediate-early (BRLF1, BZLF1), early (BMRF1), and late (gp350) genes, gp350 surface expression, and virus release. The protein phosphatase 1 a (PP1a)-specific phosphatase inhibitor Salubrinal (SAL) synergized with TG to induce EBV lytic genes; however, TG treatment alone was sufficient to activate EBV lytic replication. SAL showed ER-stress-dependent and -independent antiviral effects, preventing virus release in human LCLs and abrogating gp350 expression in 12-O-tetradecanoylphorbol-13-acetate (TPA)-treated B95-8 cells. TG resulted in sustained BCL6 but not BLIMP1 or CD138 expression, which is consistent with maintenance of a germinal center B-cell, rather than plasma-cell, phenotype. Microarray analysis identified candidate genes governing lytic replication in LCLs undergoing ER stress.

  4. Mutations reducing replication from R-loops suppress the defects of growth, chromosome segregation and DNA supercoiling in cells lacking topoisomerase I and RNase HI activity.

    Science.gov (United States)

    Usongo, Valentine; Martel, Makisha; Balleydier, Aurélien; Drolet, Marc

    2016-04-01

    R-loop formation occurs when the nascent RNA hybridizes with the template DNA strand behind the RNA polymerase. R-loops affect a wide range of cellular processes and their use as origins of replication was the first function attributed to them. In Escherichia coli, R-loop formation is promoted by the ATP-dependent negative supercoiling activity of gyrase (gyrA and gyrB) and is inhibited by topoisomerase (topo) I (topA) relaxing transcription-induced negative supercoiling. RNase HI (rnhA) degrades the RNA moiety of R-loops. The depletion of RNase HI activity in topA null mutants was previously shown to lead to extensive DNA relaxation, due to DNA gyrase inhibition, and to severe growth and chromosome segregation defects that were partially corrected by overproducing topo III (topB). Here, DNA gyrase assays in crude cell extracts showed that the ATP-dependent activity (supercoiling) of gyrase but not its ATP-independent activity (relaxation) was inhibited in topA null cells lacking RNase HI. To characterize the cellular event(s) triggered by the absence of RNase HI, we performed a genetic screen for suppressors of the growth defect of topA rnhA null cells. Suppressors affecting genes in replication (holC2::aph and dnaT18::aph) nucleotide metabolism (dcd49::aph), RNA degradation (rne59::aph) and fimbriae synthesis (fimD22::aph) were found to reduce replication from R-loops and to restore supercoiling, thus pointing to a correlation between R-loop-dependent replication in topA rnhA mutants and the inhibition of gyrase activity and growth. Interestingly, the position of fimD on the E. coli chromosome corresponds to the site of one of the five main putative origins of replication from R-loops in rnhA null cells recently identified by next-generation sequencing, thus suggesting that the fimD22::aph mutation inactivated one of these origins. Furthermore, we show that topo III overproduction is unable to complement the growth defect of topA rnhA null mutants at low

  5. Fisetin-induced apoptosis of human oral cancer SCC-4 cells through reactive oxygen species production, endoplasmic reticulum stress, caspase-, and mitochondria-dependent signaling pathways.

    Science.gov (United States)

    Su, Chen-Hsuan; Kuo, Chao-Lin; Lu, Kung-Wen; Yu, Fu-Shun; Ma, Yi-Shih; Yang, Jiun-Long; Chu, Yung-Lin; Chueh, Fu-Shin; Liu, Kuo-Ching; Chung, Jing-Gung

    2017-06-01

    Oral cancer is one of the cancer-related diseases in human populations and its incidence rates are rising worldwide. Fisetin, a flavonoid from natural products, has been shown to exhibit anticancer activities in many human cancer cell lines but the molecular mechanism of fisetin-induced apoptosis in human oral cancer cells is still unclear; thus, in this study, we investigated fisetin-induced cell death and associated signal pathways on human oral cancer SCC-4 cells in vitro. We examined cell morphological changes, total viable cells, and cell cycle distribution by phase contrast microscopy and flow cytometry assays. Reactive oxygen species (ROS), Ca 2+ , mitochondria membrane potential (ΔΨ m ), and caspase-8, -9, and -3 activities were also measured by flow cytometer. Results indicate that fisetin induced cell death through the cell morphological changes, caused G2/M phase arrest, induction of apoptosis, promoted ROS and Ca 2+ production, and decreased the level of ΔΨ m and increased caspase-3, -8, and -9 activities in SCC-4 cells. DAPI staining and DNA gel electrophoresis were also used to confirm fisetin-induced cell apoptosis in SCC-4 cells. Western blotting also found out that Fisetin increased the proapoptotic proteins such as Bax and Bid and decreased the antiapoptotic proteins such as Bcl-2. Furthermore, results also showed that Fisetin increased the cytochrome c, AIF, and Endo G release from mitochondria in SCC-4 cells. We also used ATF-6α, ATF-6β, GADD153, and GRP78 which indicated that fisetin induced cell death through ER stress. Based on those observations, we suggest that fisetin induced cell apoptosis through ER stress, mitochondria-, and caspase-dependent pathways. © 2017 Wiley Periodicals, Inc.

  6. Endoplasmic Reticulum Stress and Obesity.

    Science.gov (United States)

    Yilmaz, Erkan

    2017-01-01

    In recent years, the world has seen an alarming increase in obesity and closely associated with insulin resistance which is a state of low-grade inflammation, the latter characterized by elevated levels of proinflammatory cytokines in blood and tissues. A shift in energy balance alters systemic metabolic regulation and the important role that chronic inflammation, endoplasmic reticulum (ER) dysfunction, and activation of the unfolded protein response (UPR) play in this process.Why obesity is so closely associated with insulin resistance and inflammation is not understood well. This suggests that there are probably other causes for obesity-related insulin resistance and inflammation. One of these appears to be endoplasmic reticulum (ER) stress.The ER is a vast membranous network responsible for the trafficking of a wide range of proteins and plays a central role in integrating multiple metabolic signals critical in cellular homeostasis. Conditions that may trigger unfolded protein response activation include increased protein synthesis, the presence of mutant or misfolded proteins, inhibition of protein glycosylation, imbalance of ER calcium levels, glucose and energy deprivation, hypoxia, pathogens or pathogen-associated components and toxins. Thus, characterizing the mechanisms contributing to obesity and identifying potential targets for its prevention and treatment will have a great impact on the control of associated conditions, particularly T2D.

  7. Solute segregation during irradiation

    International Nuclear Information System (INIS)

    Wiedersich, H.; Okamoto, P.R.; Lam, N.Q.

    1977-01-01

    Irradiation at elevated temperature induces redistribution of the elements in alloys on a microstructural level. This phenomenon is caused by differences in the coupling of the various alloy constituents to the radiation-induced defect fluxes. A simple model of the segregation process based on coupled reaction-rate and diffusion equations is discussed. The model gives a good description of the experimentally observed consequences of radiation-induced segregation, including enrichment or depletion of solute elements near defect sinks such as surfaces, voids and dislocations; precipitation of second phases in solid solutions; precipitate redistribution in two-phase alloys; and effects of defect-production rates on void-swelling rates in alloys with minor solute additions

  8. Applied Thermodynamics: Grain Boundary Segregation

    Directory of Open Access Journals (Sweden)

    Pavel Lejček

    2014-03-01

    Full Text Available Chemical composition of interfaces—free surfaces and grain boundaries—is generally described by the Langmuir–McLean segregation isotherm controlled by Gibbs energy of segregation. Various components of the Gibbs energy of segregation, the standard and the excess ones as well as other thermodynamic state functions—enthalpy, entropy and volume—of interfacial segregation are derived and their physical meaning is elucidated. The importance of the thermodynamic state functions of grain boundary segregation, their dependence on volume solid solubility, mutual solute–solute interaction and pressure effect in ferrous alloys is demonstrated.

  9. Segregation in Religion Networks

    OpenAIRE

    Hu, Jiantao; Zhang, Qian-Ming; Zhou, Tao

    2018-01-01

    Religious beliefs could facilitate human cooperation [1-6], promote civic engagement [7-10], improve life satisfaction [11-13] and even boom economic development [14-16]. On the other side, some aspects of religion may lead to regional violence, intergroup conflict and moral prejudice against atheists [17-23]. Analogous to the separation of races [24], the religious segregation is a major ingredient resulting in increasing alienation, misunderstanding, cultural conflict and even violence amon...

  10. Zeaxanthin concentrations co-segregate with the magnitude of the blue light response of adaxial guard cells and leaf stomatal conductances in an F2 population of pima cotton

    Energy Technology Data Exchange (ETDEWEB)

    Quinones, M.A.; Lu Zhenmin; Zeiger, E. (Univ. of California, Los Angeles (United States))

    1993-05-01

    A blue light (BL) response of adaxial (AD) guard cells was investigated in two cotton lines with contrasting rates of stomatal conductances (g). This response is expressed as an enhancement of the red light-induced chlorophyll a fluorescence quenching by BL, and has an action spectrum indicative of a carotenoid photoreceptor. Ad guard cell from the high g, advanced line Pima S-6 have a higher carotenoid content and a larger BL response than those from the low g, primitive cotton, B368. In a growth chamber-grown F2 population of a cross between the two lines (n=30), g of individual plants segregated over a range exceeding the average g of the parental populations. Carotenoid content and the BL response of ad guard cell also segregated. There was a positive, strong correlation (r=0.71) between leaf g and the magnitude of the BL response of ad guard cells, indicating that both parameters are under genetic control, and that the BL response of guard cells contributes to the modulation of g. The concentration of all xanthopylls and [beta]-carotene in the ad guard cells correlated poorly with the BL response, except for zeaxanthin (r=0.71). In all green systems, xanthophylls are located inside the chloroplast which suggests that zeaxanthin functions in these organelle as a blue light photoreceptor for cotton guard cells.

  11. Segregation by onset asynchrony.

    Science.gov (United States)

    Hancock, P J B; Walton, L; Mitchell, G; Plenderleith, Y; Phillips, W A

    2008-08-05

    We describe a simple psychophysical paradigm for studying figure-ground segregation by onset asynchrony. Two pseudorandom arrays of Gabor patches are displayed, to left and right of fixation. Within one array, a subset of elements form a figure, such as a randomly curving path, that can only be reliably detected when their onset is not synchronized with that of the background elements. Several findings are reported. First, for most participants, segregation required an onset asynchrony of 20-40 ms. Second, detection was no better when the figure was presented first, and thus by itself, than when the background elements were presented first, even though in the latter case the figure could not be detected in either of the two successive displays alone. Third, asynchrony segregated subsets of randomly oriented elements equally well. Fourth, asynchronous onsets aligned with the path could be discriminated from those lying on the path but not aligned with it. Fifth, both transient and sustained neural activity contribute to detection. We argue that these findings are compatible with neural signaling by synchronized rate codes. Finally, schizophrenic disorganization is associated with reduced sensitivity. Thus, in addition to bearing upon basic theoretical issues, this paradigm may have clinical utility.

  12. Bitter melon extract ameliorates palmitate-induced apoptosis via inhibition of endoplasmic reticulum stress in HepG2 cells and high-fat/high-fructose-diet-induced fatty liver

    Directory of Open Access Journals (Sweden)

    Hwa Joung Lee

    2018-03-01

    Full Text Available Background: Bitter melon (BM improves glucose level, lipid homeostasis, and insulin resistance in vivo. However, the preventive mechanism of BM in nonalcoholic fatty liver disease (NAFLD has not been elucidated yet. Aim & Design: To determine the protective mechanism of bitter melon extract (BME, we performed experiments in vitro and in vivo. BME were treated palmitate (PA-administrated HepG2 cells. C57BL/6J mice were divided into two groups: high-fat/high-fructose (HF/HFr without or with BME supplementation (100 mg/kg body weight. Endoplasmic reticulum (ER stress, apoptosis, and biochemical markers were then examined by western blot and real-time PCR analyses. Results: BME significantly decreased expression levels of ER-stress markers (including phospho-eIF2α, CHOP, and phospho-JNK [Jun N-terminal kinases] in PA-treated HepG2 cells. BME also significantly decreased the activity of cleaved caspase-3 (a well known apoptotic-induced molecule and DNA fragmentation. The effect of BME on ER stress–mediated apoptosis in vitro was similarly observed in HF/HFr-fed mice in vivo. BME significantly reduced HF/HFr-induced hepatic triglyceride (TG and serum alanine aminotransferase (ALT as markers of hepatic damage in mice. In addition, BME ameliorated HF/HFr-induced serum TG and serum-free fatty acids. Conclusion: These data indicate that BME has protective effects against ER stress mediated apoptosis in HepG2 cells as well as in HF/HFr-induced fatty liver of mouse. Therefore, BME might be useful for preventing and treating NAFLD.

  13. The Predominant Pathway of Apoptosis in THP-1 Macrophage-Derived Foam Cells Induced by 5-Aminolevulinic Acid-Mediated Sonodynamic Therapy is the Mitochondria-Caspase Pathway Despite the Participation of Endoplasmic Reticulum Stress

    Directory of Open Access Journals (Sweden)

    Huan Wang

    2014-05-01

    Full Text Available Background: In advanced atherosclerosis, chronic endoplasmic reticulum (ER stress induces foam cells apoptosis and generates inflammatory reactions. Methods: THP-1 macrophage-derived foam cells (FC were incubated with 1 mM 5-aminolevulinic acid (ALA. After ALA mediated sonodynamic therapy (ALA-SDT, apoptosis of FC was assayed by Annexin V-PI staining. Intracellular reactive oxygen species (ROS and mitochondrial membrane potential were detected by staining with CellROX® Green Reagent and jc-1. Pretreatment of FC with N-acetylcysteine (NAC, Z-VAD-FMK or 4-phenylbutyrate (4-PBA, mitochondria apoptotic pathway associated proteins and C/EBP-homologous (CHOP expressions were assayed by wertern blotting. Results: Burst of apoptosis of FC was observed at 5-hour after ALA-SDT with 6-hour incubation of ALA and 0.4 W/cm2 ultrasound. After ALA-SDT, intracellular ROS level increased and mitochondrial membrane potential collapsed. Translocations of cytochrome c from mitochondria into cytosol and Bax from cytosol into mitochondria, cleaved caspase 9, cleaved caspase 3, upregulation of CHOP, as well as downregulation of Bcl-2 after ALA-SDT were detected, which could be suppressed by NAC. Activation of mitochondria-caspase pathway could not be inhibited by 4-PBA. Cleaved caspase 9 and caspase 3 as well as apoptosis induced by ALA-SDT could be inhibited by Z-VAD-FMK. Conclusion: The mitochondria-caspase pathway is predominant in the apoptosis of FC induced by ALA-SDT though ER stress participates in.

  14. Novel quinazolinone MJ-29 triggers endoplasmic reticulum stress and intrinsic apoptosis in murine leukemia WEHI-3 cells and inhibits leukemic mice.

    Directory of Open Access Journals (Sweden)

    Chi-Cheng Lu

    Full Text Available The present study was to explore the biological responses of the newly compound, MJ-29 in murine myelomonocytic leukemia WEHI-3 cells in vitro and in vivo fates. We focused on the in vitro effects of MJ-29 on ER stress and mitochondria-dependent apoptotic death in WEHI-3 cells, and to hypothesize that MJ-29 might fully impair the orthotopic leukemic mice. Our results indicated that a concentration-dependent decrease of cell viability was shown in MJ-29-treated cells. DNA content was examined utilizing flow cytometry, whereas apoptotic populations were determined using annexin V/PI, DAPI staining and TUNEL assay. Increasing vital factors of mitochondrial dysfunction by MJ-29 were further investigated. Thus, MJ-29-provaked apoptosis of WEHI-3 cells is mediated through the intrinsic pathway. Importantly, intracellular Ca(2+ release and ER stress-associated signaling also contributed to MJ-29-triggered cell apoptosis. We found that MJ-29 stimulated the protein levels of calpain 1, CHOP and p-eIF2α pathways in WEHI-3 cells. In in vivo experiments, intraperitoneal administration of MJ-29 significantly improved the total survival rate, enhanced body weight and attenuated enlarged spleen and liver tissues in leukemic mice. The infiltration of immature myeloblastic cells into splenic red pulp was reduced in MJ-29-treated leukemic mice. Moreover, MJ-29 increased the differentiations of T and B cells but decreased that of macrophages and monocytes. Additionally, MJ-29-stimulated immune responses might be involved in anti-leukemic activity in vivo. Based on these observations, MJ-29 suppresses WEHI-3 cells in vitro and in vivo, and it is proposed that this potent and selective agent could be a new chemotherapeutic candidate for anti-leukemia in the future.

  15. Endoplasmic reticulum proteostasis impairment in aging.

    Science.gov (United States)

    Martínez, Gabriela; Duran-Aniotz, Claudia; Cabral-Miranda, Felipe; Vivar, Juan P; Hetz, Claudio

    2017-08-01

    Perturbed neuronal proteostasis is a salient feature shared by both aging and protein misfolding disorders. The proteostasis network controls the health of the proteome by integrating pathways involved in protein synthesis, folding, trafficking, secretion, and their degradation. A reduction in the buffering capacity of the proteostasis network during aging may increase the risk to undergo neurodegeneration by enhancing the accumulation of misfolded proteins. As almost one-third of the proteome is synthetized at the endoplasmic reticulum (ER), maintenance of its proper function is fundamental to sustain neuronal function. In fact, ER stress is a common feature of most neurodegenerative diseases. The unfolded protein response (UPR) operates as central player to maintain ER homeostasis or the induction of cell death of chronically damaged cells. Here, we discuss recent evidence placing ER stress as a driver of brain aging, and the emerging impact of neuronal UPR in controlling global proteostasis at the whole organismal level. Finally, we discuss possible therapeutic interventions to improve proteostasis and prevent pathological brain aging. © 2017 The Authors. Aging Cell published by the Anatomical Society and John Wiley & Sons Ltd.

  16. Asymmetric strand segregation: epigenetic costs of genetic fidelity?

    Directory of Open Access Journals (Sweden)

    Diane P Genereux

    2009-06-01

    Full Text Available Asymmetric strand segregation has been proposed as a mechanism to minimize effective mutation rates in epithelial tissues. Under asymmetric strand segregation, the double-stranded molecule that contains the oldest DNA strand is preferentially targeted to the somatic stem cell after each round of DNA replication. This oldest DNA strand is expected to have fewer errors than younger strands because some of the errors that arise on daughter strands during their synthesis fail to be repaired. Empirical findings suggest the possibility of asymmetric strand segregation in a subset of mammalian cell lineages, indicating that it may indeed function to increase genetic fidelity. However, the implications of asymmetric strand segregation for the fidelity of epigenetic information remain unexplored. Here, I explore the impact of strand-segregation dynamics on epigenetic fidelity using a mathematical-modelling approach that draws on the known molecular mechanisms of DNA methylation and existing rate estimates from empirical methylation data. I find that, for a wide range of starting methylation densities, asymmetric -- but not symmetric -- strand segregation leads to systematic increases in methylation levels if parent strands are subject to de novo methylation events. I found that epigenetic fidelity can be compromised when enhanced genetic fidelity is achieved through asymmetric strand segregation. Strand segregation dynamics could thus explain the increased DNA methylation densities that are observed in structured cellular populations during aging and in disease.

  17. Salubrinal protects human skin fibroblasts against UVB-induced cell death by blocking endoplasmic reticulum (ER) stress and regulating calcium homeostasis.

    Science.gov (United States)

    Ji, Chao; Yang, Bo; Huang, Shu-Ying; Huang, Jin-Wen; Cheng, Bo

    2017-12-02

    The role of UVB in skin photo damages has been widely reported. Overexposure to UVB will induce severe DNA damages in epidermal cells and cause most cytotoxic symptoms. In the present study, we tested the potential activity of salubrinal, a selective inhibitor of Eukaryotic Initiation Factor 2 (eIF2) -alpha phosphatase, against UV-induced skin cell damages. We first exposed human fibroblasts to UVB radiation and evaluated the cytosolic Ca 2+ level as well as the induction of ER stress. We found that UVB radiation induced the depletion of ER Ca 2+ and increased the expression of ER stress marker including phosphorylated PERK, CHOP, and phosphorylated IRE1α. We then determined the effects of salubrinal in skin cell death induced by UVB radiation. We observed that cells pre-treated with salubrinal had a higher survival rate compared to cells treated with UVB alone. Pre-treatment with salubrinal successfully re-established the ER function and Ca 2+ homeostasis. Our results suggest that salubrinal can be a potential therapeutic agents used in preventing photoaging and photo damages. Copyright © 2017 Elsevier Inc. All rights reserved.

  18. Patterns of Residential Segregation.

    Directory of Open Access Journals (Sweden)

    Rémi Louf

    Full Text Available The spatial distribution of income shapes the structure and organisation of cities and its understanding has broad societal implications. Despite an abundant literature, many issues remain unclear. In particular, all definitions of segregation are implicitely tied to a single indicator, usually rely on an ambiguous definition of income classes, without any consensus on how to define neighbourhoods and to deal with the polycentric organization of large cities. In this paper, we address all these questions within a unique conceptual framework. We avoid the challenge of providing a direct definition of segregation and instead start from a definition of what segregation is not. This naturally leads to the measure of representation that is able to identify locations where categories are over- or underrepresented. From there, we provide a new measure of exposure that discriminates between situations where categories co-locate or repel one another. We then use this feature to provide an unambiguous, parameter-free method to find meaningful breaks in the income distribution, thus defining classes. Applied to the 2014 American Community Survey, we find 3 emerging classes-low, middle and higher income-out of the original 16 income categories. The higher-income households are proportionally more present in larger cities, while lower-income households are not, invalidating the idea of an increased social polarisation. Finally, using the density-and not the distance to a center which is meaningless in polycentric cities-we find that the richer class is overrepresented in high density zones, especially for larger cities. This suggests that density is a relevant factor for understanding the income structure of cities and might explain some of the differences observed between US and European cities.

