Hepatic overexpression of SIRT1 in mice attenuates endoplasmic reticulum stress and insulin resistance in the liver

PubMed Central

Li, Yu; Xu, Shanqin; Giles, Amber; Nakamura, Kazuto; Lee, Jong Woo; Hou, Xiuyun; Donmez, Gizem; Li, Ji; Luo, Zhijun; Walsh, Kenneth; Guarente, Leonard; Zang, Mengwei

2011-01-01

Endoplasmic reticulum (ER) stress has been implicated in the pathophysiology of human type 2 diabetes (T2DM). Although SIRT1 has a therapeutic effect on metabolic deterioration in T2DM, the precise mechanisms by which SIRT1 improves insulin resistance remain unclear. Here, we demonstrate that adenovirus-mediated overexpression of SIRT1 in the liver of diet-induced insulin-resistant low-density lipoprotein receptor-deficient mice and of genetically obese ob/ob mice attenuates hepatic steatosis and ameliorates systemic insulin resistance. These beneficial effects were associated with decreased mammalian target of rapamycin complex 1 (mTORC1) activity, inhibited the unfolded protein response (UPR), and enhanced insulin receptor signaling in the liver, leading to decreased hepatic gluconeogenesis and improved glucose tolerance. The tunicamycin-induced splicing of X-box binding protein-1 and expression of GRP78 and CHOP were reduced by resveratrol in cultured cells in a SIRT1-dependent manner. Conversely, SIRT1-deficient mouse embryonic fibroblasts challenged with tunicamycin exhibited markedly increased mTORC1 activity and impaired ER homeostasi and insulin signaling. These effects were abolished by mTORC1 inhibition by rapamycin in human HepG2 cells. These studies indicate that SIRT1 serves as a negative regulator of UPR signaling in T2DM and that SIRT1 attenuates hepatic steatosis, ameliorates insulin resistance, and restores glucose homeostasis, largely through the inhibition of mTORC1 and ER stress.—Li, Y., Xu, S., Giles, A., Nakamura, K., Lee, J. W., Hou, X., Donmez, G., Li, J., Luo, Z., Walsh, K., Guarente, L., Zang, M. Hepatic overexpression of SIRT1 in mice attenuates endoplasmic reticulum stress and insulin resistance in the liver. PMID:21321189

Caveolin-1 is a negative regulator of caveolae-mediated endocytosis to the endoplasmic reticulum.

PubMed

Le, Phuong U; Guay, Ginette; Altschuler, Yoram; Nabi, Ivan R

2002-02-01

Caveolae are flask-shaped invaginations at the plasma membrane that constitute a subclass of detergent-resistant membrane domains enriched in cholesterol and sphingolipids and that express caveolin, a caveolar coat protein. Autocrine motility factor receptor (AMF-R) is stably localized to caveolae, and the cholesterol extracting reagent, methyl-beta-cyclodextrin, inhibits its internalization to the endoplasmic reticulum implicating caveolae in this distinct receptor-mediated endocytic pathway. Curiously, the rate of methyl-beta-cyclodextrin-sensitive endocytosis of AMF-R to the endoplasmic reticulum is increased in ras- and abl-transformed NIH-3T3 cells that express significantly reduced levels of caveolin and few caveolae. Overexpression of the dynamin K44A dominant negative mutant via an adenovirus expression system induces caveolar invaginations sensitive to methyl-beta-cyclodextrin extraction in the transformed cells without increasing caveolin expression. Dynamin K44A expression further inhibits AMF-R-mediated endocytosis to the endoplasmic reticulum in untransformed and transformed NIH-3T3 cells. Adenoviral expression of caveolin-1 also induces caveolae in the transformed NIH-3T3 cells and reduces AMF-R-mediated endocytosis to the endoplasmic reticulum to levels observed in untransformed NIH-3T3 cells. Cholesterol-rich detergent-resistant membrane domains or glycolipid rafts therefore invaginate independently of caveolin-1 expression to form endocytosis-competent caveolar vesicles via rapid dynamin-dependent detachment from the plasma membrane. Caveolin-1 stabilizes the plasma membrane association of caveolae and thereby acts as a negative regulator of the caveolae-mediated endocytosis of AMF-R to the endoplasmic reticulum.

Assessing the acidity of high silica chabazite H-SSZ-13 by FTIR using CO as molecular probe: Comparison with H-SAPO-34.

PubMed

Bordiga, Silvia; Regli, Laura; Cocina, Donato; Lamberti, Carlo; Bjørgen, Morten; Lillerud, Karl Petter

2005-02-24

Zeolitic materials based on the chabazite topology, such as H-SAPO-34, possess unique shape-selectivity properties for converting methanol into light olefins. In addition to the topology, zeolite acidity is inherently linked to catalyst activity and selectivity. The acidic properties of high silica chabazite (H-SSZ-13) have attracted much attention in the past decade because the material represents an idealized model system having one acidic site per cage. Conclusions drawn so far have essentially been founded on quantum chemical methods. An experimentally based benchmark of the acidity of H-SSZ-13 has hitherto not been available. In this work, transmission FTIR spectroscopy provides a description of the different acidic sites of H-SSZ-13 by using CO as molecular probe at 70 K. The results demonstrate that H-SSZ-13 is a strongly Brønsted acidic material, essentially having two distinct families of acidic sites. In contrast to numerous preceding reports, we find it fundamental to consider proton distributions among all four possible sites, and do not delimit the interpretations to only two sites. The present data consistently suggest the most abundant family of protons to have three members being located on different crystalline positions on the eight-membered-ring window giving access to the chabazite cage. Consequently, these protons are exposed to two neighboring cages. The second, and less abundant family, is constituted by only one site that is situated on the six-membered ring defining the top/bottom of the barrel-shaped chabazite cage. This proton is therefore only exposed to one cage and requires a higher CO pressure to form adducts. Toward CO, both families of sites possess the same acidity. Parallel experiments were also carried out for the isostructural and commercially important H-SAPO-34 having an equal density of acidic sites. This is the first attempt to directly compare, on an experimental basis, the acidity of these two materials.

Endoplasmic reticulum stress: a novel mechanism and therapeutic target for cardiovascular diseases

PubMed Central

Liu, Mei-qing; Chen, Zhe; Chen, Lin-xi

2016-01-01

Endoplasmic reticulum is a principal organelle responsible for folding, post-translational modifications and transport of secretory, luminal and membrane proteins, thus palys an important rale in maintaining cellular homeostasis. Endoplasmic reticulum stress (ERS) is a condition that is accelerated by accumulation of unfolded/misfolded proteins after endoplasmic reticulum environment disturbance, triggered by a variety of physiological and pathological factors, such as nutrient deprivation, altered glycosylation, calcium depletion, oxidative stress, DNA damage and energy disturbance, etc. ERS may initiate the unfolded protein response (UPR) to restore cellular homeostasis or lead to apoptosis. Numerous studies have clarified the link between ERS and cardiovascular diseases. This review focuses on ERS-associated molecular mechanisms that participate in physiological and pathophysiological processes of heart and blood vessels. In addition, a number of drugs that regulate ERS was introduced, which may be used to treat cardiovascular diseases. This review may open new avenues for studying the pathogenesis of cardiovascular diseases and discovering novel drugs targeting ERS. PMID:26838072

[Endoplasmic reticulum stress in INS-1-3 cell associated with the expression changes of MODY gene pathway].

PubMed

Liu, Y T; Li, S R; Wang, Z; Xiao, J Z

2016-09-13

Objective: To profile the gene expression changes associated with endoplasmic reticulum stress in INS-1-3 cells induced by thapsigargin (TG) and tunicamycin (TM). Methods: Normal cultured INS-1-3 cells were used as a control. TG and TM were used to induce endoplasmic reticulum stress in INS-1-3 cells. Digital gene expression profiling technique was used to detect differentially expressed gene. The changes of gene expression were detected by expression pattern clustering analysis, gene ontology (GO) function and pathway enrichment analysis. Real time polymerase chain reaction (RT-PCR) was used to verify the key changes of gene expression. Results: Compared with the control group, there were 57 (45 up-regulated, 12 down-regulated) and 135 (99 up-regulated, 36 down-regulated) differentially expressed genes in TG and TM group, respectively. GO function enrichment analyses indicated that the main enrichment was in the endoplasmic reticulum. In signaling pathway analysis, the identified pathways were related with endoplasmic reticulum stress, antigen processing and presentation, protein export, and most of all, the maturity onset diabetes of the young (MODY) pathway. Conclusion: Under the condition of endoplasmic reticulum stress, the related expression changes of transcriptional factors in MODY signaling pathway may be related with the impaired function in islet beta cells.