  19. Endoplasmic Reticulum Stress in Reproductive Function

    Directory of Open Access Journals (Sweden)

    Kang-sheng LIU

    2016-09-01

    Full Text Available Normal folding requires that unique conditions should be maintained within the endoplasmic reticulum (ER lumen, and nascent proteins are initially bound to Ca2+dependent chaperone proteins. Proteins synthesized in the ER are properly folded with the assistance of ER chaperones. misfolded proteins are disposed by ER-associated protein degradation. Accumulation of misfolded proteins in the ER triggers an adaptive ER stress response, which leads to activation of the unfolded protein response (UPR, a conserved pathway that transmits signals to restore homeostasis or eliminate the irreparably damaged cells. It has been shown that ER stress involves in pathophysiological development of many diseases, including neurological diseases. However, nowadays, a few studies have begun to focus on the possibility that the accumulation of misfolded proteins can also contribute to reproductive diseases. In this article, we mainly introduced the involvement of ER stress response in preimplantation embryos, placental development, intrauterine growth restriction (IUGR and testicular germ cells so as to provide important insights for the molecular mechanisms of ER stress-induced apoptosis in reproductive diseases.

  20. The actin cytoskeleton modulates the activation of iNKT cells by segregating CD1d nanoclusters on antigen-presenting cells

    Science.gov (United States)

    Torreno-Pina, Juan A.; Manzo, Carlo; Salio, Mariolina; Aichinger, Michael C.; Oddone, Anna; Lakadamyali, Melike; Shepherd, Dawn; Besra, Gurdyal S.; Cerundolo, Vincenzo

    2016-01-01

    Invariant natural killer T (iNKT) cells recognize endogenous and exogenous lipid antigens presented in the context of CD1d molecules. The ability of iNKT cells to recognize endogenous antigens represents a distinct immune recognition strategy, which underscores the constitutive memory phenotype of iNKT cells and their activation during inflammatory conditions. However, the mechanisms regulating such “tonic” activation of iNKT cells remain unclear. Here, we show that the spatiotemporal distribution of CD1d molecules on the surface of antigen-presenting cells (APCs) modulates activation of iNKT cells. By using superresolution microscopy, we show that CD1d molecules form nanoclusters at the cell surface of APCs, and their size and density are constrained by the actin cytoskeleton. Dual-color single-particle tracking revealed that diffusing CD1d nanoclusters are actively arrested by the actin cytoskeleton, preventing their further coalescence. Formation of larger nanoclusters occurs in the absence of interactions between CD1d cytosolic tail and the actin cytoskeleton and correlates with enhanced iNKT cell activation. Importantly and consistently with iNKT cell activation during inflammatory conditions, exposure of APCs to the Toll-like receptor 7/8 agonist R848 increases nanocluster density and iNKT cell activation. Overall, these results define a previously unidentified mechanism that modulates iNKT cell autoreactivity based on the tight control by the APC cytoskeleton of the sizes and densities of endogenous antigen-loaded CD1d nanoclusters. PMID:26798067

  1. Induction of cortical endoplasmic reticulum by dimerization of a coatomer-binding peptide anchored to endoplasmic reticulum membranes

    OpenAIRE

    Lavieu, Grégory; Orci, Lelio; Shi, Lei; Geiling, Michael; Ravazzola, Mariella; Wieland, Felix; Cosson, Pierre; Rothman, James E.

    2010-01-01

    Cortical endoplasmic reticulum (cER) is a permanent feature of yeast cells but occurs transiently in most animal cell types. Ist2p is a transmembrane protein that permanently localizes to the cER in yeast. When Ist2 is expressed in mammalian cells, it induces abundant cER containing Ist2. Ist2 cytoplasmic C-terminal peptide is necessary and sufficient to induce cER. This peptide sequence resembles classic coat protein complex I (COPI) coatomer protein-binding KKXX signals, and indeed the dime...

  2. Inhibition of nuclear factor-kappaB or Bax prevents endoplasmic reticulum stress- but not nitric oxide-mediated apoptosis in INS-1E cells

    DEFF Research Database (Denmark)

    Tonnesen, Morten F; Grunnet, Lars G; Friberg, Josefine

    2009-01-01

    . Exposure of INS-1E cells to TG or SNAP caused caspase-3 cleavage and apoptosis. Both TG and SNAP induced activation of the proapoptotic transcription factor CCAAT/enhancer-binding protein homologous protein (CHOP). However, other classical ER stress-induced markers such as up-regulation of ER chaperone Bip...

  3. Endoplasmic reticulum-directed recombinant mRNA displays subcellular localization equal to endogenous mRNA during transient expression in CHO cells

    DEFF Research Database (Denmark)

    Beuchert Kallehauge, Thomas; Kol, Stefan; Andersen, Mikael Rørdam

    2016-01-01

    When expressing pharmaceutical recombinant proteins in mammalian cells, the protein is commonly directed through the secretory pathway, in a signal peptide-dependent manner, to acquire specific post-translational modifications and to facilitate secretion into the culture medium. One key premise...

  4. Chromosomal organization and segregation in Pseudomonas aeruginosa.

    Directory of Open Access Journals (Sweden)

    Isabelle Vallet-Gely

    2013-05-01

    Full Text Available The study of chromosomal organization and segregation in a handful of bacteria has revealed surprising variety in the mechanisms mediating such fundamental processes. In this study, we further emphasized this diversity by revealing an original organization of the Pseudomonas aeruginosa chromosome. We analyzed the localization of 20 chromosomal markers and several components of the replication machinery in this important opportunistic γ-proteobacteria pathogen. This technique allowed us to show that the 6.3 Mb unique circular chromosome of P. aeruginosa is globally oriented from the old pole of the cell to the division plane/new pole along the oriC-dif axis. The replication machinery is positioned at mid-cell, and the chromosomal loci from oriC to dif are moved sequentially to mid-cell prior to replication. The two chromosomal copies are subsequently segregated at their final subcellular destination in the two halves of the cell. We identified two regions in which markers localize at similar positions, suggesting a bias in the distribution of chromosomal regions in the cell. The first region encompasses 1.4 Mb surrounding oriC, where loci are positioned around the 0.2/0.8 relative cell length upon segregation. The second region contains at least 800 kb surrounding dif, where loci show an extensive colocalization step following replication. We also showed that disrupting the ParABS system is very detrimental in P. aeruginosa. Possible mechanisms responsible for the coordinated chromosomal segregation process and for the presence of large distinctive regions are discussed.

  5. Endoplasmic Reticulum-Associated rht-PA Processing in CHO Cells: Influence of Mild Hypothermia and Specific Growth Rates in Batch and Chemostat Cultures

    OpenAIRE

    Vergara, Mauricio; Berrios, Julio; Mart?nez, Irene; D?az-Barrera, Alvaro; Acevedo, Cristian; Reyes, Juan G.; Gonzalez, Ramon; Altamirano, Claudia

    2015-01-01

    Background Chinese hamster ovary (CHO) cells are the main host for producing recombinant proteins with human therapeutic applications mainly because of their capability to perform proper folding and glycosylation processes. In addition, mild hypothermia is one of the main strategies for maximising the productivity of these systems. However, little information is available on the effect of culture temperature on the folding and degradation processes of recombinant proteins that takes place in ...

  6. Waste segregation procedures and benefits

    International Nuclear Information System (INIS)

    Fish, J.D.; Massey, C.D.; Ward, S.J.

    1990-01-01

    Segregation is a critical first step in handling hazardous and radioactive materials to minimize the generation of regulated wastes. In addition, segregation can significantly reduce the complexity and the total cost of managing waste. Procedures at Sandia National Laboratories, Albuquerque require that wastes be segregated, first, by waste type (acids, solvents, low level radioactive, mixed, classified, etc.). Higher level segregation requirements, currently under development, are aimed at enhancing the possibilities for recovery, recycle and reapplication; reducing waste volumes; reducing waste disposal costs, and facilitating packaging storage, shipping and disposal. 2 tabs

  7. Dynamics of chromosome segregation in Escherichia coli

    DEFF Research Database (Denmark)

    Nielsen, Henrik Jørck

    2007-01-01

    Since the 1960’es the conformation and segregation of the chromosome in Escherichia coli has been a subject of interest for many scientists. However, after 40 years of research, we still know incredibly little about how the chromosome is organized inside the cell, how it manages to duplicate...... this incredibly big molecule and separate the two daughter chromosomes and how it makes sure that the daughter cells receives one copy each. The fully extended chromosome is two orders of magnitude larger than the cell in which it is contained. Hence the chromosome is heavily compacted in the cell...

  8. Met receptor inhibitor SU11274 localizes in the endoplasmic reticulum.

    Science.gov (United States)

    Wiest, Edwin J; Smith, Heather Jensen; Hollingsworth, Michael A

    2018-07-02

    We discovered that SU11274, a class I c-Met inhibitor, fluoresces when excited by 488 nm laser light and showed rapid specific accumulation in distinct subcellular compartments. Given that SU11274 reduces cancer cell viability, we exploited these newly identified spectral properties to determine SU11274 intracellular distribution and accumulation in human pancreatic cancer cells. The aim of the studies reported here was to identify organelle(s) to which SU11274 is trafficked. We conclude that SU11274 rapidly and predominantly accumulates in the endoplasmic reticulum. Copyright © 2018. Published by Elsevier Inc.

  9. Conditions for spatial segregation: some European perspectives.

    NARCIS (Netherlands)

    Musterd, S.; de Winter, M.

    1998-01-01

    Evaluates some theses on the theme of spatial segregation in Europe. Spatial segregation as an important issue on the political agendas of European nations; Two views of segregation in Europe; Strategies of European nations to deal with segregation; Segregation in European cities

  10. Spatial Reorganization of the Endoplasmic Reticulum during Mitosis Relies on Mitotic Kinase Cyclin A in the Early Drosophila Embryo

    Science.gov (United States)

    Bergman, Zane J.; Mclaurin, Justin D.; Eritano, Anthony S.; Johnson, Brittany M.; Sims, Amanda Q.; Riggs, Blake

    2015-01-01

    Mitotic cyclin-dependent kinase with their cyclin partners (cyclin:Cdks) are the master regulators of cell cycle progression responsible for regulating a host of activities during mitosis. Nuclear mitotic events, including chromosome condensation and segregation have been directly linked to Cdk activity. However, the regulation and timing of cytoplasmic mitotic events by cyclin:Cdks is poorly understood. In order to examine these mitotic cytoplasmic events, we looked at the dramatic changes in the endoplasmic reticulum (ER) during mitosis in the early Drosophila embryo. The dynamic changes of the ER can be arrested in an interphase state by inhibition of either DNA or protein synthesis. Here we show that this block can be alleviated by micro-injection of Cyclin A (CycA) in which defined mitotic ER clusters gathered at the spindle poles. Conversely, micro-injection of Cyclin B (CycB) did not affect spatial reorganization of the ER, suggesting CycA possesses the ability to initiate mitotic ER events in the cytoplasm. Additionally, RNAi-mediated simultaneous inhibition of all 3 mitotic cyclins (A, B and B3) blocked spatial reorganization of the ER. Our results suggest that mitotic ER reorganization events rely on CycA and that control and timing of nuclear and cytoplasmic events during mitosis may be defined by release of CycA from the nucleus as a consequence of breakdown of the nuclear envelope. PMID:25689737

  11. Shaping Segregation: Convexity vs. concavity

    NARCIS (Netherlands)

    Gonzalez Briones, Sebastián; Windows-Yule, Kit; Luding, Stefan; Parker, D.J.; Thornton, Anthony Richard

    2014-01-01

    Controlling segregation is both a practical and a theoretical challenge. In this Letter we demonstrate a manner in which rotation-induced segregation may be controlled by altering the geometry of the rotating containers in which granular systems are housed. Using a novel drum design comprising

  12. 4-Phenylbutyric Acid Reveals Good Beneficial Effects on Vital Organ Function via Anti-Endoplasmic Reticulum Stress in Septic Rats.

    Science.gov (United States)

    Liu, Liangming; Wu, Huiling; Zang, JiaTao; Yang, Guangming; Zhu, Yu; Wu, Yue; Chen, Xiangyun; Lan, Dan; Li, Tao

    2016-08-01

    Sepsis and septic shock are the common complications in ICUs. Vital organ function disorder contributes a critical role in high mortality after severe sepsis or septic shock, in which endoplasmic reticulum stress plays an important role. Whether anti-endoplasmic reticulum stress with 4-phenylbutyric acid is beneficial to sepsis and the underlying mechanisms are not known. Laboratory investigation. State Key Laboratory of Trauma, Burns and Combined Injury. Sprague-Dawley rats. Using cecal ligation and puncture-induced septic shock rats, lipopolysaccharide-treated vascular smooth muscle cells, and cardiomyocytes, effects of 4-phenylbutyric acid on vital organ function and the relationship with endoplasmic reticulum stress and endoplasmic reticulum stress-mediated inflammation, apoptosis, and oxidative stress were observed. Conventional treatment, including fluid resuscitation, vasopressin, and antibiotic, only slightly improved the hemodynamic variable, such as mean arterial blood pressure and cardiac output, and slightly improved the vital organ function and the animal survival of septic shock rats. Supplementation of 4-phenylbutyric acid (5 mg/kg; anti-endoplasmic reticulum stress), especially administered at early stage, significantly improved the hemodynamic variables, vital organ function, such as liver, renal, and intestinal barrier function, and animal survival in septic shock rats. 4-Phenylbutyric acid application inhibited the endoplasmic reticulum stress and endoplasmic reticulum stress-related proteins, such as CCAAT/enhancer-binding protein homologous protein in vital organs, such as heart and superior mesenteric artery after severe sepsis. Further studies showed that 4-phenylbutyric acid inhibited endoplasmic reticulum stress-mediated cytokine release, apoptosis, and oxidative stress via inhibition of nuclear factor-κB, caspase-3 and caspase-9, and increasing glutathione peroxidase and superoxide dismutase expression, respectively. Anti

  13. Acrolein Induces Endoplasmic Reticulum Stress and Causes Airspace Enlargement

    Science.gov (United States)

    Hanaoka, Masayuki; Natarajan, Ramesh; Kraskauskas, Donatas; Voelkel, Norbert F.

    2012-01-01

    Background Given the relative abundance and toxic potential of acrolein in inhaled cigarette smoke, it is surprising how little is known about the pulmonary and systemic effects of acrolein. Here we test the hypothesis whether systemic administration of acrolein could cause endoplasmic reticulum (ER) stress, and lung cell apoptosis, leading to the enlargement of the alveolar air spaces in rats. Methods Acute and chronic effects of intraperitoneally administered acrolein were tested. Mean alveolar airspace area was measured by using light microscopy and imaging system software. TUNEL staining and immunohistochemistry (IHC) for active caspase 3 and Western blot analysis for active caspase 3, and caspase 12 were performed to detect apoptosis. The ER-stress related gene expression in the lungs was determined by Quantitative real-time PCR analysis. Acrolein-protein adducts in the lung tissue were detected by IHC. Results Acute administration of acrolein caused a significant elevation of activated caspase 3, upregulation of VEGF expression and induced ER stress proteins in the lung tissue. The chronic administration of acrolein in rats led to emphysematous lung tissue remodeling. TUNEL staining and IHC for cleaved caspase 3 showed a large number of apoptotic septal cells in the acrolein-treated rat lungs. Chronic acrolein administration cause the endoplasmic reticulum stress response manifested by significant upregulation of ATF4, CHOP and GADd34 expression. In smokers with COPD there was a considerable accumulation of acrolein-protein adducts in the inflammatory, airway and vascular cells. Conclusions Systemic administration of acrolein causes endoplasmic reticulum stress response, lung cell apoptosis, and chronic administration leads to the enlargement of the alveolar air spaces and emphysema in rats. The substantial accumulation of acrolein-protein adducts in the lungs of COPD patients suggest a role of acrolein in the pathogenesis of emphysema. PMID:22675432

  14. The dissociation of tumor-induced weight loss from hypoglycemia in a transplantable pluripotent rat islet tumor results in the segregation of stable alpha- and beta-cell tumor phenotypes

    DEFF Research Database (Denmark)

    Madsen, O D; Karlsen, C; Nielsen, E

    1993-01-01

    in NEDH rats resulted in stable hypoglycemic insulinoma tumor lines, such as MSL-G2-IN. Occasionally, hypoglycemia as well as severe weight loss were observed in the early tumor passages of MSL-G and the subclone, NHI-5B, which carry the transfected neomycin and human insulin genes as unique clonal...... markers. By selective transplantation, it was possible to segregate stable anorectic normoglycemic tumor lines, MSL-G-AN and NHI-5B-AN, from both clones. These tumors cause an abrupt onset of anorexia when they reach a size of 400-500 mg (loss parallels...... a common clonal origin of pluripotent MSL cells, thus supporting the existence of a cell lineage relationship between islet alpha- and beta-cell during ontogeny; and 2) that our glucagonomas release an anorexigenic substance(s) of unknown nature that causes a severe weight loss comparable to that reported...

  15. Prokaryotic DNA segregation by an actin-like filament

    DEFF Research Database (Denmark)

    Møller-Jensen, Jakob; Bugge Jensen, Rasmus; Löwe, Jan

    2002-01-01

    The mechanisms responsible for prokaryotic DNA segregation are largely unknown. The partitioning locus (par) encoded by the Escherichia coli plasmid R1 actively segregates its replicon to daughter cells. We show here that the ParM ATPase encoded by par forms dynamic actin-like filaments with prop...... point for ParM polymerization. Hence, we provide evidence for a simple prokaryotic analogue of the eukaryotic mitotic spindle apparatus.......The mechanisms responsible for prokaryotic DNA segregation are largely unknown. The partitioning locus (par) encoded by the Escherichia coli plasmid R1 actively segregates its replicon to daughter cells. We show here that the ParM ATPase encoded by par forms dynamic actin-like filaments...

  16. Ceramide transport from endoplasmic reticulum to Golgi apparatus is not vesicle-mediated

    NARCIS (Netherlands)

    Kok, JW; Babia, T; Klappe, K; Egea, G; Hoekstra, D

    1998-01-01

    Ceramide (Cer) transfer from the endoplasmic reticulum (ER) to the Golgi apparatus was measured under conditions that block vesicle-mediated protein transfer. This was done either in intact cells by reducing the incubation temperature to 15 degrees C, or in streptolysin O-permeabilized cells by

  17. Sequential Introduction of Cations Deriving Large-Grain Csx FA1-x PbI3 Thin Film for Planar Hybrid Solar Cells: Insight into Phase-Segregation and Thermal-Healing Behavior.

    Science.gov (United States)

    Huang, Jiahao; Xu, Pan; Liu, Jian; You, Xiao-Zeng

    2017-03-01

    Composition engineering of perovskite materials has been demonstrated to be important for high-performance solar cells. Recently, the energy favorable hybridization of formamidinium (FA) and cesium (Cs) in three dimension lead halide perovskites has been attracting increasing attention due to its potential benefit on durability. Herein, we reported a simple and effective method to produce phase-pure CsxFA1-xPbI3 thin film via sequential introduction of cations, in which the FA cation was introduced by interdiffusion annealing in the presence of N-methylimidazole (NMI). NMI was employed as an additive to slow down the crystallization and thus drive the formation of CsxFA1-xPbI3 with micrometer grain size, which probably facilitate the charge dissociation and transportation in photovoltaic devices. More importantly, composition dependent phase-segregation has been revealed and investigated for the first time during the phase-pure mixed-cation perovskites CsxFA1-xPbI3. The present findings demonstrated that suppressing phase-segregation of mixed-cation perovskites by meticulous composition engineering is significant for further development of efficient photovoltaics. It also suggested that phase-pure Cs0.15FA0.85PbI3 may be a promising candidate with superior phase-durability, which performed an efficiency over 16% in planar perovskite solar cells. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. Sister chromatid segregation in meiosis II

    Science.gov (United States)

    Wassmann, Katja

    2013-01-01

    Meiotic divisions (meiosis I and II) are specialized cell divisions to generate haploid gametes. The first meiotic division with the separation of chromosomes is named reductional division. The second division, which takes place immediately after meiosis I without intervening S-phase, is equational, with the separation of sister chromatids, similar to mitosis. This meiotic segregation pattern requires the two-step removal of the cohesin complex holding sister chromatids together: cohesin is removed from chromosome arms that have been subjected to homologous recombination in meiosis I and from the centromere region in meiosis II. Cohesin in the centromere region is protected from removal in meiosis I, but this protection has to be removed—deprotected”—for sister chromatid segregation in meiosis II. Whereas the mechanisms of cohesin protection are quite well understood, the mechanisms of deprotection have been largely unknown until recently. In this review I summarize our current knowledge on cohesin deprotection. PMID:23574717

  19. Surface segregation during irradiation

    International Nuclear Information System (INIS)

    Rehn, L.E.; Lam, N.Q.

    1985-10-01

    Gibbsian adsorption is known to alter the surface composition of many alloys. During irradiation, four additional processes that affect the near-surface alloy composition become operative: preferential sputtering, displacement mixing, radiation-enhanced diffusion and radiation-induced segregation. Because of the mutual competition of these five processes, near-surface compositional changes in an irradiation environment can be extremely complex. Although ion-beam induced surface compositional changes were noted as long as fifty years ago, it is only during the past several years that individual mechanisms have been clearly identified. In this paper, a simple physical description of each of the processes is given, and selected examples of recent important progress are discussed. With the notable exception of preferential sputtering, it is shown that a reasonable qualitative understanding of the relative contributions from the individual processes under various irradiation conditions has been attained. However, considerably more effort will be required before a quantitative, predictive capability can be achieved. 29 refs., 8 figs

  20. Applied thermodynamics: Grain boundary segregation

    Czech Academy of Sciences Publication Activity Database

    Lejček, Pavel; Zheng, L.; Hofmann, S.; Šob, Mojmír

    2014-01-01

    Roč. 16, č. 3 (2014), s. 1462-1484 ISSN 1099-4300 R&D Projects: GA ČR(CZ) GAP108/12/0311; GA ČR GAP108/12/0144; GA MŠk(CZ) ED1.1.00/02.0068 Institutional support: RVO:68378271 ; RVO:68081723 Keywords : interfacial segregation * Gibbs energy of segregation * enthalpy * entropy * volume * grain boundaries * iron Subject RIV: BM - Solid Matter Physics ; Magnetism Impact factor: 1.502, year: 2014

  1. Habitat segregation in fish assemblages

    OpenAIRE

    Ibbotson, A.T.

    1990-01-01

    The segregation of habitats of fish assemblages found in the chalk streams and rivers within the Wessex, South West and Southern Water Authority boundaries in southern England have been examined. Habitat segregation is the most frequent type of resource partitioning in natural communities. The habitat of individual fish species will be defined in order to determine the following: (1) the requirements of each species in terms of depth, current velocity, substrate, cover etc.; (2) identify the ...

  2. The Consequences of Chromosome Segregation Errors in Mitosis and Meiosis

    Directory of Open Access Journals (Sweden)

    Tamara Potapova

    2017-02-01

    Full Text Available Mistakes during cell division frequently generate changes in chromosome content, producing aneuploid or polyploid progeny cells. Polyploid cells may then undergo abnormal division to generate aneuploid cells. Chromosome segregation errors may also involve fragments of whole chromosomes. A major consequence of segregation defects is change in the relative dosage of products from genes located on the missegregated chromosomes. Abnormal expression of transcriptional regulators can also impact genes on the properly segregated chromosomes. The consequences of these perturbations in gene expression depend on the specific chromosomes affected and on the interplay of the aneuploid phenotype with the environment. Most often, these novel chromosome distributions are detrimental to the health and survival of the organism. However, in a changed environment, alterations in gene copy number may generate a more highly adapted phenotype. Chromosome segregation errors also have important implications in human health. They may promote drug resistance in pathogenic microorganisms. In cancer cells, they are a source for genetic and phenotypic variability that may select for populations with increased malignance and resistance to therapy. Lastly, chromosome segregation errors during gamete formation in meiosis are a primary cause of human birth defects and infertility. This review describes the consequences of mitotic and meiotic errors focusing on novel concepts and human health.

  3. Income Segregation between Schools and School Districts

    Science.gov (United States)

    Owens, Ann; Reardon, Sean F.; Jencks, Christopher

    2016-01-01

    Although trends in the racial segregation of schools are well documented, less is known about trends in income segregation. We use multiple data sources to document trends in income segregation between schools and school districts. Between-district income segregation of families with children enrolled in public school increased by over 15% from…

  4. Strand-seq : A unifying tool for studies of chromosome segregation

    NARCIS (Netherlands)

    Falconer, Ester; Lansdorp, Peter M.

    2013-01-01

    Non random segregation of sister chromatids has been implicated to help specify daughter cell fate (the Silent Sister Hypothesis [1]) or to protect the genome of long-lived stem cells (the Immortal Strand Hypothesis [2]). The idea that sister chromatids are non-randomly segregated into specific

  5. The endoplasmic reticulum stress response in disease ...

    African Journals Online (AJOL)

    Rafael Vincent M. Manalo

    2017-07-12

    Jul 12, 2017 ... Review. The endoplasmic reticulum stress response in disease pathogenesis and pathophysiology .... This is an open access article under the CC BY-NC-ND license ... chain binding protein (BIP); however, ER stress permits the release, .... drugs designed to alleviate it often cause more harm long-term.

  6. Chlorhexidine-induced apoptosis or necrosis in L929 fibroblasts: A role for endoplasmic reticulum stress

    International Nuclear Information System (INIS)

    Faria, Gisele; Cardoso, Cristina R.B.; Larson, Roy E.; Silva, Joao S.; Rossi, Marcos A.

    2009-01-01

    Chlorhexidine (CHX), widely used as antiseptic and therapeutic agent in medicine and dentistry, has a toxic effect both in vivo and in vitro. The intrinsic mechanism underlying CHX-induced cytotoxicity in eukaryotic cells is, however, still unknown. A recent study from our laboratory has suggested that CHX may induce death in cultured L929 fibroblasts via endoplasmic reticulum (ER) stress. This hypothesis was further tested by means of light and electron microscopy, quantification of apoptosis and necrosis by flow cytometry, fluorescence visualization of the cytoskeleton and endoplasmic reticulum, and evaluation of the expression of 78-kDa glucose-regulated protein 78 (Grp78), a marker of activation of the unfolded protein response (UPR) in cultured L929 fibroblasts. Our finding showing increased Grp 78 expression in CHX-treated cells and the results of flow cytometry, cytoskeleton and endoplasmic reticulum fluorescence visualization, and scanning and transmission electron microscopy allowed us to suggest that CHX elicits accumulation of proteins in the endoplasmic reticulum, which causes ER overload, resulting in ER stress and cell death either by necrosis or apoptosis. It must be pointed out, however, that this does not necessarily mean that ER stress is the only way that CHX kills L929 fibroblasts, but rather that ER stress is an important target or indicator of cell death induced by this drug

  7. Image segregation in strabismic amblyopia.

    Science.gov (United States)

    Levi, Dennis M

    2007-06-01

    Humans with naturally occurring amblyopia show deficits thought to involve mechanisms downstream of V1. These include excessive crowding, abnormal global image processing, spatial sampling and symmetry detection and undercounting. Several recent studies suggest that humans with naturally occurring amblyopia show deficits in global image segregation. The current experiments were designed to study figure-ground segregation in amblyopic observers with documented deficits in crowding, symmetry detection, spatial sampling and counting, using similar stimuli. Observers had to discriminate the orientation of a figure (an "E"-like pattern made up of 17 horizontal Gabor patches), embedded in a 7x7 array of Gabor patches. When the 32 "background" patches are vertical, the "E" pops-out, due to segregation by orientation and performance is perfect; however, if the background patches are all, or mostly horizontal, the "E" is camouflaged, and performance is random. Using a method of constant stimuli, we varied the number of "background" patches that were vertical and measured the probability of correct discrimination of the global orientation of the E (up/down/left/right). Surprisingly, amblyopes who showed strong crowding and deficits in symmetry detection and counting, perform normally or very nearly so in this segregation task. I therefore conclude that these deficits are not a consequence of abnormal segregation of figure from background.

  8. Endoplasmic Reticulum Stress-Associated Lipid Droplet Formation and Type II Diabetes

    OpenAIRE

    Zhang, Xuebao; Zhang, Kezhong

    2012-01-01

    Diabetes mellitus (DM), a metabolic disorder characterized by hyperglycemia, is caused by insufficient insulin production due to excessive loss of pancreatic β cells (type I diabetes) or impaired insulin signaling due to peripheral insulin resistance (type II diabetes). Pancreatic β cell is the only insulin-secreting cell type that has highly developed endoplasmic reticulum (ER) to cope with high demands of insulin synthesis and secretion. Therefore, ER homeostasis is crucial to the proper fu...

  9. [The influence of N-, S-containing chinasolone derivatives (NC-224) on the biochemical and physicochemical parameters of membrane endoplasmatic reticulum and nuclear chromatine fractions of rats liver cells in conditions of its injury by tetrachloromethane].

    Science.gov (United States)

    Gubs'kyî, Iu I; Goriushko, G G; Belenichev, I F; Kovalenko, S I; Litvinova, N V; Marchenko, O M; Kurapova, T M; Babenko, L P; Velychko, O M

    2010-01-01

    Using biochemical and physicochemical methods of investigation in vivo, the effect of the substance NC-224, N-, S-chinasolone-derivative, on the lipoperoxidation activity in rat liver endoplasmatic reticulum membranes and nuclear chromatin fractions under tetrachloromethane intoxication have been studied. It was shown that NC-224 has pronounced antioxidant activity which is the biochemical basis of the substance membrane- and genome-protective effects and its ability to restore physicochemical properties of the surface and hydrophobic zones of hepatocyte membranes and structural parameter nuclear chromatin fractions in the conditions of chemical liver injury.

  10. Segregation in a Galton Board

    International Nuclear Information System (INIS)

    Benito, J G; Vidales, A M; Ippolito, I

    2009-01-01

    This work deals with a numerical study of the problem of separation of particles with different elastic properties. The separation procedure uses a Galton Board which consist in a bidimensional system of obstacles arranged in a triangular lattice. Disks of equal diameters but different elastic properties are launched from the top of the device. The Galton Board is commonly used for mixing particles, but here, we intend to find special conditions under which one can use it as a segregating device. We introduce a mixture of particles and generate, through simulations, different conditions to favor the segregation process based on the different elastic coefficients of the particles. We inspect which is the best configuration of size, density of obstacles and wall separation to favor the separations of particles. Our results prove that the Galton Board can be used as a segregation device under certain conditions.

  11. Grain Boundary Segregation in Metals

    CERN Document Server

    Lejcek, Pavel

    2010-01-01

    Grain boundaries are important structural components of polycrystalline materials used in the vast majority of technical applications. Because grain boundaries form a continuous network throughout such materials, their properties may limit their practical use. One of the serious phenomena which evoke these limitations is the grain boundary segregation of impurities. It results in the loss of grain boundary cohesion and consequently, in brittle fracture of the materials. The current book deals with fundamentals of grain boundary segregation in metallic materials and its relationship to the grain boundary structure, classification and other materials properties.

  12. Ricin A chain reaches the endoplasmic reticulum after endocytosis

    International Nuclear Information System (INIS)

    Liu Qiong; Zhan Jinbiao; Chen Xinhong; Zheng Shu

    2006-01-01

    Ricin is a potent ribosome inactivating protein and now has been widely used for synthesis of immunotoxins. To target ribosome in the mammalian cytosol, ricin must firstly retrograde transport from the endomembrane system to reach the endoplasmic reticulum (ER) where the ricin A chain (RTA) is recognized by ER components that facilitate its membrane translocation to the cytosol. In the study, the fusion gene of enhanced green fluorescent protein (EGFP)-RTA was expressed with the pET-28a (+) system in Escherichia coli under the control of a T7 promoter. The fusion protein showed a green fluorescence. The recombinant protein can be purified by metal chelated affinity chromatography on a column of NTA. The rabbit anti-GFP antibody can recognize the fusion protein of EGFP-RTA just like the EGFP protein. The cytotoxicity of EGFP-RTA and RTA was evaluated by the MTT assay in HeLa and HEP-G2 cells following fluid-phase endocytosis. The fusion protein had a similar cytotoxicity of RTA. After endocytosis, the subcellular location of the fusion protein can be observed with the laser scanning confocal microscopy and the immuno-gold labeling Electro Microscopy. This study provided important evidence by a visualized way to prove that RTA does reach the endoplasmic reticulum

  13. Endoplasmic Reticulum Stress-Related Factors Protect against Diabetic Retinopathy

    Directory of Open Access Journals (Sweden)

    Wei-Kun Hu

    2012-01-01

    Full Text Available The endoplasmic reticulum (ER is a principal mediator of signal transduction in the cell, and disruption of its normal function (a mechanism known as ER stress has been associated with the pathogenesis of several diseases. ER stress has been demonstrated to contribute to onset and progression of diabetic retinopathy (DR by induction of multiple inflammatory signaling pathways. Recent studies have begun to describe the gene expression profile of ER stress-related genes in DR; moreover, genes that play a protective role against DR have been identified. P58IPK was determined to be able to reduce retinal vascular leakage under high glucose conditions, thus protecting retinal cells. It has also been found by our lab that ER-associated protein degradation factors exhibit significantly different expression patterns in rat retinas under sustained high glucose conditions. Future research based upon these collective genomic findings will contribute to our overall understanding of DR pathogenesis as well as identify potential therapeutic targets.

  14. Maternal mRNAs of PEM and macho 1, the ascidian muscle determinant, associate and move with a rough endoplasmic reticulum network in the egg cortex.

    Science.gov (United States)

    Sardet, Christian; Nishida, Hiroki; Prodon, Francois; Sawada, Kaichiro

    2003-12-01

    Localization of maternal mRNAs in the egg cortex is an essential feature of polarity in embryos of Drosophila, Xenopus and ascidians. In ascidians, maternal mRNAs such as macho 1, a determinant of primary muscle-cell fate, belong to a class of postplasmic RNAs that are located along the animal-vegetal gradient in the egg cortex. Between fertilization and cleavage, these postplasmic RNAs relocate in two main phases. They further concentrate and segregate in small posterior blastomeres into a cortical structure, the centrosome-attracting body (CAB), which is responsible for unequal cleavages. By using high-resolution, fluorescent, in situ hybridization in eggs, zygotes and embryos of Halocynthia roretzi, we showed that macho 1 and HrPEM are localized on a reticulated structure situated within 2 mum of the surface of the unfertilized egg, and within 8 mum of the surface the vegetal region and then posterior region of the zygote. By isolating cortices from eggs and zygotes we demonstrated that this reticulated structure is a network of cortical rough endoplasmic reticulum (cER) that is tethered to the plasma membrane. The postplasmic RNAs macho 1 and HrPEM were located on the cER network and could be detached from it. We also show that macho 1 and HrPEM accumulated in the CAB and the cER network. We propose that these postplasmic RNAs relocalized after fertilization by following the microfilament- and microtubule-driven translocations of the cER network to the poles of the zygote. We also suggest that the RNAs segregate and concentrate in posterior blastomeres through compaction of the cER to form the CAB. A multimedia BioClip 'Polarity inside the egg cortex' tells the story and can be downloaded at www.bioclips.com/bioclip.html

  15. Endoplasmic Reticulum Stress and Associated ROS

    Directory of Open Access Journals (Sweden)

    Hafiz Maher Ali Zeeshan

    2016-03-01

    Full Text Available The endoplasmic reticulum (ER is a fascinating network of tubules through which secretory and transmembrane proteins enter unfolded and exit as either folded or misfolded proteins, after which they are directed either toward other organelles or to degradation, respectively. The ER redox environment dictates the fate of entering proteins, and the level of redox signaling mediators modulates the level of reactive oxygen species (ROS. Accumulating evidence suggests the interrelation of ER stress and ROS with redox signaling mediators such as protein disulfide isomerase (PDI-endoplasmic reticulum oxidoreductin (ERO-1, glutathione (GSH/glutathione disuphide (GSSG, NADPH oxidase 4 (Nox4, NADPH-P450 reductase (NPR, and calcium. Here, we reviewed persistent ER stress and protein misfolding-initiated ROS cascades and their significant roles in the pathogenesis of multiple human disorders, including neurodegenerative diseases, diabetes mellitus, atherosclerosis, inflammation, ischemia, and kidney and liver diseases.

  16. Regulation of calcium release from the endoplasmic reticulum by the serine hydrolase ABHD2.

    Science.gov (United States)

    Yun, Bogeon; Lee, HeeJung; Powell, Roger; Reisdorph, Nichole; Ewing, Heather; Gelb, Michael H; Hsu, Ku-Lung; Cravatt, Benjamin F; Leslie, Christina C

    2017-09-02

    The serine hydrolase inhibitors pyrrophenone and KT195 inhibit cell death induced by A23187 and H 2 O 2 by blocking the release of calcium from the endoplasmic reticulum and mitochondrial calcium uptake. The effect of pyrrophenone and KT195 on these processes is not due to inhibition of their known targets, cytosolic phospholipase A 2 and α/β-hydrolase domain-containing (ABHD) 6, respectively, but represent off-target effects. To identify targets of KT195, fibroblasts were treated with KT195-alkyne to covalently label protein targets followed by click chemistry with biotin azide, enrichment on streptavidin beads and tryptic peptide analysis by mass spectrometry. Although several serine hydrolases were identified, α/β-hydrolase domain-containing 2 (ABHD2) was the only target in which both KT195 and pyrrophenone competed for binding to KT195-alkyne. ABHD2 is a serine hydrolase with a predicted transmembrane domain consistent with its pull-down from the membrane proteome. Subcellular fractionation showed localization of ABHD2 to the endoplasmic reticulum but not to mitochondria or mitochondrial-associated membranes. Knockdown of ABHD2 with shRNA attenuated calcium release from the endoplasmic reticulum, mitochondrial calcium uptake and cell death in fibroblasts stimulated with A23187. The results describe a novel mechanism for regulating calcium transfer from the endoplasmic reticulum to mitochondria that involves the serine hydrolase ABHD2. Copyright © 2017 Elsevier Inc. All rights reserved.

  17. The endoplasmic reticulum is a hub to sort proteins toward unconventional traffic pathways and endosymbiotic organelles.

    Science.gov (United States)

    Bellucci, Michele; De Marchis, Francesca; Pompa, Andrea

    2017-12-18

    The discovery that much of the extracellular proteome in eukaryotic cells consists of proteins lacking a signal peptide, which cannot therefore enter the secretory pathway, has led to the identification of alternative protein secretion routes bypassing the Golgi apparatus. However, proteins harboring a signal peptide for translocation into the endoplasmic reticulum can also be transported along these alternative routes, which are still far from being well elucidated in terms of the molecular machineries and subcellular/intermediate compartments involved. In this review, we first try to provide a definition of all the unconventional protein secretion pathways in eukaryotic cells, as those pathways followed by proteins directed to an 'external space' bypassing the Golgi, where 'external space' refers to the extracellular space plus the lumen of the secretory route compartments and the inner space of mitochondria and plastids. Then, we discuss the role of the endoplasmic reticulum in sorting proteins toward unconventional traffic pathways in plants. In this regard, various unconventional pathways exporting proteins from the endoplasmic reticulum to the vacuole, plasma membrane, apoplast, mitochondria, and plastids are described, including the short routes followed by the proteins resident in the endoplasmic reticulum. © The Author(s) 2017. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  18. Gender Segregation: Separate but Effective?

    Science.gov (United States)

    Holthouse, David

    2010-01-01

    In 2002, only 11 public schools in the United States had gender-segregated classrooms. As of December 2009, there were more than 550. The movement is based on the hypothesis that hard-wired differences in the ways that male and female brains develop and function in childhood through adolescence require classrooms in which boys and girls are not…

  19. Sexual orientation, prejudice and segregation

    NARCIS (Netherlands)

    Plug, E.; Webbink, D.; Martin, N.

    2014-01-01

    This article examines whether gay and lesbian workers sort into tolerant occupations. With information on sexual orientation, prejudice, and occupational choice taken from Australian Twin Registers, we find that gays and lesbians shy away from prejudiced occupations. We show that our segregation

  20. Respiratory metabolism and calorie restriction relieve persistent endoplasmic reticulum stress induced by calcium shortage in yeast

    OpenAIRE

    Busti, Stefano; Mapelli, Valeria; Tripodi, Farida; Sanvito, Rossella; Magni, Fulvio; Coccetti, Paola; Rocchetti, Marcella; Nielsen, Jens; Alberghina, Lilia; Vanoni, Marco

    2016-01-01

    Calcium homeostasis is crucial to eukaryotic cell survival. By acting as an enzyme cofactor and a second messenger in several signal transduction pathways, the calcium ion controls many essential biological processes. Inside the endoplasmic reticulum (ER) calcium concentration is carefully regulated to safeguard the correct folding and processing of secretory proteins. By using the model organism Saccharomyces cerevisiae we show that calcium shortage leads to a slowdown of cell growth and met...

  1. Dietary toxins, endoplasmic reticulum (ER) stress and diabetes.

    Science.gov (United States)

    Hettiarachchi, Kalindi D; Zimmet, Paul Z; Myers, Mark A

    2008-05-01

    The incidence of Type 1 diabetes has been increasing at a rate too rapid to be due to changes in genetic risk. Instead changes in environmental factors are the likely culprit. The endoplasmic reticulum (ER) plays an important role in the production of newly synthesized proteins and interference with these processes leads to ER stress. The insulin-producing beta cells are particularly prone to ER stress as a result of their heavy engagement in insulin production. Increasing evidence suggests ER stress is central to initiation and progression of Type 1 diabetes. An early environmental exposure, such as toxins and viral infections, can impart a significant physiological load on beta cells to initiate abnormal processing of proinsulin, ER stress and insulin secretory defects. Release of altered proinsulin from the beta cells early in life may trigger autoimmunity in those with genetic susceptibility leading to cytokine-induced nitric oxide production and so exacerbating ER stress in beta cells, ultimately leading to apoptosis of beta cells and diabetes. Here we suggest that ER stress is an inherent cause of beta cell dysfunction and environmental factors, in particular dietary toxins derived from Streptomyces in infected root vegetables, can impart additional stress that aggravates beta cell death and progression to diabetes. Furthermore, we propose that the increasing incidence of Type 1 diabetes may be accounted for by increased dietary exposure to ER-stress-inducing Streptomyces toxins.

  2. 36 CFR 254.6 - Segregative effect.

    Science.gov (United States)

    2010-07-01

    ... 36 Parks, Forests, and Public Property 2 2010-07-01 2010-07-01 false Segregative effect. 254.6... ADJUSTMENTS Land Exchanges § 254.6 Segregative effect. (a) If a proposal is made to exchange Federal lands... segregative effect terminates as follows: (1) Automatically, upon issuance of a patent or other document of...

  3. Dynamic Changes in Sarcoplasmic Reticulum Structure in Ventricular Myocytes

    Directory of Open Access Journals (Sweden)

    Amanda L. Vega

    2011-01-01

    sarcoplasmic reticulum (SR and the sarcolemma where Ca2+ release is activated. Here, we tested the hypothesis that the SR is a structurally inert organelle in ventricular myocytes. Our data suggest that rather than being static, the SR undergoes frequent dynamic structural changes. SR boutons expressing functional ryanodine receptors moved throughout the cell, approaching or moving away from the sarcolemma of ventricular myocytes. These changes in SR structure occurred in the absence of changes in [Ca2+] during EC coupling. Microtubules and the molecular motors dynein and kinesin 1(Kif5b were important regulators of SR motility. These findings support a model in which the SR is a motile organelle capable of molecular motor protein-driven structural changes.