From Endoplasmic Reticulum to Mitochondria: Absence of the Arabidopsis ATP Antiporter Endoplasmic Reticulum Adenylate Transporter1 Perturbs Photorespiration[W

PubMed Central

Hoffmann, Christiane; Plocharski, Bartolome; Haferkamp, Ilka; Leroch, Michaela; Ewald, Ralph; Bauwe, Hermann; Riemer, Jan; Herrmann, Johannes M.; Neuhaus, H. Ekkehard

2013-01-01

The carrier Endoplasmic Reticulum Adenylate Transporter1 (ER-ANT1) resides in the endoplasmic reticulum (ER) membrane and acts as an ATP/ADP antiporter. Mutant plants lacking ER-ANT1 exhibit a dwarf phenotype and their seeds contain reduced protein and lipid contents. In this study, we describe a further surprising metabolic peculiarity of the er-ant1 mutants. Interestingly, Gly levels in leaves are immensely enhanced (26×) when compared with that of wild-type plants. Gly accumulation is caused by significantly decreased mitochondrial glycine decarboxylase (GDC) activity. Reduced GDC activity in mutant plants was attributed to oxidative posttranslational protein modification induced by elevated levels of reactive oxygen species (ROS). GDC activity is crucial for photorespiration; accordingly, morphological and physiological defects in er-ant1 plants were nearly completely abolished by application of high environmental CO2 concentrations. The latter observation demonstrates that the absence of ER-ANT1 activity mainly affects photorespiration (maybe solely GDC), whereas basic cellular metabolism remains largely unchanged. Since ER-ANT1 homologs are restricted to higher plants, it is tempting to speculate that this carrier fulfils a plant-specific function directly or indirectly controlling cellular ROS production. The observation that ER-ANT1 activity is associated with cellular ROS levels reveals an unexpected and critical physiological connection between the ER and other organelles in plants. PMID:23860249

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

DOE Office of Scientific and Technical Information (OSTI.GOV)

Luo, Ying; Li, Shu-Jun; Yang, Jian

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 reticulummore » 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.« less

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

PubMed

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.

Thioredoxin-1 Protects Spinal Cord from Demyelination Induced by Methamphetamine through Suppressing Endoplasmic Reticulum Stress and Inflammation

PubMed Central

Yang, Lihua; Guo, Yinli; Huang, Mengbin; Wu, Xiaoli; Li, Xiang; Chen, Guobing; Li, Ye; Bai, Jie

2018-01-01

Methamphetamine (METH) is a psychostimulant abused around the world. Emerging evidence indicates that METH causes brain damage. However, there are very few reports on METH-induced demyelination. Thioredoxin-1 (Trx-1) is a redox regulating protein and plays the roles in protecting neurons from various stresses. However, whether Trx-1 resists demyelination induced by METH has not been reported. In this study, we found that METH-induced thin myelin sheaths in spinal cord, whereas Trx-1 overexpression transgenic (TG) mice restored the myelin sheaths thickness. The expressions of myelin-associated glycoprotein, myelin basic protein, and cyclin-dependent kinase 5 were decreased by METH, whereas these alterations were blocked in Trx-1 TG mice. The expressions of procaspase-12 and procaspase-3 were decreased by METH, the expression of calpain1 was increased by METH, whereas the alterations were suppressed in Trx-1 TG mice. As same as, the expressions of the extracellular signal-regulated kinase, nuclear factor κB, tumor necrosis factor-alpha, and interleukin-1beta were induced by METH, which were suppressed in Trx-1 TG mice. These data suggest that Trx-1 may play a critical role in resisting the METH-mediated demyelination in spinal cord through regulating endoplasmic reticulum stress and inflammation pathways. PMID:29467717

NO Chemisorption on Cu/SSZ-13: a Comparative Study from Infrared Spectroscopy and DFT Calculations

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhang, Renqin; McEwen, Jean-Sabin; Kollar, Marton

The locations and energies of Cu ions in a Cu/SSZ-13 zeolite catalyst were investigated by density functional theory (DFT) calculations. For 'naked' Cu2+ ions (i.e., Cu2+ ions with no ligands in their coordination spheres other than zeolite lattice oxygen atoms), the more energetically favorable sites are within a 6-membered ring. However, with the presence of various adsorbates, the energy difference between 6- and 8-membered ring locations greatly diminishes. Specifically, Cu2+ ions are substantially stabilized by -OH ligands (as [CuII(OH)]+), making the extra-framework sites in an 8-membered ring energetically more favorable than 6-membered ring sites. Under fully dehydrated high vacuum conditionsmore » with different Si/Al and Cu/Al ratios, three chemisorbed NO species coexist upon exposure of NO to Cu/SSZ-13: NO+, Cu2+-NO and Cu+-NO. The relative signal intensities for these bands vary greatly with Si/Al ratios. The vibrational frequency of chemisorbed NO was found to be very sensitive to the location of Cu2+ ions. On the one hand, with the aid from DFT calculations, the nature for these vibrations can be assigned in detail. On the other hand, the relative intensities for various Cu2+-NO species provide a good measure of the nature of Cu2+ ions as functions of Si/Al and Cu/Al ratios and the presence of humidity. These new findings cast doubt on the generally accepted proposal that only Cu2+ ions located in 6-membered rings are catalytically active for NH3-SCR.« less

Sirtuin 1 protects the aging heart from contractile dysfunction mediated through the inhibition of endoplasmic reticulum stress-mediated apoptosis in cardiac-specific Sirtuin 1 knockout mouse model.

PubMed

Hsu, Yu-Juei; Hsu, Shih-Che; Hsu, Chiao-Po; Chen, Yen-Hui; Chang, Yung-Lung; Sadoshima, Junichi; Huang, Shih-Ming; Tsai, Chien-Sung; Lin, Chih-Yuan

2017-02-01

The longevity regulator Sirtuin 1 is an NAD + -dependent histone deacetylase that regulates endoplasmic reticulum stress and influences cardiomyocyte apoptosis during cardiac contractile dysfunction induced by aging. The mechanism underlying Sirtuin 1 function in cardiac contractile dysfunction related to aging has not been completely elucidated. We evaluated cardiac contractile function, endoplasmic reticulum stress, apoptosis, and oxidative stress in 6- and 12month-old cardiac-specific Sirtuin 1 knockout (Sirt1 -/- ) and control (Sirt1 f/f ) mice using western blotting and immunohistochemistry. Mice were injected with a protein disulphide isomerase inhibitor. For in vitro analysis, cultured H9c2 cardiomyocytes were exposed to either a Sirtuin 1 inhibitor or activator, with or without a mitochondrial inhibitor, to evaluate the effects of Sirtuin 1 on endoplasmic reticulum stress, nitric oxide synthase expression, and apoptosis. The effects of protein disulphide isomerase inhibition on oxidative stress and ER stress-related apoptosis were also investigated. Compared with 6-month-old Sirt1 f/f mice, marked impaired contractility was observed in 12-month-old Sirt1 -/- mice. These findings were consistent with increased endoplasmic reticulum stress and apoptosis in the myocardium. Measures of oxidative stress and nitric oxide synthase expression were significantly higher in Sirt1 -/- mice compared with those in Sirt1 f/f mice at 6months. In vitro experiments revealed increased endoplasmic reticulum stress-mediated apoptosis in H9c2 cardiomyocytes treated with a Sirtuin 1 inhibitor; the effects were ameliorated by a Sirtuin 1 activator. Moreover, consistent with the in vitro findings, impaired cardiac contractility was demonstrated in Sirt1 -/- mice injected with a protein disulphide isomerase inhibitor. The present study demonstrates that the aging heart is characterized by contractile dysfunction associated with increased oxidative stress and endoplasmic reticulum

Prolyl 3-hydroxylase 1 and CRTAP are mutually stabilizing in the endoplasmic reticulum collagen prolyl 3-hydroxylation complex.