  4. Osteochondritis dissecans (OCD), an endoplasmic reticulum storage disease?

    DEFF Research Database (Denmark)

    Skagen, Peter Storgaard; Horn, T; Kruse, H A

    2011-01-01

    in chondrocytes and extracellular matrix of cartilage from OCD patients. Abnormal type II collagen heterofibrils in "bundles" and chondrocytes with abnormal accumulation of matrix proteins in distended rough endoplasmic reticulum were typical findings. Further, Von Kossa staining and TEM showed empty lacunae...... close to mineralized "islands" in the cartilage and hypertrophic chondrocytes containing accumulated matrix proteins. Immunostaining revealed: (1) that types I, II, VI and X collagens and aggrecans were deposited intracellulary and (2) co-localization within the islands of types I, II, X collagens...... and aggrecan indicating that hypertrophic chondrocytes express a phenotype of bone cells during endochondral ossification. Types I, VI and X collagens were also present across the entire dissecates suggesting that chondrocytes were dedifferentiated. DNA sequencings were non-conclusive, only single nucleotide...

  5. Homocysteine inhibits hepatocyte proliferation via endoplasmic reticulum stress.

    Directory of Open Access Journals (Sweden)

    Xue Yu

    Full Text Available Homocysteine is an independent risk factor for coronary, cerebral, and peripheral vascular diseases. Recent studies have shown that levels of homocysteine are elevated in patients with impaired hepatic function, but the precise role of homocysteine in the development of hepatic dysfunction is unclear. In this study, we examined the effect of homocysteine on hepatocyte proliferation in vitro. Our results demonstrated that homocysteine inhibited hepatocyte proliferation by up-regulating protein levels of p53 as well as mRNA and protein levels of p21(Cip1 in primary cultured hepatocytes. Homocysteine induced cell growth arrest in p53-positive hepatocarcinoma cell line HepG2, but not in p53-null hepatocarcinoma cell line Hep3B. A p53 inhibitor pifithrin-α inhibited the expression of p21(Cip1 and attenuated homocysteine-induced cell growth arrest. Homocysteine induced TRB3 expression via endoplasmic reticulum stress pathway, resulting in Akt dephosphorylation. Knock-down of endogenous TRB3 significantly suppressed the inhibitory effect of homocysteine on cell proliferation and the phosphorylation of Akt. LiCl reversed homocysteine-mediated cell growth arrest by inhibiting TRB3-mediated Akt dephosphorylation. These results demonstrate that both TRB3 and p21(Cip1 are critical molecules in the homocysteine signaling cascade and provide a mechanistic explanation for impairment of liver regeneration in hyperhomocysteinemia.

  6. Strand-seq: A unifying tool for studies of chromosome segregation

    OpenAIRE

    Falconer, Ester; Lansdorp, Peter M.

    2013-01-01

    Non random segregation of sister chromatids has been implicated to help specify daughter cell fate (the Silent Sister Hypothesis [1]) or to protect the genome of long-lived stem cells (the Immortal Strand Hypothesis [2]). The idea that sister chromatids are non-randomly segregated into specific daughter cells is only marginally supported by data in sporadic and often contradictory studies. As a result, the field has moved forward rather slowly. The advent of being able to directly label and d...

  7. Nonequilibrium Segregation in Petroleum Reservoirs

    DEFF Research Database (Denmark)

    Shapiro, Alexander; Stenby, Erling Halfdan

    1999-01-01

    We analyse adsorption of a multicomponent mixture at high pressure on the basis of the potential theory of adsorption. The adsorbate is considered as a segregated mixture in the external field produced by a solid adsorbent. we derive an analytical equation for the thickness of a multicomponent fi...... close to a dew point. This equation (asymptotic adsorption equation, AAE) is a first order approximation with regard to the distance from a phase envelope....

  8. Segregation and the gender wage gaps in the private and the public sector

    DEFF Research Database (Denmark)

    Albæk, Karsten; Larsen, Mona; Stage Thomsen, Lars

    2016-01-01

    This paper examines the relation between segregation and the gender wage gap in the public and the private sectors in Denmark from 2002 to 2012. The analysis shows that male–female differences in the share of females in occupations, industries,establishments and job cells (occupations within...... establishments) constitute 46% of the raw gender wage gap in the private sector, while segregation in t he public sector accounts for as much as 63 %. Segregation thus plays a substantially more important role in accounting for the gender wage gap in the public sector than in the private sector. While...... the importance of segregation for wage formation decreased substantially in the public sector over time, it only decreased slightly in the private sector. Although the remaining gender wage gap, after controlling for segregation, is close to zero in the public sector, a substantial within-job cell differential...

  9. Nitric oxide induces segregation of decay accelerating factor (DAF or CD55) from the membrane lipid-rafts and its internalization in human endometrial cells.

    Science.gov (United States)

    Banadakoppa, Manu; Goluszko, Pawel; Liebenthal, Daniel; Yallampalli, Chandra

    2012-10-01

    Recent studies suggest that DAF (decay accelerating factor), a complement regulatory protein, present in lipid rafts, is utilized by Dr fimbriated Escherichia coli for their binding and internalization. Previous studies in our laboratory have shown that NO (nitric oxide) can reduce the invasion of Dr(+) E. coli and the severity of uterine infection in pregnant rats. Also, the expression level of DAF both at the mRNA and protein levels has been shown to be reduced by NO. Therefore NO mediated down-regulation of DAF appears to be an important factor in reducing the susceptibility to E. coli infection. However, it is unclear if NO can actually modulate the membrane association of DAF and therefore initial bacterial binding to cells. We found that NO induces the delocalization of DAF from the G(M1)-rich lipid rafts. Using biochemical and cell biological approaches in a uterine epithelial cell model (Ishikawa cells), DAF accumulates in caveolae upon exposure to NO. Interaction of DAF with the caveolar protein, caveolin1, leads to their internalization by endosomes. NO-induced delocalization of DAF from the lipid raft and its accumulation in caveolae are mediated through a cGMP (cyclic guanosine monophosphate) pathway. The acute localized synthesis of NO and its influence on DAF localization may represent an important unrecognized phenomenon of host defence against Dr(+) E. coli bacteria, as well as many disease conditions that involve complement system.

  10. Sphingosine inhibits the sarco(endo)plasmic reticulum Ca"2"+-ATPase (SERCA) activity

    International Nuclear Information System (INIS)

    Benaim, Gustavo; Pimentel, Adriana A.; Felibertt, Pimali; Mayora, Adriana; Colman, Laura; Sojo, Felipe; Rojas, Héctor; De Sanctis, Juan B.

    2016-01-01

    The increase in the intracellular Ca"2"+ concentration ([Ca"2"+]_i) is the key variable for many different processes, ranging from regulation of cell proliferation to apoptosis. In this work we demonstrated that the sphingolipid sphingosine (Sph) increases the [Ca"2"+]_i by inhibiting the sarco(endo)plasmic reticulum Ca"2"+-ATPase (SERCA), in a similar manner to thapsigargin (Tg), a specific inhibitor of this Ca"2"+ pump. The results showed that addition of sphingosine produced a release of Ca"2"+ from the endoplasmic reticulum followed by a Ca"2"+ entrance from the outside mileu. The results presented in this work support that this sphingolipid could control the activity of the SERCA, and hence sphingosine may participate in the regulation of [Ca"2"+]_I in mammalian cells.

  11. Sphingosine inhibits the sarco(endo)plasmic reticulum Ca{sup 2+}-ATPase (SERCA) activity

    Energy Technology Data Exchange (ETDEWEB)

    Benaim, Gustavo, E-mail: gbenaim@idea.gob.ve [Instituto de Estudios Avanzados (IDEA), Caracas (Venezuela, Bolivarian Republic of); Instituto de Biología Experimental, Facultad de Ciencias, Universidad Central de Venezuela (UCV), Caracas (Venezuela, Bolivarian Republic of); Pimentel, Adriana A., E-mail: adriana.pimentel@ucv.ve [Facultad de Farmacia, Universidad Central de Venezuela (UCV), Caracas (Venezuela, Bolivarian Republic of); Felibertt, Pimali [Facultad de Ciencias, Universidad de Carabobo, Valencia (Venezuela, Bolivarian Republic of); Mayora, Adriana [Instituto de Biología Experimental, Facultad de Ciencias, Universidad Central de Venezuela (UCV), Caracas (Venezuela, Bolivarian Republic of); Colman, Laura [Instituto Pasteur de Montevideo, Montevideo (Uruguay); Sojo, Felipe [Instituto de Biología Experimental, Facultad de Ciencias, Universidad Central de Venezuela (UCV), Caracas (Venezuela, Bolivarian Republic of); Rojas, Héctor [Instituto de Inmunología, Universidad Central de Venezuela (UCV), Caracas (Venezuela, Bolivarian Republic of); Centro de Biofísica y Bioquímica, Instituto Venezolano de Investigaciones Científicas (IVIC), Caracas (Venezuela, Bolivarian Republic of); De Sanctis, Juan B. [Instituto de Inmunología, Universidad Central de Venezuela (UCV), Caracas (Venezuela, Bolivarian Republic of)

    2016-04-29

    The increase in the intracellular Ca{sup 2+} concentration ([Ca{sup 2+}]{sub i}) is the key variable for many different processes, ranging from regulation of cell proliferation to apoptosis. In this work we demonstrated that the sphingolipid sphingosine (Sph) increases the [Ca{sup 2+}]{sub i} by inhibiting the sarco(endo)plasmic reticulum Ca{sup 2+}-ATPase (SERCA), in a similar manner to thapsigargin (Tg), a specific inhibitor of this Ca{sup 2+} pump. The results showed that addition of sphingosine produced a release of Ca{sup 2+} from the endoplasmic reticulum followed by a Ca{sup 2+} entrance from the outside mileu. The results presented in this work support that this sphingolipid could control the activity of the SERCA, and hence sphingosine may participate in the regulation of [Ca{sup 2+}]{sub I} in mammalian cells.

  12. Combined age and segregated kinetic model for industrial-scale penicillin fed-batch cultivation

    NARCIS (Netherlands)

    Wang, Z.F.; Lauwerijssen, M.J.C.; Yuan, J.Q.

    2005-01-01

    This paper proposes a cell age model forPenicillium chrysogenum fed-batch cultivation to supply a qualitative insight into morphology-associated dynamics. The average ages of the segregated cell populations, such as growing cells, non-growing cells and intact productive cells, were estimated by this

  13. F-actin-like filaments formed by plasmid segregation protein ParM

    DEFF Research Database (Denmark)

    van den Ent, Fusinita; Møller-Jensen, Jakob; Amos, Linda A.

    2002-01-01

    It was the general belief that DNA partitioning in prokaryotes is independent of a cytoskeletal structure, which in eukaryotic cells is indispensable for DNA segregation. Recently, however, immunofluorescence microscopy revealed highly dynamic, filamentous structures along the longitudinal axis...

  14. Are segregated sports classes scientifically justified?

    OpenAIRE

    Lawson, Sian; Hall, Edward

    2014-01-01

    School sports classes are a key part of physical and mental development, yet in many countries these classes are gender segregated. Before institutionalised segregation can be condoned it is important to tackle assumptions and check for an evidence-based rationale. This presentation aims to analyse the key arguments for segregation given in comment-form response to a recent media article discussing mixed school sports (Lawson, 2013).\\ud \\ud The primary argument given was division for strength...

  15. Digital morphogenesis via Schelling segregation

    Science.gov (United States)

    Barmpalias, George; Elwes, Richard; Lewis-Pye, Andrew

    2018-04-01

    Schelling’s model of segregation looks to explain the way in which particles or agents of two types may come to arrange themselves spatially into configurations consisting of large homogeneous clusters, i.e. connected regions consisting of only one type. As one of the earliest agent based models studied by economists and perhaps the most famous model of self-organising behaviour, it also has direct links to areas at the interface between computer science and statistical mechanics, such as the Ising model and the study of contagion and cascading phenomena in networks. While the model has been extensively studied it has largely resisted rigorous analysis, prior results from the literature generally pertaining to variants of the model which are tweaked so as to be amenable to standard techniques from statistical mechanics or stochastic evolutionary game theory. In Brandt et al (2012 Proc. 44th Annual ACM Symp. on Theory of Computing) provided the first rigorous analysis of the unperturbed model, for a specific set of input parameters. Here we provide a rigorous analysis of the model’s behaviour much more generally and establish some surprising forms of threshold behaviour, notably the existence of situations where an increased level of intolerance for neighbouring agents of opposite type leads almost certainly to decreased segregation.

  16. From particle segregation to the granular clock

    International Nuclear Information System (INIS)

    Lambiotte, R.; Salazar, J.M.; Brenig, L.

    2005-01-01

    Recently several authors studied the segregation of particles for a system composed of mono-dispersed inelastic spheres contained in a box divided by a wall in the middle. The system exhibited a symmetry breaking leading to an overpopulation of particles in one side of the box. Here we study the segregation of a mixture of particles composed of inelastic hard spheres and fluidized by a vibrating wall. Our numerical simulations show a rich phenomenology: horizontal segregation and periodic behavior. We also propose an empirical system of ODEs representing the proportion of each type of particles and the segregation flux of particles. These equations reproduce the major features observed by the simulations

  17. From particle segregation to the granular clock

    Energy Technology Data Exchange (ETDEWEB)

    Lambiotte, R. [Physique Statistique, Plasmas et Optique Non-lineaire, Universite Libre de Bruxelles, Campus Plaine, Boulevard du Triomphe, Code Postal 231, 1050 Brussels (Belgium)]. E-mail: rlambiot@ulb.ac.be; Salazar, J.M. [Universite De Bougogne-LRRS UMR-5613 CNRS, Faculte des Sciences Mirande, 9 Av. Alain Savary, 21078 Dijon Cedex (France)]. E-mail: jmarcos@u-bourgogne.fr; Brenig, L. [Physique Statistique, Plasmas et Optique Non-lineaire, Universite Libre de Bruxelles, Campus Plaine, Boulevard du Triomphe, Code Postal 231, 1050 Brussels (Belgium)]. E-mail: lbrenig@ulb.ac.be

    2005-08-01

    Recently several authors studied the segregation of particles for a system composed of mono-dispersed inelastic spheres contained in a box divided by a wall in the middle. The system exhibited a symmetry breaking leading to an overpopulation of particles in one side of the box. Here we study the segregation of a mixture of particles composed of inelastic hard spheres and fluidized by a vibrating wall. Our numerical simulations show a rich phenomenology: horizontal segregation and periodic behavior. We also propose an empirical system of ODEs representing the proportion of each type of particles and the segregation flux of particles. These equations reproduce the major features observed by the simulations.

  18. Thapsigargin, a tumor promoter, discharges intracellular Ca2+ stores by specific inhibition of the endoplasmic reticulum Ca2(+)-ATPase

    DEFF Research Database (Denmark)

    Thastrup, Ole; Cullen, P J; Drøbak, B K

    1990-01-01

    . This hypothesis is strongly supported by the demonstration that thapsigargin causes a rapid inhibition of the Ca2(+)-activated ATPase activity of rat liver microsomes, with an identical dose dependence to that seen in whole cell or isolated microsome Ca2+ discharge. The inhibition of the endoplasmic reticulum...

  19. Endoplasmic-reticulum-mediated microtubule alignment governs cytoplasmic streaming.

    Science.gov (United States)

    Kimura, Kenji; Mamane, Alexandre; Sasaki, Tohru; Sato, Kohta; Takagi, Jun; Niwayama, Ritsuya; Hufnagel, Lars; Shimamoto, Yuta; Joanny, Jean-François; Uchida, Seiichi; Kimura, Akatsuki

    2017-04-01

    Cytoplasmic streaming refers to a collective movement of cytoplasm observed in many cell types. The mechanism of meiotic cytoplasmic streaming (MeiCS) in Caenorhabditis elegans zygotes is puzzling as the direction of the flow is not predefined by cell polarity and occasionally reverses. Here, we demonstrate that the endoplasmic reticulum (ER) network structure is required for the collective flow. Using a combination of RNAi, microscopy and image processing of C. elegans zygotes, we devise a theoretical model, which reproduces and predicts the emergence and reversal of the flow. We propose a positive-feedback mechanism, where a local flow generated along a microtubule is transmitted to neighbouring regions through the ER. This, in turn, aligns microtubules over a broader area to self-organize the collective flow. The proposed model could be applicable to various cytoplasmic streaming phenomena in the absence of predefined polarity. The increased mobility of cortical granules by MeiCS correlates with the efficient exocytosis of the granules to protect the zygotes from osmotic and mechanical stresses.

  20. Residential Segregation in Texas in 1980.

    Science.gov (United States)

    Hwang, Sean-Shong; Murdock, Steve H.

    1982-01-01

    Between 1970 and 1980 racial and ethnic segregation for major Texas cities declined for all groups, but declines were small between Anglo and Spanish groups. Segregation is unaffected by variation in size of city, percent of population that is Spanish or Black, or central city status. (Author/AM)

  1. Housing Systems and Ethnic Spatial Segregation

    DEFF Research Database (Denmark)

    Andersen, Hans Skifter; Andersson, Roger; Wessel, Terje

    Residential spatial segregation is related to housing markets and housing policies. In this paper, ethnic segregation is compared across four Nordic capitals and explanations for the differences are examined by comparing the housing markets and housing policies of the countries. The housing markets...

  2. Losing Ground: School Segregation in Massachuestts

    Science.gov (United States)

    Ayscue, Jennifer B.; Greenberg, Alyssa

    2013-01-01

    Though once a leader in school integration, Massachusetts has regressed over the last two decades as its students of color have experienced intensifying school segregation. This report investigates trends in school segregation in Massachusetts by examining concentration, exposure, and evenness measures by both race and class. First, the report…

  3. Occupational Segregation by Sex: Determinants and Changes.

    Science.gov (United States)

    Beller, Andrea H.

    1982-01-01

    This study found that occupational sex segregation began to diminish during the 1970s, in conjunction with enforcement of the equal employment opportunity laws against sex discrimination in employment. The success of these laws suggests that discrimination was originally a determinant of occupational segregation. (Author/SK)

  4. Sex Segregation in Undergraduate Engineering Majors

    Science.gov (United States)

    Litzler, Elizabeth

    2010-01-01

    Gender inequality in engineering persists in spite of women reaching parity in college enrollments and degrees granted. To date, no analyses of educational sex segregation have comprehensively examined segregation within one discipline. To move beyond traditional methods of studying the long-standing stratification by field of study in higher…

  5. Administrative Segregation for Mentally Ill Inmates

    Science.gov (United States)

    O'Keefe, Maureen L.

    2007-01-01

    Largely the result of prison officials needing to safely and efficiently manage a volatile inmate population, administrative segregation or supermax facilities are criticized as violating basic human needs, particularly for mentally ill inmates. The present study compared Colorado offenders with mental illness (OMIs) to nonOMIs in segregated and…

  6. Class, Kinship Density, and Conjugal Role Segregation.

    Science.gov (United States)

    Hill, Malcolm D.

    1988-01-01

    Studied conjugal role segregation in 150 married women from intact families in working-class community. Found that, although involvement in dense kinship networks was associated with conjugal role segregation, respondents' attitudes toward marital roles and phase of family cycle when young children were present were more powerful predictors of…

  7. Towards understanding the molecular basis of bacterial DNA segregation

    DEFF Research Database (Denmark)

    Leonard, Thomas A.; Møller-Jensen, Jakob; Löwe, Jan

    2005-01-01

    Bacteria ensure the fidelity of genetic inheritance by the coordinated control of chromosome segregation and cell division. Here, we review the molecules and mechanisms that govern the correct subcellular positioning and rapid separation of newly replicated chromosomes and plasmids towards the ce...... common to the two processes. Finally, we discuss the role that the bacterial cytoskeleton plays in DNA partitioning and the missing link between prokaryotes and eukaryotes that is bacterial mechano-chemical motor proteins. Udgivelsesdato: Mar 29...

  8. The Role of Endoplasmic Reticulum Stress in Diabetic Nephropathy.

    Science.gov (United States)

    Fan, Ying; Lee, Kyung; Wang, Niansong; He, John Cijiang

    2017-03-01

    Diabetic nephropathy (DN) has become the leading cause of end-stage renal disease (ESRD) worldwide. Accumulating evidence suggests that endoplasmic reticulum (ER) stress plays a major role in the development and progression of DN. Recent findings suggested that many attributes of DN, such as hyperglycemia, proteinuria, and increased advanced glycation end products and free fatty acids, can all trigger unfolded protein response (UPR) in kidney cells. Herein, we review the current knowledge on the role of ER stress in the setting of kidney injury with a specific emphasis on DN. As maladaptive ER stress response caused by excessively prolonged UPR will eventually cause cell death and increase kidney injury, several ER stress inhibitors have been shown to improve DN in animal models, albeit blocking both adaptive and maladaptive UPR. More recently, reticulon-1A (RTN1A), an ER-associated protein, was shown to be increased in both human and mouse diabetic kidneys. Its expression correlates with the progression of DN, and its polymorphisms are associated with kidney disease in people with diabetes. Increased RTN1A expression heightened the ER stress response and renal cell apoptosis, and conversely reduced RTN1A in renal cells decreased apoptosis and ameliorated kidney injury and DN progression, suggesting that RTN1A may be a novel target to specifically restrain the maladaptive UPR. These findings suggest that ER stress response in renal cells is a key driver of progression of DN and that the inhibition of the unchecked ER stress response in DN, such as by inhibition of RTN1A function, may be a promising therapeutic approach against DN.

  9. Grain boundary segregation and intergranular failure

    International Nuclear Information System (INIS)

    White, C.L.