PubMed

Chang, Weizhong; Barnes, Aileen M; Cabral, Wayne A; Bodurtha, Joann N; Marini, Joan C

2010-01-15

Null mutations in cartilage-associated protein (CRTAP) and prolyl 3-hydroxylase 1 (P3H1/LEPRE1) cause types VII and VIII OI, respectively, two novel recessive forms of osteogenesis imperfecta (OI) with severe to lethal bone dysplasia and overmodification of the type I collagen helical region. CRTAP and P3H1 form a complex with cyclophilin B (CyPB) in the endoplasmic reticulum (ER) which 3-hydroxylates the Pro986 residue of alpha1(I) and alpha1(II) collagen chains. We investigated the interaction of complex components in fibroblasts from types VII and VIII OI patients. Both CRTAP and P3H1 are absent or reduced on western blots and by immunofluorescence microscopy in cells containing null mutations in either gene. Levels of LEPRE1 or CRTAP transcripts, however, are normal in CRTAP- or LEPRE1-null cells, respectively. Stable transfection of a CRTAP or LEPRE1 expression construct into cells with null mutations for the transfected cDNA restored both CRTAP and P3H1 protein levels. Normalization of collagen helical modification in transfected CRTAP-null cells demonstrated that the restored proteins functioned effectively as a complex. These data indicate that CRTAP and P3H1 are mutually stabilized in the collagen prolyl 3-hydroxylation complex. CyPB levels were unaffected by mutations in either CRTAP or LEPRE1. Proteasomal inhibitors partially rescue P3H1 protein in CRTAP-null cells. In LEPRE1-null cells, secretion of CRTAP is increased compared with control cells and accounts for 15-20% of the decreased CRTAP detected in cells. Thus, mutual stabilization of P3H1 and CRTAP in the ER collagen modification complex is an underlying mechanism for the overlapping phenotype of types VII and VIII OI.

Transformation of Active Sites in Fe/SSZ-13 SCR Catalysts during Hydrothermal Aging: A Spectroscopic, Microscopic, and Kinetics Study

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kovarik, Libor; Washton, Nancy M.; Kukkadapu, Ravi

Fe/SSZ-13 catalysts (Si/Al = 12, Fe loadings 0.37% and 1.20%) were prepared via solution ion-exchange, and hydrothermally aged at 600, 700 and 800 C. The fresh and aged catalysts were characterized with surface area/pore volume analysis, Mössbauer, solid-state MAS NMR, NO titration FTIR spectroscopies, and TEM and APT imaging. Hydrothermal aging causes dealumination of the catalysts, and transformation of various Fe sites. The latter include conversion of free Fe2+ ions to dimeric Fe(III) species, the agglomeration of isolated Fe-ions to Fe-oxide clusters, and incorporation of Al into the Fe-oxide species. These changes result in complex influences on standard SCR andmore » NO/NH3 oxidation reactions. In brief, mild aging causes catalyst performance enhancement for SCR, while harsh aging at 800 C deteriorates SCR performance. In comparison to Fe/zeolites more prone to hydrothermal degradation, this study demonstrates that via the utilization of highly hydrothermally stable Fe/SSZ-13 catalysts, more accurate correlations between various Fe species and their roles in SCR related chemistries can be made. The authors gratefully acknowledge the US Department of Energy (DOE), Energy Efficiency and Renewable Energy, Vehicle Technologies Office for the support of this work. The research described in this paper was performed in the Environmental Molecular Sciences Laboratory (EMSL), a national scientific user facility sponsored by the DOE’s Office of Biological and Environmental Research and located at Pacific Northwest National Laboratory (PNNL). PNNL is operated for the US DOE by Battelle.« less

Activation of autophagy by unfolded proteins during endoplasmic reticulum stress.

PubMed

Yang, Xiaochen; Srivastava, Renu; Howell, Stephen H; Bassham, Diane C

2016-01-01

Endoplasmic reticulum stress is defined as the accumulation of unfolded proteins in the endoplasmic reticulum, and is caused by conditions such as heat or agents that cause endoplasmic reticulum stress, including tunicamycin and dithiothreitol. Autophagy, a major pathway for degradation of macromolecules in the vacuole, is activated by these stress agents in a manner dependent on inositol-requiring enzyme 1b (IRE1b), and delivers endoplasmic reticulum fragments to the vacuole for degradation. In this study, we examined the mechanism for activation of autophagy during endoplasmic reticulum stress in Arabidopsis thaliana. The chemical chaperones sodium 4-phenylbutyrate and tauroursodeoxycholic acid were found to reduce tunicamycin- or dithiothreitol-induced autophagy, but not autophagy caused by unrelated stresses. Similarly, over-expression of BINDING IMMUNOGLOBULIN PROTEIN (BIP), encoding a heat shock protein 70 (HSP70) molecular chaperone, reduced autophagy. Autophagy activated by heat stress was also found to be partially dependent on IRE1b and to be inhibited by sodium 4-phenylbutyrate, suggesting that heat-induced autophagy is due to accumulation of unfolded proteins in the endoplasmic reticulum. Expression in Arabidopsis of the misfolded protein mimics zeolin or a mutated form of carboxypeptidase Y (CPY*) also induced autophagy in an IRE1b-dependent manner. Moreover, zeolin and CPY* partially co-localized with the autophagic body marker GFP-ATG8e, indicating delivery to the vacuole by autophagy. We conclude that accumulation of unfolded proteins in the endoplasmic reticulum is a trigger for autophagy under conditions that cause endoplasmic reticulum stress. © 2015 The Authors The Plant Journal © 2015 John Wiley & Sons Ltd.

The Role of Endoplasmic Reticulum Stress in Human Pathology

PubMed Central

Oakes, Scott A.; Papa, Feroz R.

2017-01-01

Numerous genetic and environmental insults impede the ability of cells to properly fold and posttranslationally modify secretory and transmembrane proteins in the endoplasmic reticulum (ER), leading to a buildup of misfolded proteins in this organelle—a condition called ER stress. ER-stressed cells must rapidly restore protein-folding capacity to match protein-folding demand if they are to survive. In the presence of high levels of misfolded proteins in the ER, an intracellular signaling pathway called the unfolded protein response (UPR) induces a set of transcriptional and translational events that restore ER homeostasis. However, if ER stress persists chronically at high levels, a terminal UPR program ensures that cells commit to self-destruction. Chronic ER stress and defects in UPR signaling are emerging as key contributors to a growing list of human diseases, including diabetes, neurodegeneration, and cancer. Hence, there is much interest in targeting components of the UPR as a therapeutic strategy to combat these ER stress–associated pathologies. PMID:25387057

Receptor-mediated Drp1 oligomerization on endoplasmic reticulum

PubMed Central

Ji, Wei-Ke; Fan, Xintao; Strack, Stefan

2017-01-01

Drp1 is a dynamin guanosine triphosphatase important for mitochondrial and peroxisomal division. Drp1 oligomerization and mitochondrial recruitment are regulated by multiple factors, including interaction with mitochondrial receptors such as Mff, MiD49, MiD51, and Fis. In addition, both endoplasmic reticulum (ER) and actin filaments play positive roles in mitochondrial division, but mechanisms for their roles are poorly defined. Here, we find that a population of Drp1 oligomers is associated with ER in mammalian cells and is distinct from mitochondrial or peroxisomal Drp1 populations. Subpopulations of Mff and Fis1, which are tail-anchored proteins, also localize to ER. Drp1 oligomers assemble on ER, from which they can transfer to mitochondria. Suppression of Mff or inhibition of actin polymerization through the formin INF2 significantly reduces all Drp1 oligomer populations (mitochondrial, peroxisomal, and ER bound) and mitochondrial division, whereas Mff targeting to ER has a stimulatory effect on division. Our results suggest that ER can function as a platform for Drp1 oligomerization, and that ER-associated Drp1 contributes to mitochondrial division. PMID:29158231

Fe/SSZ-13 as an NH3-SCR Catalyst: A Reaction Kinetics and FTIR/Mössbauer Spectroscopic Study

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gao, Feng; Kollar, Marton; Kukkadapu, Ravi K.

2015-03-01

Using a traditional aqueous solution ion-exchange method under a protecting atmosphere of N2, an Fe/SSZ-13 catalyst active in NH3-SCR was synthesized. Mössbauer and FTIR spectroscopies were used to probe the nature of the Fe sites. In the fresh sample, the majority of Fe species are extra-framework cations. The likely monomeric and dimeric ferric ions in hydrated form are [Fe(OH)2]+ and [HO-Fe-O-Fe-OH]2+, based on Mössbauer measurements. During the severe hydrothermal aging (HTA) applied in this study, a majority of cationic Fe species convert to FeAlOx and clustered FeOx species, accompanied by severe dealumination of the SSZ-13 framework. The clustered FeOx speciesmore » do not give a sextet Mössbauer spectrum, indicating that these are highly disordered. However, some Fe species in cationic positions remain after aging as determined from Mössbauer measurements and CO/NO FTIR titrations. NO/NH3 oxidation reaction tests reveal that dehydrated cationic Fe are substantially more active in catalyzing oxidation reactions than the hydrated ones. For NH3-SCR, enhancement of NO oxidation under ‘dry’ conditions promotes SCR rates below ~300 • C. This is due mainly to contribution from the “fast” SCR channel. Above ~300 • C, enhancement of NH3 oxidation under ‘dry’ conditions, however, becomes detrimental to NOx conversions. The HTA sample loses much of the SCR activity below ~300 • C; however, above ~400 • C much of the activity remains. This may suggest that the FeAlOx and FeOx species become active at such elevated temperatures. Alternatively, the high-temperature activity may be maintained by the remaining extra-framework cationic species. For potential practical applications, Fe/SSZ-13 may be used as a co-catalyst for Cu/CHA as integral aftertreatment SCR catalysts on the basis of the stable high temperature activity after hydrothermal aging. The authors gratefully acknowledge the US Department of Energy (DOE), Energy Efficiency and Renewable

Local Environment Sensitivity of the Cu K-Edge XANES Features in Cu-SSZ-13: Analysis from First-Principles.

PubMed

Zhang, Renqin; McEwen, Jean-Sabin

2018-05-22

Cu K-edge X-ray absorption near-edge spectra (XANES) have been widely used to study the properties of Cu-SSZ-13. In this Letter, the sensitivity of the XANES features to the local environment for a Cu + cation with a linear configuration and a Cu 2+ cation with a square-linear configuration in Cu-SSZ-13 is reported. When a Cu + cation is bonded to H 2 O or NH 3 in a linear configuration, the XANES has a strong peak at around 8983 eV. The intensity of this peak decreases as the linear configuration is broken. As for the Cu 2+ cations in a square-planar configuration with a coordination number of 4, two peaks at around 8986 and 8993 eV are found. An intensity decrease for both peaks at around 8986 and 8993 eV is found in an NH 3 _4_Z 2 Cu model as the N-Cu-N angle changes from 180 to 100°. We correlate these features to the variation of the 4p state by PDOS analysis. In addition, the feature peaks for both the Cu + cation and Cu 2+ cation do not show a dependence on the Cu-N bond length. We further show that the feature peaks also change when the coordination number of the Cu cation is varied, while these feature peaks are independent of the zeolite topology. These findings help elucidate the experimental XANES features at an atomic and an electronic level.

Sel1L is indispensable for mammalian endoplasmic reticulum-associated degradation, endoplasmic reticulum homeostasis, and survival

PubMed Central

Sun, Shengyi; Shi, Guojun; Han, Xuemei; Francisco, Adam B.; Ji, Yewei; Mendonça, Nuno; Liu, Xiaojing; Locasale, Jason W.; Simpson, Kenneth W.; Duhamel, Gerald E.; Kersten, Sander; Yates, John R.; Long, Qiaoming; Qi, Ling

2014-01-01

Suppressor/Enhancer of Lin-12-like (Sel1L) is an adaptor protein for the E3 ligase hydroxymethylglutaryl reductase degradation protein 1 (Hrd1) involved in endoplasmic reticulum-associated degradation (ERAD). Sel1L’s physiological importance in mammalian ERAD, however, remains to be established. Here, using the inducible Sel1L knockout mouse and cell models, we show that Sel1L is indispensable for Hrd1 stability, ER homeostasis, and survival. Acute loss of Sel1L leads to premature death in adult mice within 3 wk with profound pancreatic atrophy. Contrary to current belief, our data show that mammalian Sel1L is required for Hrd1 stability and ERAD function both in vitro and in vivo. Sel1L deficiency disturbs ER homeostasis, activates ER stress, attenuates translation, and promotes cell death. Serendipitously, using a biochemical approach coupled with mass spectrometry, we found that Sel1L deficiency causes the aggregation of both small and large ribosomal subunits. Thus, Sel1L is an indispensable component of the mammalian Hrd1 ERAD complex and ER homeostasis, which is essential for protein translation, pancreatic function, and cellular and organismal survival. PMID:24453213

New insights into Cu/SSZ-13 SCR catalyst acidity. Part I: Nature of acidic sites probed by NH 3 titration

DOE Office of Scientific and Technical Information (OSTI.GOV)

Luo, Jinyong; Gao, Feng; Kamasamudram, Krishna

In this work we investigated an unusual acidity feature of a Cu/SSZ-13 catalyst used in selective catalytic reduction of NOx with NH3 (NH3-SCR). In particular, this catalyst showed two distinct NH3 desorption peaks in NH3-TPD measurements, in contrast to single, unresolved desorption peaks observed for other Cu-exchanged zeolites conventionally used in the SCR studies, including its isostructural but chemically different analogue Cu/SAPO-34. We further observed that the intensities of the two TPD peaks, which represented the amount of stored NH3, changed in opposite directions in response to progressive mild hydrothermal aging, while the total storage capacity was preserved. We proposedmore » an explanation for this remarkable behavior, by using model reference samples and additional characterization techniques. At least three NH3 storage sites were identified: two distinct populations of Cu sites responsible for low-temperature NH3 storage, and Brønsted acid sites responsible for high-temperature NH3 storage. Contrary to the commonly accepted mechanism that Brønsted acid site loss during hydrothermal aging is driven by dealumination, we concluded that the decline in the number of Brønsted acid sites upon mild hydrothermal aging for Cu/SSZ-13 was not due to dealumination, but rather transformation of Cu sites, i.e., gradual conversion of ZCuOH (Cu2+ singly coordinated with Zeolite) to Z2Cu (Cu2+ doubly coordinated with Zeolite). This transformation was responsible for the increased low-temperature desorption peak in NH3-TPD since each ZCuOH adsorbed ~1 NH3 molecule while each Z2Cu adsorbed ~2 NH3 molecules under the conditions used here. These findings were used in Part II of this series of studies to develop a method for quantifying hydrothermal ageing of industrial Cu/SSZ-13 SCR catalysts. Authors would like to thank Randall Jines for his help with collecting the reactor data, Nancy W. Washton for measuring the NMR data and Tamas Varga for in-situ XRD

circHIPK2-mediated σ-1R promotes endoplasmic reticulum stress in human pulmonary fibroblasts exposed to silica.

PubMed

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.

Penfluridol induces endoplasmic reticulum stress leading to autophagy in pancreatic cancer.