    1980-01-01

    Trace elements and impurities often segregate strongly to grain boundaries in metals and alloys. Concentrations of these elements at grain boundaries are often 10 3 to 10 5 times as great as their overall concentration in the alloy. Because of such segregation, certain trace elements can exert a disproportionate influence on material properties. One frequently observed consequence of trace element segregation to grain boundaries is the occurrence of grain boundary failure and low ductility. Less well known are incidences of improved ductility and inhibition of grain boundary fracture resulting from trace element segregation to grain boundaries in certain systems. An overview of trace element segregation and intergranular failure in a variety of alloy systems as well as preliminary results from studies on Al 3% Li will be presented

  10. Regulation of endoplasmic reticulum turnover by selective autophagy

    NARCIS (Netherlands)

    Khaminets, Aliaksandr; Heinrich, Theresa; Mari, Muriel; Grumati, Paolo; Huebner, Antje K; Akutsu, Masato; Liebmann, Lutz; Stolz, Alexandra; Nietzsche, Sandor; Koch, Nicole; Mauthe, Mario; Katona, Istvan; Qualmann, Britta; Weis, Joachim; Reggiori, Fulvio; Kurth, Ingo; Hübner, Christian A; Dikic, Ivan

    2015-01-01

    The endoplasmic reticulum (ER) is the largest intracellular endomembrane system, enabling protein and lipid synthesis, ion homeostasis, quality control of newly synthesized proteins and organelle communication. Constant ER turnover and modulation is needed to meet different cellular requirements and

  11. Regulation of endoplasmic reticulum turnover by selective autophagy

    NARCIS (Netherlands)

    Khaminets, Aliaksandr; Heinrich, Theresa; Mari, Muriel; Grumati, Paolo; Huebner, Antje K.; Akutsu, Masato; Liebmann, Lutz; Stolz, Alexandra; Nietzsche, Sandor; Koch, Nicole; Mauthe, Mario; Katona, Istvan; Qualmann, Britta; Weis, Joachim; Reggiori, Fulvio; Kurth, Ingo; Huebner, Christian A.; Dikic, Ivan

    2015-01-01

    The endoplasmic reticulum (ER) is the largest intracellular endomembrane system, enabling protein and lipid synthesis, ion homeostasis, quality control of newly synthesized proteins and organelle communication(1). Constant ER turnover and modulation is needed to meet different cellular requirements

  12. CENP-A regulates chromosome segregation during the first meiosis of mouse oocytes.

    Science.gov (United States)

    Li, Li; Qi, Shu-Tao; Sun, Qing-Yuan; Chen, Shi-Ling

    2017-06-01

    Proper chromosome separation in both mitosis and meiosis depends on the correct connection between kinetochores of chromosomes and spindle microtubules. Kinetochore dysfunction can lead to unequal distribution of chromosomes during cell division and result in aneuploidy, thus kinetochores are critical for faithful segregation of chromosomes. Centromere protein A (CENP-A) is an important component of the inner kinetochore plate. Multiple studies in mitosis have found that deficiencies in CENP-A could result in structural and functional changes of kinetochores, leading to abnormal chromosome segregation, aneuploidy and apoptosis in cells. Here we report the expression and function of CENP-A during mouse oocyte meiosis. Our study found that microinjection of CENP-A blocking antibody resulted in errors of homologous chromosome segregation and caused aneuploidy in eggs. Thus, our findings provide evidence that CENP-A is critical for the faithful chromosome segregation during mammalian oocyte meiosis.

  13. Structural polarity and dynamics of male germline stem cells in the milkweed bug (Oncopeltus fasciatus).

    Science.gov (United States)

    Schmidt, Esther D; Dorn, August

    2004-11-01

    The male germline stem cells (GSCs) of the milkweed bug present an extraordinary structural polarity that is, to our knowledge, unequalled by any other type of stem cells. They consist of a perikaryon and numerous projections arising from the cell pole directed toward the apical cells, the proposed niche of the GSCs. The projections can traverse a considerable distance until their terminals touch the apical cells. From hatching until death, the GSC projections undergo conspicuous changes, the sequence of which has been deduced from observations of all developmental stages. Projection formation starts from lobular cell protrusions showing trabecular ingrowths of the cell membrane. Finger-like projections result from a process of growth and "carving out". The newly formed projections contain mostly only free ribosomes other than a few mitochondria. A stereotyped degradation process commences in the projection terminals: profiles of circular, often concentric, cisternae of rough endoplasmic reticulum appear and turn into myelin bodies, whereas mitochondria become more numerous. The cytoplasm vesiculates, lysosomal bodies appear, and mitochondria become swollen. At the same time, the projection terminals are segregated by transverse ingrowths of the cell membrane. Finally, autophagic vacuoles and myelin bodies fill the segregated terminals, which then rupture. Simultaneously, new projections seem to sprout from the perikaryon of the GSCs. These dynamics, which are not synchronized among the GSCs, indicate that a novel type of signal exchange and transduction between the stem cells and their niche is involved in the regulation of asymmetric versus symmetric division of GSCs.

  14. Endoplasmic reticulum stress in wake-active neurons progresses with aging.

    Science.gov (United States)

    Naidoo, Nirinjini; Zhu, Jingxu; Zhu, Yan; Fenik, Polina; Lian, Jie; Galante, Ray; Veasey, Sigrid

    2011-08-01

    Fragmentation of wakefulness and sleep are expected outcomes of advanced aging. We hypothesize that wake neurons develop endoplasmic reticulum dyshomeostasis with aging, in parallel with impaired wakefulness. In this series of experiments, we sought to more fully characterize age-related changes in wakefulness and then, in relevant wake neuronal populations, explore functionality and endoplasmic reticulum homeostasis. We report that old mice show greater sleep/wake transitions in the active period with markedly shortened wake periods, shortened latencies to sleep, and less wake time in the subjective day in response to a novel social encounter. Consistent with sleep/wake instability and reduced social encounter wakefulness, orexinergic and noradrenergic wake neurons in aged mice show reduced c-fos response to wakefulness and endoplasmic reticulum dyshomeostasis with increased nuclear translocation of CHOP and GADD34. We have identified an age-related unfolded protein response injury to and dysfunction of wake neurons. It is anticipated that these changes contribute to sleep/wake fragmentation and cognitive impairment in aging. © 2011 The Authors. Aging Cell © 2011 Blackwell Publishing Ltd/Anatomical Society of Great Britain and Ireland.

  15. The quality control of glycoprotein folding in the endoplasmic reticulum, a trip from trypanosomes to mammals

    Directory of Open Access Journals (Sweden)

    A.J. Parodi

    1998-05-01

    Full Text Available The present review deals with the stages of synthesis and processing of asparagine-linked oligosaccharides occurring in the lumen of the endoplasmic reticulum and their relationship to the acquisition by glycoproteins of their proper tertiary structures. Special emphasis is placed on reactions taking place in trypanosomatid protozoa since their study has allowed the detection of the transient glucosylation of glycoproteins catalyzed by UDP-Glc:glycoprotein glucosyltransferase and glucosidase II. The former enzyme has the unique property of covalently tagging improperly folded conformations by catalyzing the formation of protein-linked Glc1Man7GlcNAc2, Glc1Man8GlcNac2 and Glc1Man9GlcNAc2 from the unglucosylated proteins. Glucosyltransferase is a soluble protein of the endoplasmic reticulum that recognizes protein domains exposed in denatured but not in native conformations (probably hydrophobic amino acids and the innermost N-acetylglucosamine unit that is hidden from macromolecular probes in most native glycoproteins. In vivo, the glucose units are removed by glucosidase II. The influence of oligosaccharides in glycoprotein folding is reviewed as well as the participation of endoplasmic reticulum chaperones (calnexin and calreticulin that recognize monoglucosylated species in the same process. A model for the quality control of glycoprotein folding in the endoplasmic reticulum, i.e., the mechanism by which cells recognize the tertiary structure of glycoproteins and only allow transit to the Golgi apparatus of properly folded species, is discussed. The main elements of this control are calnexin and calreticulin as retaining components, the UDP-Glc:glycoprotein glucosyltransferase as a sensor of tertiary structures and glucosidase II as the releasing agent.

  16. Purification of a sarcoplasmic reticulum protein that binds Ca2+ and plasma lipoproteins

    International Nuclear Information System (INIS)

    Hofmann, S.L.; Brown, M.S.; Lee, E.; Pathak, R.K.; Anderson, R.G.; Goldstein, J.L.

    1989-01-01

    A protein in the sarcoplasmic reticulum of rabbit skeletal and cardiac muscle was identified because of its ability to bind 125I-labeled low density lipoprotein (LDL) with high affinity after sodium dodecyl sulfate-polyacrylamide gel electrophoresis. This protein, referred to as the 165-kDa protein, is restricted to striated muscle. It was not detected in 14 other tissues, including several that contain smooth muscle, but it appears in rat L6 myoblasts when they differentiate into myocytes. Immunofluorescence and immunoelectron microscopic studies revealed that the protein is present throughout the sarcoplasmic reticulum and the terminal cisternae. It binds 45Ca2+ on nitrocellulose blots and stains metachromatically with Stains-all, a cationic dye that stains Ca2+-binding proteins. It does not appear to be a glycoprotein, and it appears slightly larger than the 160-kDa glycoprotein previously described in sarcoplasmic reticulum. The 165-kDa protein binds LDL, beta-migrating very low density lipoprotein, and a cholesterol-induced high density lipoprotein particle that contains apoprotein E as its sole apoprotein with much higher affinity than it binds high density lipoprotein. The protein is stable to boiling and to treatment with sodium dodecyl sulfate, but it becomes sensitive to these treatments when its cystine residues are reduced and alkylated. The protein was purified 1300-fold to apparent homogeneity from rabbit skeletal muscle membranes. It differs from the cell surface LDL receptor in that (1) its apparent molecular weight is not changed by reduction and alkylation; (2) it is present in Watanabe-heritable hyperlipidemic rabbits, which lack functional LDL receptors; (3) binding of lipoproteins is not inhibited by EDTA; and (4) it is located within the lumen of the sarcoplasmic reticulum where it has no access to plasma lipoproteins

  17. STIM proteins and the endoplasmic reticulum-plasma membrane junctions.

    Science.gov (United States)

    Carrasco, Silvia; Meyer, Tobias

    2011-01-01

    Eukaryotic organelles can interact with each other through stable junctions where the two membranes are kept in close apposition. The junction that connects the endoplasmic reticulum to the plasma membrane (ER-PM junction) is unique in providing a direct communication link between the ER and the PM. In a recently discovered signaling process, STIM (stromal-interacting molecule) proteins sense a drop in ER Ca(2+) levels and directly activate Orai PM Ca(2+) channels across the junction space. In an inverse process, a voltage-gated PM Ca(2+) channel can directly open ER ryanodine-receptor Ca(2+) channels in striated-muscle cells. Although ER-PM junctions were first described 50 years ago, their broad importance in Ca(2+) signaling, as well as in the regulation of cholesterol and phosphatidylinositol lipid transfer, has only recently been realized. Here, we discuss research from different fields to provide a broad perspective on the structures and unique roles of ER-PM junctions in controlling signaling and metabolic processes.

  18. Rapid mass segregation in small stellar clusters

    Science.gov (United States)

    Spera, Mario; Capuzzo-Dolcetta, Roberto

    2017-12-01

    In this paper we focus our attention on small-to-intermediate N-body systems that are, initially, distributed uniformly in space and dynamically `cool' (virial ratios Q=2T/|Ω| below ˜0.3). In this work, we study the mass segregation that emerges after the initial violent dynamical evolution. At this scope, we ran a set of high precision N-body simulations of isolated clusters by means of HiGPUs, our direct summation N-body code. After the collapse, the system shows a clear mass segregation. This (quick) mass segregation occurs in two phases: the first shows up in clumps originated by sub-fragmentation before the deep overall collapse; this segregation is partly erased during the deep collapse to re-emerge, abruptly, during the second phase, that follows the first bounce of the system. In this second stage, the proper clock to measure the rate of segregation is the dynamical time after virialization, which (for cold and cool systems) may be significantly different from the crossing time evaluated from initial conditions. This result is obtained for isolated clusters composed of stars of two different masses (in the ratio mh/ml=2), at varying their number ratio, and is confirmed also in presence of a massive central object (simulating a black hole of stellar size). Actually, in stellar systems starting their dynamical evolution from cool conditions, the fast mass segregation adds to the following, slow, secular segregation which is collisionally induced. The violent mass segregation is an effect persistent over the whole range of N (128 ≤ N ≤1,024) investigated, and is an interesting feature on the astronomical-observational side, too. The semi-steady state reached after virialization corresponds to a mass segregated distribution function rather than that of equipartition of kinetic energy per unit mass as it should result from violent relaxation.

  19. Spillover-mediated feedforward-inhibition functionally segregates interneuron activity

    Science.gov (United States)

    Coddington, Luke T.; Rudolph, Stephanie; Lune, Patrick Vande; Overstreet-Wadiche, Linda; Wadiche, Jacques I.

    2013-01-01

    Summary Neurotransmitter spillover represents a form of neural transmission not restricted to morphologically defined synaptic connections. Communication between climbing fibers (CFs) and molecular layer interneurons (MLIs) in the cerebellum is mediated exclusively by glutamate spillover. Here, we show how CF stimulation functionally segregates MLIs based on their location relative to glutamate release. Excitation of MLIs that reside within the domain of spillover diffusion coordinates inhibition of MLIs outside the diffusion limit. CF excitation of MLIs is dependent on extrasynaptic NMDA receptors that enhance the spatial and temporal spread of CF signaling. Activity mediated by functionally segregated MLIs converges onto neighboring Purkinje cells (PCs) to generate a long-lasting biphasic change in inhibition. These data demonstrate how glutamate release from single CFs modulates excitability of neighboring PCs, thus expanding the influence of CFs on cerebellar cortical activity in a manner not predicted by anatomical connectivity. PMID:23707614

  20. Genes that bias Mendelian segregation.

    Science.gov (United States)

    Grognet, Pierre; Lalucque, Hervé; Malagnac, Fabienne; Silar, Philippe

    2014-01-01

    Mendel laws of inheritance can be cheated by Meiotic Drive Elements (MDs), complex nuclear genetic loci found in various eukaryotic genomes and distorting segregation in their favor. Here, we identify and characterize in the model fungus Podospora anserina Spok1 and Spok2, two MDs known as Spore Killers. We show that they are related genes with both spore-killing distorter and spore-protecting responder activities carried out by the same allele. These alleles act as autonomous elements, exert their effects independently of their location in the genome and can act as MDs in other fungi. Additionally, Spok1 acts as a resistance factor to Spok2 killing. Genetical data and cytological analysis of Spok1 and Spok2 localization during the killing process suggest a complex mode of action for Spok proteins. Spok1 and Spok2 belong to a multigene family prevalent in the genomes of many ascomycetes. As they have no obvious cellular role, Spok1 and Spok2 Spore Killer genes represent a novel kind of selfish genetic elements prevalent in fungal genome that proliferate through meiotic distortion.

  1. Genes that bias Mendelian segregation.

    Directory of Open Access Journals (Sweden)

    Pierre Grognet

    Full Text Available Mendel laws of inheritance can be cheated by Meiotic Drive Elements (MDs, complex nuclear genetic loci found in various eukaryotic genomes and distorting segregation in their favor. Here, we identify and characterize in the model fungus Podospora anserina Spok1 and Spok2, two MDs known as Spore Killers. We show that they are related genes with both spore-killing distorter and spore-protecting responder activities carried out by the same allele. These alleles act as autonomous elements, exert their effects independently of their location in the genome and can act as MDs in other fungi. Additionally, Spok1 acts as a resistance factor to Spok2 killing. Genetical data and cytological analysis of Spok1 and Spok2 localization during the killing process suggest a complex mode of action for Spok proteins. Spok1 and Spok2 belong to a multigene family prevalent in the genomes of many ascomycetes. As they have no obvious cellular role, Spok1 and Spok2 Spore Killer genes represent a novel kind of selfish genetic elements prevalent in fungal genome that proliferate through meiotic distortion.

  2. Veil: A Wall of Segregation

    Directory of Open Access Journals (Sweden)

    Tayebeh Nowrouzi

    2015-08-01

    Full Text Available Moving behind the confines of the race has been the continuous efforts of African-Americans so as to reveal and confirm their true humanity and abilities to white race as well as their own race. African-Americans, Dubois posited, are shut out of the white America, inhabiting behind a vast veil which creates a deep division between the races. Veil is made of the fabric of racism interwoven thread by thread and imposed by white world. It is thrown discourteously and forcibly to the African-Americans whom their distorted images are imposed on them and their true humanity and identity are hidden behind the veil. This study overtakes to present how Loraine Hansberry, in her first and the most outstanding drama, A Raisin in the Sun examines the world within the veil. She demonstrated that Duboisian metaphoric veil is operating in the racist American society so that not only African-Americans are segregated physically and psychologically from the rest of the world but also are inflicted with obscurity of vision that are neither able to see themselves clearly nor be seen truly. On the other hand, it presents how the veil provides blacks with the second sight to observe and comprehend the racist nature of whites which is hidden and incomprehensible for them.

  3. Full-length Ebola glycoprotein accumulates in the endoplasmic reticulum

    Directory of Open Access Journals (Sweden)

    Bhattacharyya Suchita

    2011-01-01

    Full Text Available Abstract The Filoviridae family comprises of Ebola and Marburg viruses, which are known to cause lethal hemorrhagic fever. However, there is no effective anti-viral therapy or licensed vaccines currently available for these human pathogens. The envelope glycoprotein (GP of Ebola virus, which mediates entry into target cells, is cytotoxic and this effect maps to a highly glycosylated mucin-like region in the surface subunit of GP (GP1. However, the mechanism underlying this cytotoxic property of GP is unknown. To gain insight into the basis of this GP-induced cytotoxicity, HEK293T cells were transiently transfected with full-length and mucin-deleted (Δmucin Ebola GP plasmids and GP localization was examined relative to the nucleus, endoplasmic reticulum (ER, Golgi, early and late endosomes using deconvolution fluorescent microscopy. Full-length Ebola GP was observed to accumulate in the ER. In contrast, GPΔmucin was uniformly expressed throughout the cell and did not localize in the ER. The Ebola major matrix protein VP40 was also co-expressed with GP to investigate its influence on GP localization. GP and VP40 co-expression did not alter GP localization to the ER. Also, when VP40 was co-expressed with the nucleoprotein (NP, it localized to the plasma membrane while NP accumulated in distinct cytoplasmic structures lined with vimentin. These latter structures are consistent with aggresomes and may serve as assembly sites for filoviral nucleocapsids. Collectively, these data suggest that full-length GP, but not GPΔmucin, accumulates in the ER in close proximity to the nuclear membrane, which may underscore its cytotoxic property.

  4. Endoplasmic reticulum stress-mediated neuronal apoptosis by acrylamide exposure

    Energy Technology Data Exchange (ETDEWEB)

    Komoike, Yuta, E-mail: komoike@research.twmu.ac.jp; Matsuoka, Masato, E-mail: matsuoka@research.twmu.ac.jp

    2016-11-01

    Acrylamide (AA) is a well-known neurotoxic compound in humans and experimental animals. However, intracellular stress signaling pathways responsible for the neurotoxicity of AA are still not clear. In this study, we explored the involvement of the endoplasmic reticulum (ER) stress response in AA-induced neuronal damage in vitro and in vivo. Exposure of SH-SY5Y human neuroblastoma cells to AA increased the levels of phosphorylated form of eukaryotic translation initiation factor 2α (eIF2α) and its downstream effector, activating transcription factor 4 (ATF4), indicating the induction of the unfolded protein response (UPR) by AA exposure. Furthermore, AA exposure increased the mRNA level of c/EBP homologous protein (CHOP), the ER stress-dependent apoptotic factor, and caused the accumulation of reactive oxygen species (ROS) in SH-SY5Y cells. Treatments of SH-SY5Y cells with the chemical chaperone, 4-phenylbutyric acid and the ROS scavenger, N-acetyl-cysteine reduced the AA-induced expression of ATF4 protein and CHOP mRNA, and resulted in the suppression of apoptosis. In addition, AA-induced eIF2α phosphorylation was also suppressed by NAC treatment. In consistent with in vitro study, exposure of zebrafish larvae at 6-day post fertilization to AA induced the expression of chop mRNA and apoptotic cell death in the brain, and also caused the disruption of brain structure. These findings suggest that AA exposure induces apoptotic neuronal cell death through the ER stress and subsequent eIF2α–ATF4–CHOP signaling cascade. The accumulation of ROS by AA exposure appears to be responsible for this ER stress-mediated apoptotic pathway. - Highlights: • Exposure of SH-SY5Y cells to AA activates the eIF2α–ATF4 pathway of the UPR. • Exposure of SH-SY5Y cells to AA induces the CHOP expression and apoptosis. • Exposure of zebrafish to AA induces the chop expression and apoptosis in the brain. • AA possibly induces apoptotic neuronal cell death through the ER

  5. CDIP1-BAP31 Complex Transduces Apoptotic Signals from Endoplasmic Reticulum to Mitochondria under Endoplasmic Reticulum Stress

    Directory of Open Access Journals (Sweden)

    Takushi Namba

    2013-10-01

    Full Text Available Resolved endoplasmic reticulum (ER stress response is essential for intracellular homeostatic balance, but unsettled ER stress can lead to apoptosis. Here, we show that a proapoptotic p53 target, CDIP1, acts as a key signal transducer of ER-stress-mediated apoptosis. We identify B-cell-receptor-associated protein 31 (BAP31 as an interacting partner of CDIP1. Upon ER stress, CDIP1 is induced and enhances an association with BAP31 at the ER membrane. We also show that CDIP1 binding to BAP31 is required for BAP31 cleavage upon ER stress and for BAP31-Bcl-2 association. The recruitment of Bcl-2 to the BAP31-CDIP1 complex, as well as CDIP1-dependent truncated Bid (tBid and caspase-8 activation, contributes to BAX oligomerization. Genetic knockout of CDIP1 in mice leads to impaired response to ER-stress-mediated apoptosis. Altogether, our data demonstrate that the CDIP1/BAP31-mediated regulation of mitochondrial apoptosis pathway represents a mechanism for establishing an ER-mitochondrial crosstalk for ER-stress-mediated apoptosis signaling.