PubMed

Ranjan, Alok; German, Nadezhda; Mikelis, Constantinos; Srivenugopal, Kalkunte; Srivastava, Sanjay K

2017-06-01

Pancreatic cancer is one of the most aggressive and difficult to treat cancers. Experimental and clinical evidence suggests that high basal state autophagy in pancreatic tumors could induce resistance to chemotherapy. Recently, we have demonstrated that penfluridol suppresses pancreatic tumor growth by autophagy-mediated apoptosis both in vitro and in vivo; however, the mechanism of autophagy induction by penfluridol was not clear. Several studies have established that endoplasmic reticulum stress could lead to autophagy and inhibit tumor progression. In this study, we demonstrated that penfluridol induced endoplasmic reticulum stress in BxPC-3, AsPC-1, and Panc-1 pancreatic cancer cell lines as indicated by upregulation of endoplasmic reticulum stress markers such as binding protein (BIP), C/EBP homologous protein (CHOP) and inositol requiring 1α (IRE1α) after treatment with penfluridol in a concentration-dependent manner. Inhibiting endoplasmic reticulum stress by pretreatment with pharmacological inhibitors such as sodium phenylbutyrate and mithramycin or by silencing CHOP using CHOP small interfering RNA, blocked penfluridol-induced autophagy. These results clearly indicate that penfluridol-induced endoplasmic reticulum stress lead to autophagy in our model. Western blot analysis of subcutaneously implanted AsPC-1 and BxPC-3 tumors as well as orthotopically implanted Panc-1 tumors demonstrated upregulation of BIP, CHOP, and IRE1α expression in the tumor lysates from penfluridol-treated mice as compared to tumors from control mice. Altogether, our study establishes that penfluridol-induced endoplasmic reticulum stress leads to autophagy resulting in reduced pancreatic tumor growth. Our study opens a new therapeutic target for advanced chemotherapies against pancreatic cancer.

Endoplasmic Reticulum Stress and Type 2 Diabetes

PubMed Central

Back, Sung Hoon; Kaufman, Randal J.

2013-01-01

Given the functional importance of the endoplasmic reticulum (ER), an organelle that performs folding, modification, and trafficking of secretory and membrane proteins to the Golgi compartment, the maintenance of ER homeostasis in insulin-secreting β-cells is very important. When ER homeostasis is disrupted, the ER generates adaptive signaling pathways, called the unfolded protein response (UPR), to maintain homeostasis of this organelle. However, if homeostasis fails to be restored, the ER initiates death signaling pathways. New observations suggest that both chronic hyperglycemia and hyperlipidemia, known as important causative factors of type 2 diabetes (T2D), disrupt ER homeostasis to induce unresolvable UPR activation and β-cell death. This review examines how the UPR pathways, induced by high glucose and free fatty acids (FFAs), interact to disrupt ER function and cause β-cell dysfunction and death. PMID:22443930

Bacteria, the endoplasmic reticulum and the unfolded protein response: friends or foes?

PubMed

Celli, Jean; Tsolis, Renée M

2015-02-01

The unfolded protein response (UPR) is a cytoprotective response that is aimed at restoring cellular homeostasis following physiological stress exerted on the endoplasmic reticulum (ER), which also invokes innate immune signalling in response to invading microorganisms. Although it has been known for some time that the UPR is modulated by various viruses, recent evidence indicates that it also has multiple roles during bacterial infections. In this Review, we describe how bacteria interact with the ER, including how bacteria induce the UPR, how subversion of the UPR promotes bacterial proliferation and how the UPR contributes to innate immune responses against invading bacteria.

Functionalization of zeolitic cavities: grafting NH2 groups in framework T sites of B-SSZ-13--a way to obtain basic solids catalysts?

PubMed

Regli, Laura; Bordiga, Silvia; Busco, Claudia; Prestipino, Carmelo; Ugliengo, Piero; Zecchina, Adriano; Lamberti, Carlo

2007-10-10

Insertion of B atoms into an Al-free zeolitic framework with CHA topology results in the formation of B-SSZ-13 zeotype with Si/B = 11. B K-edge NEXAFS testifies that B forms [B(OSi)4] units in a Td-like geometry (sp3-hybridized B atoms). According to B K-edge NEXAFS and IR, template burning results in the formation of [B(OSi)3] units in a D3h-like geometry (sp2-hybridized B atoms) with a break of a B-O-Si bond and the formation of a Si-OH group. The activated material contains B(III) Lewis acid centers able to specifically coordinate bases like NH3. Such [B(OSi)3] units are reactive toward ammonia, resulting in the formation of B-NH2 surface functionality inside the pores of B-SSZ-13 already under mild conditions, i.e., 35 mbar of NH3 at 373 K for 30 min and without crystallinity degradation. A minor fraction of Si-NH2 cannot be excluded owing to the presence of two IR doublets at 3500 and 3430 cm-1 and at 1600 and 1550 cm-1. Ab initio B3LYP/6-31+G(d,p) calculations on a cluster model, supported by a single-point MP2 on B3LYP/6-31+G(D,P) optimized structures, found the break by NH3 of a B-O-Si bond of the [B(OSi)3] unit with formation of [SiOH] and [H2N-B(OSi)2] species to be energetically favored. Comparison between experimental and computed frequency shifts shows them to be in semiquantitative agreement. The high stability of the B-NH2 surface functionality is probed by N K-edge NEXAFS spectra collected under UHV conditions. These findings can open a new route in the preparation of shape selective solid basic catalysts.

[Effect and mechanism of endoplasmic reticulum stress on cisplatin resistance in ovarian carcinoma].

PubMed

Tian, Jing; Hu, Xiaoming; Qu, Quanxin

2014-05-01

The study intended to investigate the effect and mechanism of endoplasmic reticulum stress on cisplatin resistance in ovarian carcinoma. RT-PCR and Western blot were used to test the expression of mTOR and Beclin1 mRNA and protein in ovarian cancer SKOV3 cells after saquinavir induction. MTT assay was used to analyze the influence of saquinavir on cisplatin sensitivity in SKOV3 cells. The IC50 of SKOV3 cells was (5.490 ± 1.148) µg/ml. After induced by Saquinavair 10 µmol/L and 20 µmol/L, the IC50 of SKOV3 cells was increased to (11.199 ± 0.984) µg/ml and (14.906 ± 2.015) µg/ml, respectively. It suggested that the sensitivity of ovarian cancer cells to cisplatin was decreased significantly (P = 0.001). The expression of mTOR and Beclin1 mRNA and protein was significantly different among the five groups: the (Saquinavair+DDP) group of, Saquinavair group, LY294002 group, DDP group and control group (P < 0.001) . The expressions of mTOR and Beclin1 mRNA were highest in the (Saquinavair+DDP) group, 0.684 ± 0.072 and 0.647 ± 0.047, respectively; Secondly, the Saquinavair group, 0.577 ± 0.016 and 0.565 ± 0.037, respectively. The expressions of mTOR and Beclin1 proteins were also highest in the (Saquinavair+DDP) group, 0.624 ± 0.058 and 0.924 ± 0.033, respectively, followed by the Saquinavair group, 0.544 ± 0.019 and 0.712 ± 0.024. 3-MA inhibited the autophagy and restored cisplatin sensitivity in the SKOV3 cells after Saquinavir induced ER stress (P < 0.001). Saquinavir can effectively induce endoplasmic reticulum stress in SKOV3 cells. Endoplasmic reticulum stress can decrease the sensitivity to cisplatin in SKOV3 cells. The mechanism of the decrease of sensitivity to cisplatin in SKOV3 cells may be that ERS regulates cell autophagy through the mTOR and Beclin1 pathways. ERS of tumor cells and autophagy may become a new target to improve the therapeutic effect of chemotherapy and to reverse the drug resistance in tumor treatment.

Arabidopsis SYT1 maintains stability of cortical endoplasmic reticulum networks and VAP27-1-enriched endoplasmic reticulum-plasma membrane contact sites.

PubMed

Siao, Wei; Wang, Pengwei; Voigt, Boris; Hussey, Patrick J; Baluska, Frantisek

2016-11-01

Arabidopsis synaptotagmin 1 (SYT1) is localized on the endoplasmic reticulum-plasma membrane (ER-PM) contact sites in leaf and root cells. The ER-PM localization of Arabidopsis SYT1 resembles that of the extended synaptotagmins (E-SYTs) in animal cells. In mammals, E-SYTs have been shown to regulate calcium signaling, lipid transfer, and endocytosis. Arabidopsis SYT1 was reported to be essential for maintaining cell integrity and virus movement. This study provides detailed insight into the subcellular localization of SYT1 and VAP27-1, another ER-PM-tethering protein. SYT1 and VAP27-1 were shown to be localized on distinct ER-PM contact sites. The VAP27-1-enriched ER-PM contact sites (V-EPCSs) were always in contact with the SYT1-enriched ER-PM contact sites (S-EPCSs). The V-EPCSs still existed in the leaf epidermal cells of the SYT1 null mutant; however, they were less stable than those in the wild type. The polygonal networks of cortical ER disassembled and the mobility of VAP27-1 protein on the ER-PM contact sites increased in leaf cells of the SYT1 null mutant. These results suggest that SYT1 is responsible for stabilizing the ER network and V-EPCSs. © The Author 2016. Published by Oxford University Press on behalf of the Society for Experimental Biology.

The VCP/p97 and YOD1 Proteins Have Different Substrate-dependent Activities in Endoplasmic Reticulum-associated Degradation (ERAD).

PubMed

Sasset, Linda; Petris, Gianluca; Cesaratto, Francesca; Burrone, Oscar R

2015-11-20

Endoplasmic reticulum-associated degradation (ERAD) is an essential quality control mechanism of the folding state of proteins in the secretory pathway that targets unfolded/misfolded polypeptides for proteasomal degradation. The cytosolic p97/valosin-containing protein is an essential ATPase for degradation of ERAD substrates. It has been considered necessary during retro-translocation to extract proteins from the endoplasmic reticulum that are otherwise supposed to accumulate in the endoplasmic reticulum lumen. The activity of the p97-associated deubiquitinylase YOD1 is also required for substrate disposal. We used the in vivo biotinylation retro-translocation assay in mammalian cells under conditions of impaired p97 or YOD1 activity to directly discriminate their requirements and diverse functions in ERAD. Using different ERAD substrates, we found that both proteins participate in two distinct retro-translocation steps. For CD4 and MHC-Iα, which are induced to degradation by the HIV-1 protein Vpu and by the CMV immunoevasins US2 and US11, respectively, p97 and YOD1 have a retro-translocation-triggering role. In contrast, for three other spontaneous ERAD model substrates (NS1, NHK-α1AT, and BST-2/Tetherin), p97 and YOD1 are required in the downstream events of substrate deglycosylation and proteasomal degradation. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Organization of transport from endoplasmic reticulum to Golgi in higher plants.

PubMed

Andreeva, A V; Zheng, H; Saint-Jore, C M; Kutuzov, M A; Evans, D E; Hawes, C R

2000-01-01

In plant cells, the organization of the Golgi apparatus and its interrelationships with the endoplasmic reticulum differ from those in mammalian and yeast cells. Endoplasmic reticulum and Golgi apparatus can now be visualized in plant cells in vivo with green fluorescent protein (GFP) specifically directed to these compartments. This makes it possible to study the dynamics of the membrane transport between these two organelles in the living cells. The GFP approach, in conjunction with a considerable volume of data about proteins participating in the transport between endoplasmic reticulum and Golgi in yeast and mammalian cells and the identification of their putative plant homologues, should allow the establishment of an experimental model in which to test the involvement of the candidate proteins in plants. As a first step towards the development of such a system, we are using Sar1, a small G-protein necessary for vesicle budding from the endoplasmic reticulum. This work has demonstrated that the introduction of Sar1 mutants blocks the transport from endoplasmic reticulum to Golgi in vivo in tobacco leaf epidermal cells and has therefore confirmed the feasibility of this approach to test the function of other proteins that are presumably involved in this step of endomembrane trafficking in plant cells.

The Endoplasmic Reticulum Stress Protein Calreticulin in Diabetic Chronic Kidney Disease

DTIC Science & Technology

2016-07-01

AWARD NUMBER: W81XWH-14-1-0203 TITLE: The Endoplasmic Reticulum Stress Protein Calreticulin in Diabetic Chronic Kidney Disease PRINCIPAL...1 July 2015- 30 June 2016 4. TITLE AND SUBTITLE The Endoplasmic Reticulum Stress Protein Calreticulin in Diabetic Chronic Kidney Disease 5a... kidney targeted microbubble/ultrasound-mediated plasmid delivery. We will also examine non-targeted CRT knockdown in these mice. Aim 2.b: We will

Complex of Fas-associated Factor 1 (FAF1) with Valosin-containing Protein (VCP)-Npl4-Ufd1 and Polyubiquitinated Proteins Promotes Endoplasmic Reticulum-associated Degradation (ERAD)*

PubMed Central

Lee, Jae-Jin; Park, Joon Kyu; Jeong, Jaeho; Jeon, Hyesung; Yoon, Jong-Bok; Kim, Eunice EunKyeong; Lee, Kong-Joo

2013-01-01

Fas-associated factor 1 (FAF1) is a ubiquitin receptor containing multiple ubiquitin-related domains including ubiquitin-associated (UBA), ubiquitin-like (UBL) 1, UBL2, and ubiquitin regulatory X (UBX). We previously showed that N-terminal UBA domain recognizes Lys48-ubiquitin linkage to recruit polyubiquitinated proteins and that a C-terminal UBX domain interacts with valosin-containing protein (VCP). This study shows that FAF1 interacts only with VCP complexed with Npl4-Ufd1 heterodimer, a requirement for the recruitment of polyubiquitinated proteins to UBA domain. Intriguingly, VCP association to C-terminal UBX domain regulates ubiquitin binding to N-terminal UBA domain without direct interaction between UBA and UBX domains. These interactions are well characterized by structural and biochemical analysis. VCP-Npl4-Ufd1 complex is known as the machinery required for endoplasmic reticulum-associated degradation. We demonstrate here that FAF1 binds to VCP-Npl4-Ufd1 complex via UBX domain and polyubiquitinated proteins via UBA domain to promote endoplasmic reticulum-associated degradation. PMID:23293021

Defective interplay between mTORC1 activity and endoplasmic reticulum stress-unfolded protein response in uremic vascular calcification.

PubMed

Panda, Dibyendu K; Bai, Xiuying; Sabbagh, Yves; Zhang, Yan; Zaun, Hans-Christian; Karellis, Angeliki; Koromilas, Antonis E; Lipman, Mark L; Karaplis, Andrew C

2018-06-01

Vascular calcification increases the risk of cardiovascular disease and death in patients with chronic kidney disease (CKD). Increased activity of mammalian target of rapamycin complex 1 (mTORC1) and endoplasmic reticulum (ER) stress-unfolded protein response (UPR) are independently reported to partake in the pathogenesis of vascular calcification in CKD. However, the association between mTORC1 activity and ER stress-UPR remains unknown. We report here that components of the uremic state [activation of the receptor for advanced glycation end products (RAGE) and hyperphosphatemia] potentiate vascular smooth muscle cell (VSMC) calcification by inducing persistent and exaggerated activity of mTORC1. This gives rise to prolonged and excessive ER stress-UPR as well as attenuated levels of sestrin 1 ( Sesn1) and Sesn3 feeding back to inhibit mTORC1 activity. Activating transcription factor 4 arising from the UPR mediates cell death via expression of CCAAT/enhancer-binding protein (c/EBP) homologous protein (CHOP), impairs the generation of pyrophosphate, a potent inhibitor of mineralization, and potentiates VSMC transdifferentiation to the osteochondrocytic phenotype. Short-term treatment of CKD mice with rapamycin, an inhibitor of mTORC1, or tauroursodeoxycholic acid, a bile acid that restores ER homeostasis, normalized mTORC1 activity, molecular markers of UPR, and calcium content of aortas. Collectively, these data highlight that increased and/or protracted mTORC1 activity arising from the uremic state leads to dysregulated ER stress-UPR and VSMC calcification. Manipulation of the mTORC1-ER stress-UPR pathway opens up new therapeutic strategies for the prevention and treatment of vascular calcification in CKD.

CaMKII-dependent endoplasmic reticulum fission by whisker stimulation and during cortical spreading depolarization.