  6. The involvement of SMILE/TMTC3 in endoplasmic reticulum stress response.

    Directory of Open Access Journals (Sweden)

    Maud Racapé

    Full Text Available The state of operational tolerance has been detected sporadically in some renal transplanted patients that stopped immunosuppressive drugs, demonstrating that allograft tolerance might exist in humans. Several years ago, a study by Brouard et al. identified a molecular signature of several genes that were significantly differentially expressed in the blood of such patients compared with patients with other clinical situations. The aim of the present study is to analyze the role of one of these molecules over-expressed in the blood of operationally tolerant patients, SMILE or TMTC3, a protein whose function is still unknown.We first confirmed that SMILE mRNA is differentially expressed in the blood of operationally tolerant patients with drug-free long term graft function compared to stable and rejecting patients. Using a yeast two-hybrid approach and a colocalization study by confocal microscopy we furthermore report an interaction of SMILE with PDIA3, a molecule resident in the endoplasmic reticulum (ER. In accordance with this observation, SMILE silencing in HeLa cells correlated with the modulation of several transcripts involved in proteolysis and a decrease in proteasome activity. Finally, SMILE silencing increased HeLa cell sensitivity to the proteasome inhibitor Bortezomib, a drug that induces ER stress via protein overload, and increased transcript expression of a stress response protein, XBP-1, in HeLa cells and keratinocytes.In this study we showed that SMILE is involved in the endoplasmic reticulum stress response, by modulating proteasome activity and XBP-1 transcript expression. This function of SMILE may influence immune cell behavior in the context of transplantation, and the analysis of endoplasmic reticulum stress in transplantation may reveal new pathways of regulation in long-term graft acceptance thereby increasing our understanding of tolerance.

  7. TMAP/CKAP2 is essential for proper chromosome segregation.

    Science.gov (United States)

    Hong, Kyung Uk; Kim, Eunhee; Bae, Chang-Dae; Park, Joobae

    2009-01-15

    Tumor-associated microtubule-associated protein (TMAP), also known as cytoskeleton associated protein 2 (CKAP2), is a novel mitotic spindle-associated protein which is frequently up-regulated in various malignances. However, its cellular functions remain unknown. Previous reports suggested that the cellular functions of TMAP/CKAP2 pertain to regulation of the dynamics and assembly of the mitotic spindle. To investigate its role in mitosis, we studied the effects of siRNA-mediated depletion of TMAP/CKAP2 in cultured mammalian cells. Unexpectedly, TMAP/CKAP2 knockdown did not result in significant alterations of the spindle apparatus. However, TMAP/CKAP2-depleted cells often exhibited abnormal nuclear morphologies, which were accompanied by abnormal organization of the nuclear lamina, and chromatin bridge formation between two daughter cell nuclei. Time lapse video microscopy revealed that the changes in nuclear morphology and chromatin bridge formations observed in TMAP/CKAP2-depleted cells are the result of defects in chromosome segregation. Consistent with this, the spindle checkpoint activity was significantly reduced in TMAP/CKAP2-depleted cells. Moreover, chromosome missegregation induced by depletion of TMAP/CKAP2 ultimately resulted in reduced cell viability and increased chromosomal instability. Our present findings demonstrate that TMAP/CKAP2 is essential for proper chromosome segregation and for maintaining genomic stability.

  8. Silver nanoparticles induce endoplasmatic reticulum stress response in zebrafish

    Energy Technology Data Exchange (ETDEWEB)

    Christen, Verena [University of Applied Sciences and Arts Northwestern Switzerland, School of Life Sciences, Gründenstrasse 40, CH-4132 Muttenz (Switzerland); Capelle, Martinus [Crucell, P.O. Box 2048, NL-2301 Leiden (Netherlands); Fent, Karl, E-mail: karl.fent@fhnw.ch [University of Applied Sciences and Arts Northwestern Switzerland, School of Life Sciences, Gründenstrasse 40, CH-4132 Muttenz (Switzerland); Swiss Federal Institute of Technology Zürich, Department of Environmental Systems Science, CH-8092 Zürich (Switzerland)

    2013-10-15

    Silver nanoparticles (AgNPs) find increasing applications, and therefore humans and the environment are increasingly exposed to them. However, potential toxicological implications are not sufficiently known. Here we investigate effects of AgNPs (average size 120 nm) on zebrafish in vitro and in vivo, and compare them to human hepatoma cells (Huh7). AgNPs are incorporated in zebrafish liver cells (ZFL) and Huh7, and in zebrafish embryos. In ZFL cells AgNPs lead to induction of reactive oxygen species (ROS), endoplasmatic reticulum (ER) stress response, and TNF-α. Transcriptional alterations also occur in pro-apoptotic genes p53 and Bax. The transcriptional profile differed in ZFL and Huh7 cells. In ZFL cells, the ER stress marker BiP is induced, concomitant with the ER stress marker ATF-6 and spliced XBP-1 after 6 h and 24 h exposure to 0.5 g/L and 0.05 g/L AgNPs, respectively. This indicates the induction of different pathways of the ER stress response. Moreover, AgNPs induce TNF-α. In zebrafish embryos exposed to 0.01, 0.1, 1 and 5 mg/L AgNPs hatching was affected and morphological defects occurred at high concentrations. ER stress related gene transcripts BiP and Synv are significantly up-regulated after 24 h at 0.1 and 5 mg/L AgNPs. Furthermore, transcriptional alterations occurred in the pro-apoptotic genes Noxa and p21. The ER stress response was strong in ZFL cells and occurred in zebrafish embryos as well. Our data demonstrate for the first time that AgNPs lead to induction of ER stress in zebrafish. The induction of ER stress can have several consequences including the activation of apoptotic and inflammatory pathways. - Highlights: • Effects of silver nanoparticles (120 nm AgNPs) are investigated in zebrafish. • AgNPs induce all ER stress reponses in vitro in zebrafish liver cells. • AgNPs induce weak ER stress in zebrafish embryos. • AgNPs induce oxidative stress and transcripts of pro-apoptosis genes.

  9. Silver nanoparticles induce endoplasmatic reticulum stress response in zebrafish

    International Nuclear Information System (INIS)

    Christen, Verena; Capelle, Martinus; Fent, Karl

    2013-01-01

    Silver nanoparticles (AgNPs) find increasing applications, and therefore humans and the environment are increasingly exposed to them. However, potential toxicological implications are not sufficiently known. Here we investigate effects of AgNPs (average size 120 nm) on zebrafish in vitro and in vivo, and compare them to human hepatoma cells (Huh7). AgNPs are incorporated in zebrafish liver cells (ZFL) and Huh7, and in zebrafish embryos. In ZFL cells AgNPs lead to induction of reactive oxygen species (ROS), endoplasmatic reticulum (ER) stress response, and TNF-α. Transcriptional alterations also occur in pro-apoptotic genes p53 and Bax. The transcriptional profile differed in ZFL and Huh7 cells. In ZFL cells, the ER stress marker BiP is induced, concomitant with the ER stress marker ATF-6 and spliced XBP-1 after 6 h and 24 h exposure to 0.5 g/L and 0.05 g/L AgNPs, respectively. This indicates the induction of different pathways of the ER stress response. Moreover, AgNPs induce TNF-α. In zebrafish embryos exposed to 0.01, 0.1, 1 and 5 mg/L AgNPs hatching was affected and morphological defects occurred at high concentrations. ER stress related gene transcripts BiP and Synv are significantly up-regulated after 24 h at 0.1 and 5 mg/L AgNPs. Furthermore, transcriptional alterations occurred in the pro-apoptotic genes Noxa and p21. The ER stress response was strong in ZFL cells and occurred in zebrafish embryos as well. Our data demonstrate for the first time that AgNPs lead to induction of ER stress in zebrafish. The induction of ER stress can have several consequences including the activation of apoptotic and inflammatory pathways. - Highlights: • Effects of silver nanoparticles (120 nm AgNPs) are investigated in zebrafish. • AgNPs induce all ER stress reponses in vitro in zebrafish liver cells. • AgNPs induce weak ER stress in zebrafish embryos. • AgNPs induce oxidative stress and transcripts of pro-apoptosis genes

  10. Granular Segregation by an Oscillating Ratchet Mechanism

    International Nuclear Information System (INIS)

    Igarashi, A.; Horiuchi, Ch.

    2004-01-01

    We report on a method to segregate granular mixtures which consist of two kinds of particles by an oscillating ''ratchet'' mechanism. The segregation system has an asymmetrical sawtooth-shaped base which is vertically oscillating. Such a ratchet base produces a directional current of particles owing to its transport property. It is a counterintuitive and interesting phenomenon that a vertically vibrated base transports particles horizontally. This system is studied with numerical simulations, and it is found that we can apply such a system to segregation of mixtures of particles with different properties (radius or mass). Furthermore, we find out that an appropriate inclination of the ratchet-base makes the quality of segregation high. (author)

  11. Gender Differences in the Effect of Residential Segregation on Workplace Segregation among Newly Arrived Immigrants

    OpenAIRE

    Tammaru, Tiit; Strömgren, Magnus; van Ham, Maarten; Danzer, Alexander M.

    2015-01-01

    Contemporary cities are becoming more and more diverse in population as a result of immigration. Research also shows that within cities residential neighborhoods are becoming ethnically more diverse, but that residential segregation has remained persistently high. High levels of segregation are often seen as negative, preventing integration of immigrants in their host society and having a negative impact on people's lives. Segregation research often focuses on residential neighborhoods, but i...

  12. Gender Segregation in the Retail Industry

    OpenAIRE

    Lynch, Samantha

    2002-01-01

    This paper examines the phenomenon of occupational gender segregation in the retail industry, with a particular focus on part time working. The empirical data was gathered through a series of 59 interviews, and a small survey of employees, with store level managers in three UK retail organisations. The paper illustrates the extent of occupational gender segregation and considers the impact of such stereotyping on the gender pay gap, training and career development.\\ud \\ud Occupational gender ...

  13. Racial segregation patterns in selective universities

    OpenAIRE

    Peter Arcidiacono; Esteban M. Aucejo; Andrew Hussey; Kenneth Spenner

    2013-01-01

    This paper examines sorting into interracial friendships at selective universities. We show significant friendship segregation, particularly for blacks. Indeed, blacks' friendships are no more diverse in college than in high school, despite the fact that the colleges that blacks attend have substantially smaller black populations. We demonstrate that the segregation patterns occur in part because affirmative action results in large differences in the academic backgrounds of students of differ...

  14. Gender Segregation in the Spanish Labor Market: An Alternative Approach

    Science.gov (United States)

    del Rio, Coral; Alonso-Villar, Olga

    2010-01-01

    The aim of this paper is to study occupational segregation by gender in Spain, which is a country where occupational segregation explains a large part of the gender wage gap. As opposed to previous studies, this paper measures not only overall segregation, but also the segregation of several population subgroups. For this purpose, this paper uses…

  15. Blocking variant surface glycoprotein synthesis alters endoplasmic reticulum exit sites/Golgi homeostasis in Trypanosoma brucei.

    Science.gov (United States)

    Ooi, Cher-Pheng; Smith, Terry K; Gluenz, Eva; Wand, Nadina Vasileva; Vaughan, Sue; Rudenko, Gloria

    2018-06-01

    The predominant secretory cargo of bloodstream form Trypanosoma brucei is variant surface glycoprotein (VSG), comprising ~10% total protein and forming a dense protective layer. Blocking VSG translation using Morpholino oligonucleotides triggered a precise pre-cytokinesis arrest. We investigated the effect of blocking VSG synthesis on the secretory pathway. The number of Golgi decreased, particularly in post-mitotic cells, from 3.5 ± 0.6 to 2.0 ± 0.04 per cell. Similarly, the number of endoplasmic reticulum exit sites (ERES) in post-mitotic cells dropped from 3.9 ± 0.6 to 2.7 ± 0.1 eight hours after blocking VSG synthesis. The secretory pathway was still functional in these stalled cells, as monitored using Cathepsin L. Rates of phospholipid and glycosylphosphatidylinositol-anchor biosynthesis remained relatively unaffected, except for the level of sphingomyelin which increased. However, both endoplasmic reticulum and Golgi morphology became distorted, with the Golgi cisternae becoming significantly dilated, particularly at the trans-face. Membrane accumulation in these structures is possibly caused by reduced budding of nascent vesicles due to the drastic reduction in the total amount of secretory cargo, that is, VSG. These data argue that the total flux of secretory cargo impacts upon the biogenesis and maintenance of secretory structures and organelles in T. brucei, including the ERES and Golgi. © 2018 The Authors. Traffic published by John Wiley & Sons Ltd.

  16. Figure-ground segregation: A fully nonlocal approach.

    Science.gov (United States)

    Dimiccoli, Mariella

    2016-09-01

    We present a computational model that computes and integrates in a nonlocal fashion several configural cues for automatic figure-ground segregation. Our working hypothesis is that the figural status of each pixel is a nonlocal function of several geometric shape properties and it can be estimated without explicitly relying on object boundaries. The methodology is grounded on two elements: multi-directional linear voting and nonlinear diffusion. A first estimation of the figural status of each pixel is obtained as a result of a voting process, in which several differently oriented line-shaped neighborhoods vote to express their belief about the figural status of the pixel. A nonlinear diffusion process is then applied to enforce the coherence of figural status estimates among perceptually homogeneous regions. Computer simulations fit human perception and match the experimental evidence that several cues cooperate in defining figure-ground segregation. The results of this work suggest that figure-ground segregation involves feedback from cells with larger receptive fields in higher visual cortical areas. Copyright © 2015 Elsevier Ltd. All rights reserved.

  17. Suppressed Sr segregation and performance of directly assembled La0.6Sr0.4Co0.2Fe0.8O3-δ oxygen electrode on Y2O3-ZrO2 electrolyte of solid oxide electrolysis cells

    Science.gov (United States)

    Ai, Na; He, Shuai; Li, Na; Zhang, Qi; Rickard, William D. A.; Chen, Kongfa; Zhang, Teng; Jiang, San Ping

    2018-04-01

    Active and stable oxygen electrode is probably the most important in the development of solid oxide electrolysis cells (SOECs) technologies. Herein, we report the successful development of mixed ionic and electronic conducting (MIEC) La0.6Sr0.4Co0.2Fe0.8O3-δ (LSCF) perovskite oxides directly assembled on barrier-layer-free yttria-stabilized zirconia (YSZ) electrolyte as highly active and stable oxygen electrodes of SOECs. Electrolysis polarization effectively induces the formation of electrode/electrolyte interface, similar to that observed under solid oxide fuel cell (SOFC) operation conditions. However, in contrast to the significant performance decay under SOFC operation conditions, the cell with directly assembled LSCF oxygen electrodes shows excellent stability, tested for 300 h at 0.5 A cm-2 and 750 °C under SOEC operation conditions. Detailed microstructure and phase analysis reveal that Sr segregation is inevitable for LSCF electrode, but anodic polarization substantially suppresses Sr segregation and migration to the electrode/electrolyte interface, leading to the formation of stable and efficient electrode/electrolyte interface for water and CO2 electrolysis under SOECs operation conditions. The present study demonstrates the feasibility of using directly assembled MIEC cobaltite based oxygen electrodes on barrier-layer-free YSZ electrolyte of SOECs.

  18. Neuromorphic VLSI vision system for real-time texture segregation.

    Science.gov (United States)

    Shimonomura, Kazuhiro; Yagi, Tetsuya

    2008-10-01

    The visual system of the brain can perceive an external scene in real-time with extremely low power dissipation, although the response speed of an individual neuron is considerably lower than that of semiconductor devices. The neurons in the visual pathway generate their receptive fields using a parallel and hierarchical architecture. This architecture of the visual cortex is interesting and important for designing a novel perception system from an engineering perspective. The aim of this study is to develop a vision system hardware, which is designed inspired by a hierarchical visual processing in V1, for real time texture segregation. The system consists of a silicon retina, orientation chip, and field programmable gate array (FPGA) circuit. The silicon retina emulates the neural circuits of the vertebrate retina and exhibits a Laplacian-Gaussian-like receptive field. The orientation chip selectively aggregates multiple pixels of the silicon retina in order to produce Gabor-like receptive fields that are tuned to various orientations by mimicking the feed-forward model proposed by Hubel and Wiesel. The FPGA circuit receives the output of the orientation chip and computes the responses of the complex cells. Using this system, the neural images of simple cells were computed in real-time for various orientations and spatial frequencies. Using the orientation-selective outputs obtained from the multi-chip system, a real-time texture segregation was conducted based on a computational model inspired by psychophysics and neurophysiology. The texture image was filtered by the two orthogonally oriented receptive fields of the multi-chip system and the filtered images were combined to segregate the area of different texture orientation with the aid of FPGA. The present system is also useful for the investigation of the functions of the higher-order cells that can be obtained by combining the simple and complex cells.

  19. Respiratory metabolism and calorie restriction relieve persistent endoplasmic reticulum stress induced by calcium shortage in yeast

    DEFF Research Database (Denmark)

    Busti, Stefano; Mapelli, Valeria; Tripodi, Farida

    2016-01-01

    respiration. Calcium homeostasis, protein biosynthesis and the unfolded protein response are tightly intertwined and the consequences of facing calcium starvation are determined by whether cellular energy production is balanced with demands for anabolic functions. Our findings confirm that the connections...... reticulum (ER stress) triggers the unfolded protein response (UPR) and generates a state of oxidative stress that decreases cell viability. These effects are severe during growth on rapidly fermentable carbon sources and can be mitigated by decreasing the protein synthesis rate or by inducing cellular...

  20. The endoplasmic reticulum stress response in disease ...

    African Journals Online (AJOL)

    These proteins are essential for cell survival, and intuitively the ER must activate stress responses to evade immediate cell dysfunction as the cell processes lag behind. This review will discuss mainly the ER and its role in the pathogenesis and pathophysiology of epidemiologically-relevant diseases, as well as updates on ...

  1. The Tr-cp 14 cysteine protease in white clover (Trifolium repens) is localized to the endoplasmic reticulum and is associated with programmed cell death during development of tracheary elements

    DEFF Research Database (Denmark)

    Mulisch, Maria; Asp, Torben; Krupinska, Karin

    2013-01-01

    family of cysteine proteases with homology to XCP1 and XCP2 from Arabidopsis thaliana and p48h-17 from Zinnia elegans, which previously have been reported to be associated with tracheary element differentiation. The proform as well as the processed form of the protein was detected in petioles, flowers....... Immunogold studies suggest that the protease prior to the burst of the vacuole was associated to the ER cisternae. After disruption of the tonoplast, it was found in the cytoplasm, and, in later stages, associated with disintegrating material dispersed throughout the cell....

  2. Friend or foe: Endoplasmic reticulum protein 29 (ERp29) in epithelial cancer

    Science.gov (United States)

    Chen, Shaohua; Zhang, Daohai

    2015-01-01

    The endoplasmic reticulum (ER) protein 29 (ERp29) is a molecular chaperone that plays a critical role in protein secretion from the ER in eukaryotic cells. Recent studies have also shown that ERp29 plays a role in cancer. It has been demonstrated that ERp29 is inversely associated with primary tumor development and functions as a tumor suppressor by inducing cell growth arrest in breast cancer. However, ERp29 has also been reported to promote epithelial cell morphogenesis, cell survival against genotoxic stress and distant metastasis. In this review, we summarize the current understanding on the biological and pathological functions of ERp29 in cancer and discuss the pivotal aspects of ERp29 as “friend or foe” in epithelial cancer. PMID:25709888

  3. Racial Segregation and the American Foreclosure Crisis.

    Science.gov (United States)

    Rugh, Jacob S; Massey, Douglas S

    2010-10-01

    Although the rise in subprime lending and the ensuing wave of foreclosures was partly a result of market forces that have been well-identified in the literature, in the United States it was also a highly racialized process. We argue that residential segregation created a unique niche of poor minority clients who were differentially marketed risky subprime loans that were in great demand for use in mortgage-backed securities that could be sold on secondary markets. We test this argument by regressing foreclosure actions in the top 100 U.S. metropolitan areas on measures of black, Hispanic, and Asian segregation while controlling for a variety of housing market conditions, including average creditworthiness, the extent of coverage under the Community Reinvestment Act, the degree of zoning regulation, and the overall rate of subprime lending. We find that black residential dissimilarity and spatial isolation are powerful predictors of foreclosures across U.S. metropolitan areas. In order to isolate subprime lending as the causal mechanism whereby segregation influences foreclosures, we estimate a two-stage least squares model that confirms the causal effect of black segregation on the number and rate of foreclosures across metropolitan areas. In the United States segregation was an important contributing cause of the foreclosure crisis, along with overbuilding, risky lending practices, lax regulation, and the bursting of the housing price bubble.

  4. Racial Segregation and the American Foreclosure Crisis

    Science.gov (United States)

    Rugh, Jacob S.; Massey, Douglas S.