PubMed

Kucharz, Krzysztof; Lauritzen, Martin

2018-04-01

Cortical spreading depolarization waves, the cause underlying migraine aura, are also the markers and mechanism of pathology in the acutely injured human brain. Propagation of spreading depolarization wave uniquely depends on the interaction between presynaptic and postsynaptic glutamate N-methyl-d-aspartate receptors (NMDARs). In the normally perfused brain, even a single wave causes a massive depolarization of neurons and glia, which results in transient loss of neuronal function and depression of the ongoing electrocorticographic activity. Endoplasmic reticulum is the cellular organelle of particular importance for modulation of neurotransmission. Neuronal endoplasmic reticulum structure is assumed to be persistently continuous in neurons, but is rapidly lost within 1 to 2 min of global cerebral ischaemia, i.e. the organelle disintegrates by fission. This phenomenon appears to be timed with the cardiac arrest-induced cortical spreading depolarizations, rather than ensuing cell death. To what extent NMDAR-dependent processes may trigger neuronal endoplasmic reticulum fission and whether fission is reversible in the normally perfused brain is unknown. We used two-photon microscopy to examine neuronal endoplasmic reticulum structural dynamics during whisker stimulation and cortical spreading depolarizations in vivo. Somatosensory stimulation triggered loss of endoplasmic reticulum continuity, a likely outcome of constriction and fission, in dendritic spines within less than 10 s of stimulation, which was spontaneously reversible and recovery to normal took 5 min. The endoplasmic reticulum fission was inhibited by blockade of NMDAR and Ca2+/calmodulin-dependent protein kinase II (CaMKII) activated downstream of the NMDARs, whereas inhibition of guanosine triphosphate hydrolases hindered regain of endoplasmic reticulum continuity, i.e. fusion. In contrast to somatosensory stimulation, endoplasmic reticulum fission during spreading depolarization was widespread and

CCPG1, a cargo receptor required for reticulophagy and endoplasmic reticulum proteostasis.

PubMed

Smith, Matthew D; Wilkinson, Simon

2018-06-19

The importance of selective macroautophagy/autophagy in cellular health is increasingly evident. The selective degradation of portions of the endoplasmic reticulum (ER), or reticulophagy, is an emerging example but requires further mechanistic detail and broad evidence of physiological relevance. In a recent study, we identified CCPG1, an ER-resident transmembrane protein that can bind to Atg8-family proteins and, independently and discretely, to RB1CC1/FIP200. Both of these interactions are required to facilitate CCPG1's function as a reticulophagy cargo receptor. CCPG1 transcripts are inducible by ER stress, providing a direct link between ER stress and reticulophagy. In vivo, CCPG1 prevents the hyper-accumulation of insoluble protein within the ER lumen of pancreatic acinar cells and alleviates ER stress. Accordingly, CCPG1 loss sensitizes the exocrine pancreas to tissue injury.

Toward Rational Design of Cu/SSZ-13 Selective Catalytic Reduction Catalysts: Implications from Atomic-Level Understanding of Hydrothermal Stability

DOE Office of Scientific and Technical Information (OSTI.GOV)

Song, James; Wang, Yilin; Walter, Eric D.

The hydrothermal stability of Cu/SSZ-13 SCR catalysts has been extensively studied, yet atomic level understanding of changes to the zeolite support and the Cu active sites during hydrothermal aging are still lacking. In this work, via the utilization of spectroscopic methods including solid-state 27Al and 29Si NMR, EPR, DRIFTS, and XPS, together with imaging and elemental mapping using STEM, detailed kinetic analyses, and theoretical calculations with DFT, various Cu species, including two types of isolated active sites and CuOx clusters, were precisely quantified for samples hydrothermally aged under varying conditions. This quantification convincingly confirms the exceptional hydrothermal stability of isolatedmore » Cu2+-2Z sites, and the gradual conversion of [Cu(OH)]+-Z to CuOx clusters with increasing aging severity. This stability difference is rationalized from the hydrolysis activation barrier difference between the two isolated sites via DFT. Discussions are provided on the nature of the CuOx clusters, and their possible detrimental roles on catalyst stability. Finally, a few rational design principles for Cu/SSZ-13 are derived rigorously from the atomic-level understanding of this catalyst obtained here. The authors gratefully acknowledge the US Department of Energy (DOE), Energy Efficiency and Renewable Energy, Vehicle Technologies Office for the support of this work. Computing time was granted by a user proposal at the William R. Wiley Environmental Molecular Sciences Laboratory (EMSL) and by the National Energy Research Scientific Computing Center (NERSC). The experimental studies described in this paper were performed in the EMSL, a national scientific user facility sponsored by the DOE’s Office of Biological and Environmental Research and located at Pacific Northwest National Laboratory (PNNL). PNNL is operated for the US DOE by Battelle.« less

Endoplasmic reticulum stress and proteasomal system in amyotrophic lateral sclerosis.

PubMed

Karademir, Betul; Corek, Ceyda; Ozer, Nesrin Kartal

2015-11-01

Protein processing including folding, unfolding and degradation is involved in the mechanisms of many diseases. Unfolded protein response and/or endoplasmic reticulum stress are accepted to be the first steps which should be completed via protein degradation. In this direction, proteasomal system and autophagy play important role as the degradation pathways and controlled via complex mechanisms. Amyotrophic lateral sclerosis is a multifactorial neurodegenerative disease which is also known as the most catastrophic one. Mutation of many different genes are involved in the pathogenesis such as superoxide dismutase 1, chromosome 9 open reading frame 72 and ubiquilin 2. These genes are mainly related to the antioxidant defense systems, endoplasmic reticulum stress related proteins and also protein aggregation, degradation pathways and therefore mutation of these genes cause related disorders.This review focused on the role of protein processing via endoplasmic reticulum and proteasomal system in amyotrophic lateral sclerosis which are the main players in the pathology. In this direction, dysfunction of endoplasmic reticulum associated degradation and related cell death mechanisms that are autophagy/apoptosis have been detailed. Copyright © 2015 Elsevier Inc. All rights reserved.

Targeting the Endoplasmic Reticulum Unfolded Protein Response to Counteract the Oxidative Stress-Induced Endothelial Dysfunction

PubMed Central

Moltedo, Ornella; Faraonio, Raffaella

2018-01-01

In endothelial cells, the tight control of the redox environment is essential for the maintenance of vascular homeostasis. The imbalance between ROS production and antioxidant response can induce endothelial dysfunction, the initial event of many cardiovascular diseases. Recent studies have revealed that the endoplasmic reticulum could be a new player in the promotion of the pro- or antioxidative pathways and that in such a modulation, the unfolded protein response (UPR) pathways play an essential role. The UPR consists of a set of conserved signalling pathways evolved to restore the proteostasis during protein misfolding within the endoplasmic reticulum. Although the first outcome of the UPR pathways is the promotion of an adaptive response, the persistent activation of UPR leads to increased oxidative stress and cell death. This molecular switch has been correlated to the onset or to the exacerbation of the endothelial dysfunction in cardiovascular diseases. In this review, we highlight the multiple chances of the UPR to induce or ameliorate oxidative disturbances and propose the UPR pathways as a new therapeutic target for the clinical management of endothelial dysfunction. PMID:29725497

Endoplasmic reticulum stress implicated in chronic traumatic encephalopathy.

PubMed

Lucke-Wold, Brandon P; Turner, Ryan C; Logsdon, Aric F; Nguyen, Linda; Bailes, Julian E; Lee, John M; Robson, Matthew J; Omalu, Bennet I; Huber, Jason D; Rosen, Charles L

2016-03-01

Chronic traumatic encephalopathy is a progressive neurodegenerative disease characterized by neurofibrillary tau tangles following repetitive neurotrauma. The underlying mechanism linking traumatic brain injury to chronic traumatic encephalopathy has not been elucidated. The authors investigate the role of endoplasmic reticulum stress as a link between acute neurotrauma and chronic neurodegeneration. The authors used pharmacological, biochemical, and behavioral tools to assess the role of endoplasmic reticulum stress in linking acute repetitive traumatic brain injury to the development of chronic neurodegeneration. Data from the authors' clinically relevant and validated rodent blast model were compared with those obtained from postmortem human chronic traumatic encephalopathy specimens from a National Football League player and World Wrestling Entertainment wrestler. The results demonstrated strong correlation of endoplasmic reticulum stress activation with subsequent tau hyperphosphorylation. Various endoplasmic reticulum stress markers were increased in human chronic traumatic encephalopathy specimens, and the endoplasmic reticulum stress response was associated with an increase in the tau kinase, glycogen synthase kinase-3β. Docosahexaenoic acid, an endoplasmic reticulum stress inhibitor, improved cognitive performance in the rat model 3 weeks after repetitive blast exposure. The data showed that docosahexaenoic acid administration substantially reduced tau hyperphosphorylation (t = 4.111, p < 0.05), improved cognition (t = 6.532, p < 0.001), and inhibited C/EBP homology protein activation (t = 5.631, p < 0.01). Additionally the data showed, for the first time, that endoplasmic reticulum stress is involved in the pathophysiology of chronic traumatic encephalopathy. Docosahexaenoic acid therefore warrants further investigation as a potential therapeutic agent for the prevention of chronic traumatic encephalopathy.