    2013-01-01

    Although the rise in subprime lending and the ensuing wave of foreclosures was partly a result of market forces that have been well-identified in the literature, in the United States it was also a highly racialized process. We argue that residential segregation created a unique niche of poor minority clients who were differentially marketed risky subprime loans that were in great demand for use in mortgage-backed securities that could be sold on secondary markets. We test this argument by regressing foreclosure actions in the top 100 U.S. metropolitan areas on measures of black, Hispanic, and Asian segregation while controlling for a variety of housing market conditions, including average creditworthiness, the extent of coverage under the Community Reinvestment Act, the degree of zoning regulation, and the overall rate of subprime lending. We find that black residential dissimilarity and spatial isolation are powerful predictors of foreclosures across U.S. metropolitan areas. In order to isolate subprime lending as the causal mechanism whereby segregation influences foreclosures, we estimate a two-stage least squares model that confirms the causal effect of black segregation on the number and rate of foreclosures across metropolitan areas. In the United States segregation was an important contributing cause of the foreclosure crisis, along with overbuilding, risky lending practices, lax regulation, and the bursting of the housing price bubble. PMID:25308973

  5. The dissociation of tumor-induced weight loss from hypoglycemia in a transplantable pluripotent rat islet tumor results in the segregation of stable alpha- and beta-cell tumor phenotypes

    DEFF Research Database (Denmark)

    Madsen, O D; Karlsen, C; Nielsen, E

    1993-01-01

    that of starved rats until death results from cachexia. After tumor resection, animals immediately resume normal feeding behavior. Comparative studies of hormone release and mRNA content in anorectic lines, MSL-G-AN and NHI-5B-AN, vs. those in the insulinoma line, MSL-G2-IN, revealed selective glucagon gene...... in animals carrying tumor necrosis factor-producing experimental tumors....... markers. By selective transplantation, it was possible to segregate stable anorectic normoglycemic tumor lines, MSL-G-AN and NHI-5B-AN, from both clones. These tumors cause an abrupt onset of anorexia when they reach a size of 400-500 mg (

  6. Towards deep learning with segregated dendrites.

    Science.gov (United States)

    Guerguiev, Jordan; Lillicrap, Timothy P; Richards, Blake A

    2017-12-05

    Deep learning has led to significant advances in artificial intelligence, in part, by adopting strategies motivated by neurophysiology. However, it is unclear whether deep learning could occur in the real brain. Here, we show that a deep learning algorithm that utilizes multi-compartment neurons might help us to understand how the neocortex optimizes cost functions. Like neocortical pyramidal neurons, neurons in our model receive sensory information and higher-order feedback in electrotonically segregated compartments. Thanks to this segregation, neurons in different layers of the network can coordinate synaptic weight updates. As a result, the network learns to categorize images better than a single layer network. Furthermore, we show that our algorithm takes advantage of multilayer architectures to identify useful higher-order representations-the hallmark of deep learning. This work demonstrates that deep learning can be achieved using segregated dendritic compartments, which may help to explain the morphology of neocortical pyramidal neurons.

  7. School Segregation and Racial Academic Achievement Gaps

    Directory of Open Access Journals (Sweden)

    Sean F. Reardon

    2016-09-01

    Full Text Available Although it is clear that racial segregation is linked to academic achievement gaps, the mechanisms underlying this link have been debated since James Coleman published his eponymous 1966 report. In this paper, I examine sixteen distinct measures of segregation to determine which is most strongly associated with academic achievement gaps. I find clear evidence that one aspect of segregation in particular—the disparity in average school poverty rates between white and black students’ schools—is consistently the single most powerful correlate of achievement gaps, a pattern that holds in both bivariate and multivariate analyses. This implies that high-poverty schools are, on average, much less effective than lower-poverty schools and suggests that strategies that reduce the differential exposure of black, Hispanic, and white students to poor schoolmates may lead to meaningful reductions in academic achievement gaps.

  8. The metabolomic signature of Leber's hereditary optic neuropathy reveals endoplasmic reticulum stress.

    Science.gov (United States)

    Chao de la Barca, Juan Manuel; Simard, Gilles; Amati-Bonneau, Patrizia; Safiedeen, Zainab; Prunier-Mirebeau, Delphine; Chupin, Stéphanie; Gadras, Cédric; Tessier, Lydie; Gueguen, Naïg; Chevrollier, Arnaud; Desquiret-Dumas, Valérie; Ferré, Marc; Bris, Céline; Kouassi Nzoughet, Judith; Bocca, Cinzia; Leruez, Stéphanie; Verny, Christophe; Miléa, Dan; Bonneau, Dominique; Lenaers, Guy; Martinez, M Carmen; Procaccio, Vincent; Reynier, Pascal

    2016-11-01

    Leber's hereditary optic neuropathy (MIM#535000), the commonest mitochondrial DNA-related disease, is caused by mutations affecting mitochondrial complex I. The clinical expression of the disorder, usually occurring in young adults, is typically characterized by subacute, usually sequential, bilateral visual loss, resulting from the degeneration of retinal ganglion cells. As the precise action of mitochondrial DNA mutations on the overall cell metabolism in Leber's hereditary optic neuropathy is unknown, we investigated the metabolomic profile of the disease. High performance liquid chromatography coupled with tandem mass spectrometry was used to quantify 188 metabolites in fibroblasts from 16 patients with Leber's hereditary optic neuropathy and eight healthy control subjects. Latent variable-based statistical methods were used to identify discriminating metabolites. One hundred and twenty-four of the metabolites were considered to be accurately quantified. A supervised orthogonal partial least squares discriminant analysis model separating patients with Leber's hereditary optic neuropathy from control subjects showed good predictive capability (Q 2cumulated = 0.57). Thirty-eight metabolites appeared to be the most significant variables, defining a Leber's hereditary optic neuropathy metabolic signature that revealed decreased concentrations of all proteinogenic amino acids, spermidine, putrescine, isovaleryl-carnitine, propionyl-carnitine and five sphingomyelin species, together with increased concentrations of 10 phosphatidylcholine species. This signature was not reproduced by the inhibition of complex I with rotenone or piericidin A in control fibroblasts. The importance of sphingomyelins and phosphatidylcholines in the Leber's hereditary optic neuropathy signature, together with the decreased amino acid pool, suggested an involvement of the endoplasmic reticulum. This was confirmed by the significantly increased phosphorylation of PERK and eIF2α, as well as

  9. Chemical segregation and self polarisation in ferroelectrics

    Directory of Open Access Journals (Sweden)

    Bernard E. Watts

    2009-06-01

    Full Text Available Chemical partitioning or segregation is commonly encountered in solid-state syntheses. It is driven by compositional, thermal and electric field gradients. These phenomena can be quite extreme in thin films and lead to notable effects on the electrical properties of ferroelectrics. The segregation in ferroelectric thin films will be illustrated and the mechanisms explained in terms of diffusion processes driven by a potential gradient of the oxygen. The hypothesis can also explain self polarisation and imprint in ferroelectric hysteresis.

  10. Particle segregation in pneumatic conveying lines

    Energy Technology Data Exchange (ETDEWEB)

    McGlinchey, D.; Marjanovic, P.; Cook, S.; Jones, M.G. [Glasgow Caledonian University, Glasgow (United Kingdom). Centre for Industrial Bulk Solids Handling

    2000-07-01

    This investigation studied segregation of particles during pneumatic transport from a theoretical and experimental perspective. Dilute phase or suspension flow and dense phase (non-suspension flow) were both considered. A computer model was generated based on the conservation equations to investigate dilute phase conditions; an initial qualitative investigation of material behaviour being conveyed in dense phase was made with plastic pellets and salt as a segregating mixture in a small test rig and the results from a full scale test rig conveying two grades of coal of different size distributions are discussed. 11 refs., 9 figs., 1 tab.

  11. Plasma membrane—endoplasmic reticulum contact sites regulate phosphatidylcholine synthesis

    NARCIS (Netherlands)

    Tavassoli, S.; Chao, J.T.; Young, B.P.; Cox, R.C.; Prinz, W.A.; de Kroon, A.I.P.M.; Loewen, C.I.R.

    2013-01-01

    Synthesis of phospholipids, sterols and sphingolipids is thought to occur at contact sites between the endoplasmic reticulum (ER) and other organelles because many lipid-synthesizing enzymes are enriched in these contacts. In only a few cases have the enzymes been localized to contacts in vivo and

  12. Tannic Acid Induces Endoplasmic Reticulum Stress-Mediated Apoptosis in Prostate Cancer.

    Science.gov (United States)

    Nagesh, Prashanth K B; Hatami, Elham; Chowdhury, Pallabita; Kashyap, Vivek K; Khan, Sheema; Hafeez, Bilal B; Chauhan, Subhash C; Jaggi, Meena; Yallapu, Murali M

    2018-03-07

    Endoplasmic reticulum (ER) stress is an intriguing target with significant clinical importance in chemotherapy. Interference with ER functions can lead to the accumulation of unfolded proteins, as detected by transmembrane sensors that instigate the unfolded protein response (UPR). Therefore, controlling induced UPR via ER stress with natural compounds could be a novel therapeutic strategy for the management of prostate cancer. Tannic acid (a naturally occurring polyphenol) was used to examine the ER stress mediated UPR pathway in prostate cancer cells. Tannic acid treatment inhibited the growth, clonogenic, invasive, and migratory potential of prostate cancer cells. Tannic acid demonstrated activation of ER stress response (Protein kinase R-like endoplasmic reticulum kinase (PERK) and inositol requiring enzyme 1 (IRE1)) and altered its regulatory proteins (ATF4, Bip, and PDI) expression. Tannic acid treatment affirmed upregulation of apoptosis-associated markers (Bak, Bim, cleaved caspase 3, and cleaved PARP), while downregulation of pro-survival proteins (Bcl-2 and Bcl-xL). Tannic acid exhibited elevated G₁ population, due to increase in p18 INK4C and p21 WAF1/CIP1 expression, while cyclin D1 expression was inhibited. Reduction of MMP2 and MMP9, and reinstated E-cadherin signifies the anti-metastatic potential of this compound. Altogether, these results demonstrate that tannic acid can promote apoptosis via the ER stress mediated UPR pathway, indicating a potential candidate for cancer treatment.

  13. Impact of high cholesterol and endoplasmic reticulum stress on metabolic diseases: An updated mini-review

    Directory of Open Access Journals (Sweden)

    Erdi Sozen

    2017-08-01

    Full Text Available Endoplasmic reticulum (ER is the major site of protein folding and calcium storage. Beside the role of ER in protein homeostasis, it controls the cholesterol production and lipid-membrane biosynthesis as well as surviving and cell death signaling mechanisms in the cell. It is well-documented that elevated plasma cholesterol induces adverse effects in cardiovascular diseases (CVDs, liver disorders, such as non-alcoholic fatty liver disease (NAFLD, non-alcoholic steatosis hepatitis (NASH, and metabolic diseases which are associated with oxidative and ER stress. Recent animal model and human studies have showed high cholesterol and ER stress as an emerging factors involved in the development of many metabolic diseases. In this review, we will summarize the crucial effects of hypercholesterolemia and ER stress response in the pathogenesis of CVDs, NAFLD/NASH, diabetes and obesity which are major health problems in western countries. Keywords: Endoplasmic reticulum stress, High cholesterol, Cardiovascular diseases, Non-alcoholic fatty liver disease, Non-alcoholic steatosis hepatitis

  14. Endoplasmic Reticulum Stress-Mediated Hippocampal Neuron Apoptosis Involved in Diabetic Cognitive Impairment

    Directory of Open Access Journals (Sweden)

    Xiaoming Zhang

    2013-01-01

    Full Text Available Poor management of DM causes cognitive impairment while the mechanism is still unconfirmed. The aim of the present study was to investigate the activation of C/EBP Homology Protein (CHOP, the prominent mediator of the endoplasmic reticulum (ER stress-induced apoptosis under hyperglycemia. We employed streptozotocin- (STZ- induced diabetic rats to explore the ability of learning and memory by the Morris water maze test. The ultrastructure of hippocampus in diabetic rats and cultured neurons in high glucose medium were observed by transmission electron microscopy and scanning electron microscopy. TUNEL staining was also performed to assess apoptotic cells while the expression of CHOP was assayed by immunohistochemistry and Western blot assay in these hippocampal neurons. Six weeks after diabetes induction, the escape latency increased and the average frequency in finding the platform decreased in diabetic rats (P<0.05. The morphology of neuron and synaptic structure was impaired; the number of TUNEL-positive cells and the expression of CHOP in hippocampus of diabetic rats and high glucose medium cultured neurons were markedly altered (P<0.05. The present results suggested that the CHOP-dependent endoplasmic reticulum (ER stress-mediated apoptosis may be involved in hyperglycemia-induced hippocampal synapses and neurons impairment and promote the diabetic cognitive impairment.

  15. The plant membrane surrounding powdery mildew haustoria shares properties with the endoplasmic reticulum membrane

    DEFF Research Database (Denmark)

    Kwaaitaal, Mark Adrianus Cornelis J; Nielsen, Mads Eggert; Böhlenius, Henrik

    2017-01-01

    Many filamentous plant pathogens place specialized feeding structures, called haustoria, inside living host cells. As haustoria grow, they are believed to manipulate plant cells to generate a specialized, still enigmatic extrahaustorial membrane (EHM) around them. Here, we focused on revealing...... properties of the EHM. With the help of membranespecific dyes and transient expression of membrane-associated proteins fused to fluorescent tags, we studied the nature of the EHM generated by barley leaf epidermal cells around powdery mildew haustoria. Observations suggesting that endoplasmic reticulum (ER...... that it is not a continuum of the ER. Furthermore, GDP-locked Sar1 and a nucleotide-free RabD2a, which block ER to Golgi exit, did not hamper haustorium formation. These results indicated that the EHM shares features with the plant ER membrane, but that the EHM membrane is not dependent on conventional secretion...

  16. Endoplasmic reticulum-dependent redox reactions control endoplasmic reticulum-associated degradation and pathogen entry.

    Science.gov (United States)

    Walczak, Christopher P; Bernardi, Kaleena M; Tsai, Billy

    2012-04-15

    Protein misfolding within the endoplasmic reticulum (ER) is managed by an ER quality control system that retro-translocates aberrant proteins into the cytosol for proteasomal destruction. This process, known as ER-associated degradation, utilizes the action of ER redox enzymes to accommodate the disulfide-bonded nature of misfolded proteins. Strikingly, various pathogenic viruses and toxins co-opt these redox components to reach the cytosol during entry. These redox factors thus regulate critical cellular homeostasis and host-pathogen interactions. Recent studies identify specific members of the protein disulfide isomerase (PDI) family, which use their chaperone and catalytic activities, in engaging both misfolded ER proteins and pathogens. The precise molecular mechanism by which a dedicated PDI family member disrupts the disulfide bonds in the misfolded ER proteins and pathogens, as well as how they act to unfold these substrates to promote their ER-to-cytosol membrane transport, remain poorly characterized. How PDI family members distinguish folded versus misfolded ER substrates remains enigmatic. What physical characteristics surrounding a substrate's disulfide bond instruct PDI that it is mispaired or native? For the pathogens, as their disulfide bonds normally serve a critical role in providing physical support, what conformational changes experienced in the host enable their disulfide bonds to be disrupted? A combination of more rigorous biochemical and high-resolution structural studies should begin to address these questions.

  17. Engineering economic evaluations of trash segregation alternatives

    International Nuclear Information System (INIS)

    Collins, H.E.

    1987-01-01

    Health physicists are becoming increasingly involved in the selection of equipment to segregate a contaminated trash from clean trash in the effort to reduce low level waste disposal costs. Although well qualified to evaluate the technical merits of different equipment, health physicists also need to be aware of the elements of economic comparisons of different alternatives that meet all technical requirements

  18. Koedukation oder Geschlechtertrennung = Coeducation or Gender Segregation.

    Science.gov (United States)

    Baumert, Jurgen

    1992-01-01

    Presents study results examining recruitment practice differences between coeducational and gender segregated secondary schools in Germany. Discusses the impact of organizational form on teacher judgments, achievement in specific subjects, school subject-related interests, and school commitment. Reports that under conditions of free school choice,…

  19. Segregation reinforced by urban planning | IDRC - International ...

    International Development Research Centre (IDRC) Digital Library (Canada)

    2015-10-28

    Oct 28, 2015 ... ... What is driving urban violence? Segregated urban planning can leave a legacy of community tension and insecurity. Potential solutions? Include vulnerable communities in city planning decisions; invest in transport infrastructure; and regularly update city development plans to reflect population growth.

  20. 49 CFR 176.83 - Segregation.

    Science.gov (United States)

    2010-10-01

    ... presence of one or more steel bulkheads or decks between them or a combination thereof. Intervening spaces... substance but vary only in their water content (for example, sodium sulfide in Division 4.2 or Class 8) or... applied. (11) Certain exceptions from segregation for waste cyanides or waste cyanide mixtures or...

  1. Plasmid and chromosome segregation in prokaryotes

    DEFF Research Database (Denmark)

    Møller-Jensen, Jakob; Bugge Jensen, Rasmus; Gerdes, Kenn

    2000-01-01

    Recent major advances in the understanding of prokaryotic DNA segregation have been achieved by using fluorescence microscopy to visualize the localization of cellular components. Plasmids and bacterial chromosomes are partitioned in a highly dynamic fashion, suggesting the presence of a mitotic...

  2. Educational Justice, Segregated Schooling and Vocational Education

    Science.gov (United States)

    Giesinger, Johannes

    2017-01-01

    The philosophical debate on educational justice currently focusses on the Anglo-American situation. This essay brings in an additional perspective. It provides a justice-oriented critique of the segregated education systems in German-speaking countries. First, arguments that are commonly put forward in favour of these systems are rejected. Second,…

  3. A Social Network Analysis of Occupational Segregation

    DEFF Research Database (Denmark)

    Buhai, Ioan Sebastian; van der Leij, Marco

    We develop a social network model of occupational segregation between different social groups, generated by the existence of positive inbreeding bias among individuals from the same group. If network referrals are important for job search, then expected homophily in the contact network structure...

  4. Sex segregation in undergraduate engineering majors

    Science.gov (United States)

    Litzler, Elizabeth

    Gender inequality in engineering persists in spite of women reaching parity in college enrollments and degrees granted. To date, no analyses of educational sex segregation have comprehensively examined segregation within one discipline. To move beyond traditional methods of studying the long-standing stratification by field of study in higher education, I explore gender stratification within one field: engineering. This dissertation investigates why some engineering disciplines have a greater representation of women than other engineering disciplines. I assess the individual and institutional factors and conditions associated with women's representation in certain engineering departments and compare the mechanisms affecting women's and men's choice of majors. I use national data from the Engineering Workforce Commission, survey data from 21 schools in the Project to Assess Climate in Engineering study, and Carnegie Foundation classification information to study sex segregation in engineering majors from multiple perspectives: the individual, major, institution, and country. I utilize correlations, t-tests, cross-tabulations, log-linear modeling, multilevel logistic regression and weighted least squares regression to test the relative utility of alternative explanations for women's disproportionate representation across engineering majors. As a whole, the analyses illustrate the importance of context and environment for women's representation in engineering majors. Hypotheses regarding hostile climate and discrimination find wide support across different analyses, suggesting that women's under-representation in certain engineering majors is not a question of choice or ability. However, individual level factors such as having engineering coursework prior to college show an especially strong association with student choice of major. Overall, the analyses indicate that institutions matter, albeit less for women, and women's under-representation in engineering is not

  5. circHIPK2-mediated σ-1R promotes endoplasmic reticulum stress in human pulmonary fibroblasts exposed to silica.

    Science.gov (United States)

    Cao, Zhouli; Xiao, Qingling; Dai, Xiaoniu; Zhou, Zewei; Jiang, Rong; Cheng, Yusi; Yang, Xiyue; Guo, Huifang; Wang, Jing; Xi, Zhaoqing; Yao, Honghong; Chao, Jie

    2017-12-13

    Silicosis is characterized by fibroblast accumulation and excessive deposition of extracellular matrix. Although the roles of SiO 2 -induced chemokines and cytokines released from alveolar macrophages have received significant attention, the direct effects of SiO 2 on protein production and functional changes in pulmonary fibroblasts have been less extensively studied. Sigma-1 receptor, which has been associated with cell proliferation and migration in the central nervous system, is expressed in the lung, but its role in silicosis remains unknown. To elucidate the role of sigma-1 receptor in fibrosis induced by silica, both the upstream molecular mechanisms and the functional effects on cell proliferation and migration were investigated. Both molecular biological assays and pharmacological techniques, combined with functional experiments, such as migration and proliferation, were applied in human pulmonary fibroblasts from adults to analyze the molecular and functional changes induced by SiO 2 . SiO 2 induced endoplasmic reticulum stress in association with enhanced expression of sigma-1 receptor. Endoplasmic reticulum stress promoted migration and proliferation of human pulmonary fibroblasts-adult exposed to SiO 2 , inducing the development of silicosis. Inhibition of sigma-1 receptor ameliorated endoplasmic reticulum stress and fibroblast functional changes induced by SiO 2 . circHIPK2 is involved in the regulation of sigma-1 receptor in human pulmonary fibroblasts-adult exposed to SiO 2 . Our study elucidated a link between SiO 2 -induced fibrosis and sigma-1 receptor signaling, thereby providing novel insight into the potential use of sigma-1 receptor/endoplasmic reticulum stress in the development of novel therapeutic strategies for silicosis treatment.

  6. Ab-initio study of surface segregation in aluminum alloys

    Energy Technology Data Exchange (ETDEWEB)

    Qin, Yifa, E-mail: yfqin10s@imr.ac.cn; Wang, Shaoqing

    2017-03-31

    Highlights: • A thorough study of surface segregation energies of 41 elements in Al is performed. • Segregation energies vary periodically with the atomic numbers of impurities. • 41 elements are classified into 3 groups according to the signs of segregation energies. • The results are validated by the surface/total concentration ratio in Al alloys. - Abstract: We have calculated surface segregation energies of 41 impurities by means of density functional theory calculations. An interesting periodical variation tendency was found for surface segregation energies derived. For the majority of main group elements, segregation energies are negative which means solute elements enrichment at Al surface is energetically more favorable than uniformly dissolution. Half of transition elements possess positive segregation energies and the energies are sensitive to surface crystallographic orientations. A strong correlation is found between the segregation energies at the Al surface and the surface energ of solute elements.

  7. Charles J. McMahon Interfacial Segregation and Embrittlement Symposium

    National Research Council Canada - National Science Library

    Vitek, Vaclav

    2003-01-01

    .... McMahon Interfacial Segregation and Embrittlement Symposium: Grain Boundary Segregation and Fracture in Steels was sponsored by ASM International, Materials Science Critical Technology Sector, Structural Materials Division, Materials Processing...

  8. Burst-time-dependent plasticity robustly guides ON/OFF segregation in the lateral geniculate nucleus.

    Directory of Open Access Journals (Sweden)

    Julijana Gjorgjieva

    2009-12-01

    Full Text Available Spontaneous retinal activity (known as "waves" remodels synaptic connectivity to the lateral geniculate nucleus (LGN during development. Analysis of retinal waves recorded with multielectrode arrays in mouse suggested that a cue for the segregation of functionally distinct (ON and OFF retinal ganglion cells (RGCs in the LGN may be a desynchronization in their firing, where ON cells precede OFF cells by one second. Using the recorded retinal waves as input, with two different modeling approaches we explore timing-based plasticity rules for the evolution of synaptic weights to identify key features underlying ON/OFF segregation. First, we analytically derive a linear model for the evolution of ON and OFF weights, to understand how synaptic plasticity rules extract input firing properties to guide segregation. Second, we simulate postsynaptic activity with a nonlinear integrate-and-fire model to compare findings with the linear model. We find that spike-time-dependent plasticity, which modifies synaptic weights based on millisecond-long timing and order of pre- and postsynaptic spikes, fails to segregate ON and OFF retinal inputs in the absence of normalization. Implementing homeostatic mechanisms results in segregation, but only with carefully-tuned parameters. Furthermore, extending spike integration timescales to match the second-long input correlation timescales always leads to ON segregation because ON cells fire before OFF cells. We show that burst-time-dependent plasticity can robustly guide ON/OFF segregation in the LGN without normalization, by integrating pre- and postsynaptic bursts irrespective of their firing order and over second-long timescales. We predict that an LGN neuron will become ON- or OFF-responsive based on a local competition of the firing patterns of neighboring RGCs connecting to it. Finally, we demonstrate consistency with ON/OFF segregation in ferret, despite differences in the firing properties of retinal waves. Our

  9. Type 2 diabetes mellitus induces congenital heart defects in murine embryos by increasing oxidative stress, endoplasmic reticulum stress, and apoptosis.

    Science.gov (United States)

    Wu, Yanqing; Reece, E Albert; Zhong, Jianxiang; Dong, Daoyin; Shen, Wei-Bin; Harman, Christopher R; Yang, Peixin

    2016-09-01

    expression; and XBP1 messenger RNA splicing, as well as increased cleaved caspase 3 and 8 in embryonic hearts. Furthermore, maternal type 2 diabetes mellitus triggered excessive apoptosis in ventricular myocardium, endocardial cushion, and outflow tract of the embryonic heart. Similar to those observations in type 1 diabetic embryopathy, maternal type 2 diabetes mellitus causes heart defects in the developing embryo manifested with oxidative stress, endoplasmic reticulum stress, and excessive apoptosis in heart cells. Copyright © 2016 Elsevier Inc. All rights reserved.

  10. 3-Bromopyruvate inhibits calcium uptake by sarcoplasmic reticulum vesicles but not SERCA ATP hydrolysis activity.

    Science.gov (United States)

    Jardim-Messeder, Douglas; Camacho-Pereira, Juliana; Galina, Antonio

    2012-05-01

    3-Bromopyruvate (3BrPA) is an antitumor agent that alkylates the thiol groups of enzymes and has been proposed as a treatment for neoplasias because of its specific reactivity with metabolic energy transducing enzymes in tumor cells. In this study, we show that the sarco/endoplasmic reticulum calcium (Ca(2+)) ATPase (SERCA) type 1 is one of the target enzymes of 3BrPA activity. Sarco/endoplasmic reticulum vesicles (SRV) were incubated in the presence of 1mM 3BrPA, which was unable to inhibit the ATPase activity of SERCA. However, Ca(2+)-uptake activity was significantly inhibited by 80% with 150 μM 3BrPA. These results indicate that 3BrPA has the ability to uncouple the ATP hydrolysis from the calcium transport activities. In addition, we observed that the inclusion of 2mM reduced glutathione (GSH) in the reaction medium with different 3BrPA concentrations promoted an increase in 40% in ATPase activity and protects the inhibition promoted by 3BrPA in calcium uptake activity. This derivatization is accompanied by a decrease of reduced cysteine (Cys), suggesting that GSH and 3BrPA increases SERCA activity and transport by pyruvylation and/or S-glutathiolation mediated by GSH at a critical Cys residues of the SERCA. Copyright © 2012 Elsevier Ltd. All rights reserved.

  11. Mitochondrial enzymes and endoplasmic reticulum calcium stores as targets of oxidative stress in neurodegenerative diseases.

    Science.gov (United States)

    Gibson, Gary E; Huang, Hsueh-Meei

    2004-08-01

    Considerable evidence indicates that oxidative stress accompanies age-related neurodegenerative diseases. Specific mechanisms by which oxidative stress leads to neurodegeneration are unknown. Two targets of oxidative stress that are known to change in neurodegenerative diseases are the mitochondrial enzyme alpha-ketoglutarate dehydrogenase complex (KGDHC) and endoplasmic reticulum calcium stores. KGDHC activities are diminished in all common neurodegenerative diseases and the changes are particularly well documented in Alzheimer's disease (AD). A second change that occurs in cells from AD patients is an exaggerated endoplasmic reticulum calcium store [i.e., bombesin-releasable calcium stores (BRCS)]. H(2)O(2), a general oxidant, changes both variables in the same direction as occurs in disease. Other oxidants selectively alter these variables. Various antioxidants were used to help define the critical oxidant species that modifies these responses. All of the antioxidants diminish the oxidant-induced carboxy-dichlorofluorescein (cDCF) detectable reactive oxygen species (ROS), but have diverse actions on these cellular processes. For example, alpha-keto-beta-methyl-n-valeric acid (KMV) diminishes the H(2)O(2) effects on BRCS, while trolox and DMSO exaggerate the response. Acute trolox treatment does not alter H(2)O(2)-induced changes in KGDHC, whereas chronic treatment with trolox increases KGDHC almost threefold. The results suggest that KGDHC and BRCS provide targets by which oxidative stress may induce neurodegeneration and a useful tool for selecting antioxidants for reversing age-related neurodegeneration.

  12. Roles of Endoplasmic Reticulum Stress in NECA-Induced Cardioprotection against Ischemia/Reperfusion Injury

    Directory of Open Access Journals (Sweden)

    Fengmei Xing

    2017-01-01

    Full Text Available Objective. This study aimed to investigate whether the nonselective A2 adenosine receptor agonist NECA induces cardioprotection against myocardial ischemia/reperfusion (I/R injury via glycogen synthase kinase 3β (GSK-3β and the mitochondrial permeability transition pore (mPTP through inhibition of endoplasmic reticulum stress (ERS. Methods and Results. H9c2 cells were exposed to H2O2 for 20 minutes. NECA significantly prevented H2O2-induced TMRE fluorescence reduction, indicating that NECA inhibited the mPTP opening. NECA blocked H2O2-induced GSK-3β phosphorylation and GRP94 expression. NECA increased GSK-3β phosphorylation and decreased GRP94 expression, which were prevented by both ERS inductor 2-DG and PKG inhibitor KT5823, suggesting that NECA may induce cardioprotection through GSK-3β and cGMP/PKG via ERS. In isolated rat hearts, both NECA and the ERS inhibitor TUDCA decreased myocardial infarction, increased GSK-3β phosphorylation, and reversed GRP94 expression at reperfusion, suggesting that NECA protected the heart by inhibiting GSK-3β and ERS. Transmission electron microscopy showed that NECA and TUDCA reduced mitochondrial swelling and endoplasmic reticulum expansion, further supporting that NECA protected the heart by preventing the mPTP opening and ERS. Conclusion. These data suggest that NECA prevents the mPTP opening through inactivation of GSK-3β via ERS inhibition. The cGMP/PKG signaling pathway is responsible for GSK-3β inactivation by NECA.

  13. p53-inducible DHRS3 Is an Endoplasmic Reticulum Protein Associated with Lipid Droplet Accumulation*

    Science.gov (United States)

    Deisenroth, Chad; Itahana, Yoko; Tollini, Laura; Jin, Aiwen; Zhang, Yanping

    2011-01-01

    The transcription factor p53 plays a critical role in maintaining homeostasis as it relates to cellular growth, proliferation, and metabolism. In an effort to identify novel p53 target genes, a microarray approach was utilized to identify DHRS3 (also known as retSDR1) as a robust candidate gene. DHRS3 is a highly conserved member of the short chain alcohol dehydrogenase/reductase superfamily with a reported role in lipid and retinoid metabolism. Here, we demonstrate that DHRS3 is an endoplasmic reticulum (ER) protein that is shuttled to the ER via an N-terminal endoplasmic reticulum targeting signal. One important function of the ER is synthesis of neutral lipids that are packaged into lipid droplets whose biogenesis occurs from ER-derived membranes. DHRS3 is enriched at focal points of lipid droplet budding where it also localizes to the phospholipid monolayer of ER-derived lipid droplets. p53 promotes lipid droplet accumulation in a manner consistent with DHRS3 enrichment in the ER. As a p53 target gene, the observations of Dhrs3 location and potential function provide novel insight into an unexpected role for p53 in lipid droplet dynamics with implications in cancer cell metabolism and obesity. PMID:21659514

  14. Methods for monitoring endoplasmic reticulum stress and the unfolded protein response.

    LENUS (Irish Health Repository)

    Samali, Afshin

    2010-01-01

    The endoplasmic reticulum (ER) is the site of folding of membrane and secreted proteins in the cell. Physiological or pathological processes that disturb protein folding in the endoplasmic reticulum cause ER stress and activate a set of signaling pathways termed the Unfolded Protein Response (UPR). The UPR can promote cellular repair and sustained survival by reducing the load of unfolded proteins through upregulation of chaperones and global attenuation of protein synthesis. Research into ER stress and the UPR continues to grow at a rapid rate as many new investigators are entering the field. There are also many researchers not working directly on ER stress, but who wish to determine whether this response is activated in the system they are studying: thus, it is important to list a standard set of criteria for monitoring UPR in different model systems. Here, we discuss approaches that can be used by researchers to plan and interpret experiments aimed at evaluating whether the UPR and related processes are activated. We would like to emphasize that no individual assay is guaranteed to be the most appropriate one in every situation and strongly recommend the use of multiple assays to verify UPR activation.

  15. Methods for Monitoring Endoplasmic Reticulum Stress and the Unfolded Protein Response

    Directory of Open Access Journals (Sweden)

    Afshin Samali

    2010-01-01

    Full Text Available The endoplasmic reticulum (ER is the site of folding of membrane and secreted proteins in the cell. Physiological or pathological processes that disturb protein folding in the endoplasmic reticulum cause ER stress and activate a set of signaling pathways termed the Unfolded Protein Response (UPR. The UPR can promote cellular repair and sustained survival by reducing the load of unfolded proteins through upregulation of chaperones and global attenuation of protein synthesis. Research into ER stress and the UPR continues to grow at a rapid rate as many new investigators are entering the field. There are also many researchers not working directly on ER stress, but who wish to determine whether this response is activated in the system they are studying: thus, it is important to list a standard set of criteria for monitoring UPR in different model systems. Here, we discuss approaches that can be used by researchers to plan and interpret experiments aimed at evaluating whether the UPR and related processes are activated. We would like to emphasize that no individual assay is guaranteed to be the most appropriate one in every situation and strongly recommend the use of multiple assays to verify UPR activation.

  16. Hypothyroidism Causes Endoplasmic Reticulum Stress in Adult Rat Hippocampus: A Mechanism Associated with Hippocampal Damage

    Directory of Open Access Journals (Sweden)

    Alejandra Paola Torres-Manzo

    2018-01-01

    Full Text Available Thyroid hormones (TH are essential for hippocampal neuronal viability in adulthood, and their deficiency causes hypothyroidism, which is related to oxidative stress events and neuronal damage. Also, it has been hypothesized that hypothyroidism causes a glucose deprivation in the neuron. This study is aimed at evaluating the temporal participation of the endoplasmic reticulum stress (ERE in hippocampal neurons of adult hypothyroid rats and its association with the oxidative stress events. Adult Wistar male rats were divided into euthyroid and hypothyroid groups. Thyroidectomy with parathyroid gland reimplementation caused hypothyroidism at three weeks postsurgery. Oxidative stress, redox environment, and antioxidant enzyme markers, as well as the expression of the ERE through the pathways of PERK, ATF6, and IRE1, were evaluated at the 3rd and 4th weeks postsurgery. We found a rise in ROS and nitrite production; also, catalase increased and glutathione peroxidase diminished their activities. These events promote an enhancement of the lipoperoxidation, as well as of γ-GT, myeloperoxidase, and caspase 3 activities. With respect to ERE, there were ATF6, IRE1, and GADD153 overexpressions with a reduction in mitochondrial activity and GSH2/GSSG ratio. We conclude that the endoplasmic reticulum stress might play a pivotal role in the activation of hypothyroidism-induced hippocampal cell death.

  17. 41 CFR 60-1.8 - Segregated facilities.

    Science.gov (United States)

    2010-07-01

    ... 41 Public Contracts and Property Management 1 2010-07-01 2010-07-01 true Segregated facilities. 60...; Compliance Reports § 60-1.8 Segregated facilities. To comply with its obligations under the Order, a contractor must ensure that facilities provided for employees are provided in such a manner that segregation...

  18. Gender Segregation in Nursery School: Predictors and Outcomes.

    Science.gov (United States)

    Maccoby, Eleanor E.; Jacklin, Carol Nagy

    Sex segregation is a powerful phenomenon in childhood. It occurs universally whenever children have a choice of playmates and is found in sub-human primates too. Adults are not directly responsible for sex segregation. Data do not support the hypothesis that the most ladylike girls and the most rough and active boys first form the segregated play…

  19. "E Pluribus"... Separation: Deepening Double Segregation for More Students

    Science.gov (United States)

    Orfield, Gary; Kucsera, John; Siegel-Hawley, Genevieve

    2012-01-01

    This report shows segregation has increased dramatically across the country for Latino students, who are attending more intensely segregated and impoverished schools than they have for generations. The segregation increases have been the most dramatic in the West. The typical Latino student in the region attends a school where less than a quarter…

  20. Segregation effects and phase developments during solidification of alloy 625

    DEFF Research Database (Denmark)

    Højerslev, Christian; Tiedje, Niels Skat; Hald, John

    2006-01-01

    contained gamma-phase, Laves phase and, if carbon was dissolved in the liquid, niobium rich carbides formed. Molybdenum and niobium showed strong tendencies to segregate. Their segregation was balanced by inverse segregation of nickel and iron. The chromium concentration remained almost constant in gamma...

  1. Requirements for the evaluation of computational speech segregation systems

    DEFF Research Database (Denmark)

    May, Tobias; Dau, Torsten

    2014-01-01

    Recent studies on computational speech segregation reported improved speech intelligibility in noise when estimating and applying an ideal binary mask with supervised learning algorithms. However, an important requirement for such systems in technical applications is their robustness to acoustic...... associated with perceptual attributes in speech segregation. The results could help establish a framework for a systematic evaluation of future segregation systems....

  2. Residential segregation of socioeconomic variables and health indices in Iran

    Directory of Open Access Journals (Sweden)

    Seyed Saeed Hashemi Nazari

    2013-01-01

    Conclusions: Correlation of segregation of determinants of socioeconomic status with segregation of health indices is an indicator of existence of hot zones of health problems across some provinces. Further studies using multilevel modeling and individual data in health outcomes at individual level and segregation measures at appropriate geographic levels are required to confirm these relations.

  3. Asymmetric Damage Segregation Constitutes an Emergent Population-Level Stress Response

    DEFF Research Database (Denmark)

    Vedel, Søren; Nunns, Harry; Košmrlj, Andrej

    2016-01-01

    Asymmetric damage segregation (ADS) is a mechanism for increasing population fitness through non-random, asymmetric partitioning of damaged macromolecules at cell division. ADS has been reported across multiple organisms, though the measured effects on fitness of individuals are often small. Here...

  4. Multivesicular body formation enhancement and exosome release during endoplasmic reticulum stress.

    Science.gov (United States)

    Kanemoto, Soshi; Nitani, Ryota; Murakami, Tatsuhiko; Kaneko, Masayuki; Asada, Rie; Matsuhisa, Koji; Saito, Atsushi; Imaizumi, Kazunori

    2016-11-11

    The endoplasmic reticulum (ER) plays a pivotal role in maintaining cellular homeostasis. However, numerous environmental and genetic factors give rise to ER stress by inducing an accumulation of unfolded proteins. Under ER stress conditions, cells initiate the unfolded protein response (UPR). Here, we demonstrate a novel aspect of the UPR by electron microscopy and immunostaining analyses, whereby multivesicular body (MVB) formation was enhanced after ER stress. This MVB formation was influenced by inhibition of ER stress transducers inositol required enzyme 1 (IRE1) and PKR-like ER kinase (PERK). Furthermore, exosome release was also increased during ER stress. However, in IRE1 or PERK deficient cells, exosome release was not upregulated, indicating that IRE1- and PERK-mediated pathways are involved in ER stress-dependent exosome release. Copyright © 2016 Elsevier Inc. All rights reserved.

  5. Vanilloid receptor expressed in the sarcoplasmic reticulum of rat skeletal muscle

    International Nuclear Information System (INIS)

    Xin Hong; Tanaka, Hideyuki; Yamaguchi, Maki; Takemori, Shigeru; Nakamura, Akio; Kohama, Kazuhiro

    2005-01-01

    Vanilloid receptor subtype 1 (VR1) was cloned as a capsaicin receptor from neuronal cells of dorsal root ganglia. VR1 was subsequently found in a few non-neuronal tissues, including skeletal muscle [Onozawa et al., Tissue distribution of capsaicin receptor in the various organs of rats, Proc. Jpn. Acad. Ser. B 76 (2000) 68-72]. We confirmed the expression of VR1 in muscle cells using the RT-PCR method and Western blot analysis. Immunostaining studies with a confocal microscope and an electron microscope indicated that VR1 was present in the sarcoplasmic reticulum (SR), a store of Ca 2+ . The SR releases Ca 2+ to cause a contraction when a muscle is excited. However, SR still releases a small amount of Ca 2+ under relaxed conditions. We found that this leakage was enhanced by capsaicin and was antagonized by capsazepine, a capsaicin blocker, indicating that leakage of Ca 2+ occurs through a channel composed of VR1

  6. Cyclosporine A-sensitive, cyclophilin B-dependent endoplasmic reticulum-associated degradation.

    Directory of Open Access Journals (Sweden)

    Riccardo Bernasconi

    2010-09-01

    Full Text Available Peptidyl-prolyl cis/trans isomerases (PPIs catalyze cis/trans isomerization of peptide bonds preceding proline residues. The involvement of PPI family members in protein refolding has been established in test tube experiments. Surprisingly, however, no data is available on the involvement of endoplasmic reticulum (ER-resident members of the PPI family in protein folding, quality control or disposal in the living cell. Here we report that the immunosuppressive drug cyclosporine A (CsA selectively inhibits the degradation of a subset of misfolded proteins generated in the ER. We identify cyclophilin B (CyPB as the ER-resident target of CsA that catalytically enhances disposal from the ER of ERAD-L(S substrates containing cis proline residues. Our manuscript presents the first evidence for enzymatic involvement of a PPI in protein quality control in the ER of living cells.

  7. Cyclosporine A-Sensitive, Cyclophilin B-Dependent Endoplasmic Reticulum-Associated Degradation

    Science.gov (United States)

    Luban, Jeremy; Molinari, Maurizio

    2010-01-01

    Peptidyl-prolyl cis/trans isomerases (PPIs) catalyze cis/trans isomerization of peptide bonds preceding proline residues. The involvement of PPI family members in protein refolding has been established in test tube experiments. Surprisingly, however, no data is available on the involvement of endoplasmic reticulum (ER)-resident members of the PPI family in protein folding, quality control or disposal in the living cell. Here we report that the immunosuppressive drug cyclosporine A (CsA) selectively inhibits the degradation of a subset of misfolded proteins generated in the ER. We identify cyclophilin B (CyPB) as the ER-resident target of CsA that catalytically enhances disposal from the ER of ERAD-LS substrates containing cis proline residues. Our manuscript presents the first evidence for enzymatic involvement of a PPI in protein quality control in the ER of living cells. PMID:20927389

  8. Management of the endoplasmic reticulum stress by activation of the heat shock response in yeast

    DEFF Research Database (Denmark)

    Hou, Jin; Tang, Hongting; Liu, Zihe

    2014-01-01

    In yeast Saccharomyces cerevisiae, accumulation of misfolded proteins in the endoplasmic reticulum (ER) causes ER stress and activates the unfolded protein response (UPR), which is mediated by Hac1p. The heat shock response (HSR) mediated by Hsf1p, mainly regulates cytosolic processes and protects...... the cell from stresses. Here, we find that a constitutive activation of the HSR could increase ER stress resistance in both wild-type and UPR-deficient cells. Activation of HSR decreased UPR activation in the WT (as shown by the decreased HAC1 mRNA splicing). We analyzed the genome-wide transcriptional...... response in order to propose regulatory mechanisms that govern the interplay between UPR and HSR and followed up for the hypotheses by experiments in vivo and in vitro. Interestingly, we found that the regulation of ER stress response via HSR is (1) only partially dependent on over-expression of Kar2p (ER...

  9. The Role of the CRL4Cdt2 Target Spd1 in Chromosome Segregation in Fission Yeast

    DEFF Research Database (Denmark)

    Landvad, Katrine

    Ddb1, a component of the E3 ubiquitin ligase CRL4Cdt2, is needed for proper chromosome segregation in fission yeast as ddb1 deleted cells show unequal distribution of DNA to daughter cells and sensitivity to the microtubule destabilising drug TBZ. In this study we show that Δddb1 cells have...

  10. Osteochondritis dissecans (OCD), an endoplasmic reticulum storage disease?

    DEFF Research Database (Denmark)

    Skagen, Peter Storgaard; Horn, T; Kruse, H A

    2011-01-01

    Osteochondritis dissecans (OCD) fragments, cartilage and blood from four patients were used for morphological and molecular analysis. Controls included articular cartilage and blood samples from healthy individuals. Light microscopy and transmission electron microscopy (TEM) showed abnormalities...... in chondrocytes and extracellular matrix of cartilage from OCD patients. Abnormal type II collagen heterofibrils in "bundles" and chondrocytes with abnormal accumulation of matrix proteins in distended rough endoplasmic reticulum were typical findings. Further, Von Kossa staining and TEM showed empty lacunae...... polymorphi