Aflatoxin B1 affects apoptosis and expression of death receptor and endoplasmic reticulum molecules in chicken spleen.

PubMed

Zhu, Panpan; Zuo, Zhicai; Zheng, Zhixiang; Wang, Fengyuan; Peng, Xi; Fang, Jing; Cui, Hengmin; Gao, Caixia; Song, Hetao; Zhou, Yi; Liu, Xici

2017-11-21

Aflatoxin B 1 (AFB 1 ) is a natural product of the Aspergillus genus of molds, which grow on several foodstuffs stored in hot moist conditions, and is among the most potent hepatocarcinogens and immunosuppression presently known. The latter was related to the up-regulated apoptosis of immune organs. However, the effect of expression of death receptor and endoplasmic reticulum molecules in AFB 1 -induced apoptosis of chicken splenocytes was largely unknown. The objective of this study was to investigate this unknown field. One hundred and forty four one-day-old chickens were randomly divided into control group (0 mg/kg AFB 1 ) and AFB 1 group (0.6 mg/kg AFB 1 ), respectively and fed with AFB 1 for 21 days. Histological observation demonstrated that AFB 1 caused slight congestion and lymphocytic depletion in the spleen. TUNEL and flow cytometry assays showed the excessive apoptosis of splenocytes provoked by AFB 1 . Moreover, quantitative real-time PCR analysis revealed that AFB 1 induced the elevated mRNA expression of Fas, FasL, TNF-α, TNF-R 1 , Caspase-3, Caspase-8, Caspase-10, Grp78 and Grp94 in the spleen. These findings suggested that AFB 1 could lead the excessive apoptosis and alter the expression of death receptor and endoplasmic reticulum molecules in chicken spleen.

Identification of Two Novel Endoplasmic Reticulum Body-Specific Integral Membrane Proteins1[W][OA

PubMed Central

Yamada, Kenji; Nagano, Atsushi J.; Nishina, Momoko; Hara-Nishimura, Ikuko; Nishimura, Mikio

2013-01-01

The endoplasmic reticulum (ER) body, a large compartment specific to the Brassicales, accumulates β-glucosidase and possibly plays a role in the defense against pathogens and herbivores. Although the ER body is a subdomain of the ER, it is unclear whether any ER body-specific membrane protein exists. In this study, we identified two integral membrane proteins of the ER body in Arabidopsis (Arabidopsis thaliana) and termed them MEMBRANE PROTEIN OF ENDOPLASMIC RETICULUM BODY1 (MEB1) and MEB2. In Arabidopsis, a basic helix-loop-helix transcription factor, NAI1, and an ER body component, NAI2, regulate ER body formation. The expression profiles of MEB1 and MEB2 are similar to those of NAI1, NAI2, and ER body β-glucosidase PYK10 in Arabidopsis. The expression of MEB1 and MEB2 was reduced in the nai1 mutant, indicating that NAI1 regulates the expression of MEB1 and MEB2 genes. MEB1 and MEB2 proteins localize to the ER body membrane but not to the ER network, suggesting that these proteins are specifically recruited to the ER body membrane. MEB1 and MEB2 physically interacted with ER body component NAI2, and they were diffused throughout the ER network in the nai2 mutant, which has no ER body. Heterologous expression of MEB1 and MEB2 in yeast (Saccharomyces cerevisiae) suppresses iron and manganese toxicity, suggesting that MEB1 and MEB2 are metal transporters. These results indicate that the membrane of ER bodies has specific membrane proteins and suggest that the ER body is involved in defense against metal stress as well as pathogens and herbivores. PMID:23166355

Endoplasmic Reticulum Stress and Associated ROS

PubMed Central

Zeeshan, Hafiz Maher Ali; Lee, Geum Hwa; Kim, Hyung-Ryong; Chae, Han-Jung

2016-01-01

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. PMID:26950115

The density of the cell sap and endoplasm of Nitellopsis and Chara

NASA Technical Reports Server (NTRS)

Wayne, R.; Staves, M. P.

1991-01-01

We measured the densities of the cell sap, endoplasm and cell wall of Nitellopsis obtusa and Chara corallina using interference microscopy, refractometry, immersion refractometry, equilibrium sedimentation and chemical microanalysis techniques. These values are important for the determination of many rheological properties of the cytoplasm as well as for understanding buoyancy regulation, dispersal mechanisms and how cells respond to gravity. The average densities of the cell sap, endoplasm and cell wall are 1,006.9, 1,016.7 and 1,371 kg m-3 for Nitellopsis and 1,005.0, 1,013.9, and 1,355.3 kg m-3 for Chara.

Induction of endoplasmic reticulum stress under endotoxin tolerance increases inflammatory responses and decreases Pseudomonas aeruginosa pneumonia.

PubMed

Kim, Sena; Joe, Yeonsoo; Park, Se-Ung; Jeong, Sun Oh; Kim, Jin-Kyung; Park, Seong Hoon; Pae, Hyun-Ock; Surh, Young-Joon; Shin, Jaekyoon; Chung, Hun Taeg

2018-06-20

Endotoxin tolerance develops in the late phase of sepsis to protect cells from an early hyperinflammatory response. Nonetheless, because it induces an immunosuppressive environment, patients with sepsis in its late phase are affected by secondary infections, particularly bacterial pneumonia. Here, we showed that induction of endoplasmic reticulum (ER) stress leads to activation of glycogen synthase kinase 3β (GSK-3β) and X-box-binding protein 1 (XBP-1) in an inositol-requiring enzyme 1α (IRE1α)-mediated manner, which in turn restores the inflammatory response in endotoxin-tolerant macrophages. Animal and in vitro models of endotoxin tolerance were studied along with a model of LPS-induced endotoxin tolerance and a model of cecal ligation and puncture (CLP)-induced endotoxin tolerance. To detect the suppressed inflammatory response during endotoxin tolerance, inflammatory-cytokine expression levels were measured by quantitative real-time PCR and an ELISA. Our research revealed that induction of ER stress alleviated lung injury in a septic host infected with Pseudomonas aeruginosa via the activation of GSK-3β and XBP-1 in an IRE1α-mediated manner. Consequently, in the lungs of the septic host infected with P. aeruginosa, symptoms of pneumonia improved and the infecting bacteria were cleared. Thus, for septic patients, determination of immune status may guide the selection of appropriate immunomodulation, and ER stress can be a novel therapeutic strategy restoring the immune response in patients with endotoxin tolerance. ©2018 Society for Leukocyte Biology.

Permethrin potentiates adipogenesis via intracellular calcium and endoplasmic reticulum stress-mediated mechanisms in 3T3-L1 adipocytes.

PubMed

Xiao, Xiao; Qi, Weipeng; Clark, John M; Park, Yeonhwa

2017-11-01

Permethrin, a pyrethroid insecticide, was previously reported to promote adipogenesis in vitro and weight gain in vivo. The mechanism by which permethrin promotes adipogenesis/obesity, however, has not been fully explored. Intracellular calcium and endoplasmic reticulum (ER) stress have been reported to be linked with adipogenesis and obesity. Because pyrethroid insecticides have been determined to influence intracellular calcium and ER stress in vitro, the purpose of this current study was to investigate whether permethrin potentiates adipogenesis via a change in intracellular calcium, leading to endoplasmic reticulum (ER) stress in 3T3-L1 adipocytes. 3T3-L1 cells were exposed to four different concentrations of permethrin (0.01, 0.1, 1 & 10 μM) for 6 days during differentiation. Treatment of permethrin increased intracellular calcium level in a concentration-dependent manner. Similarly, permethrin treatment increased protein levels of ER stress markers in a concentration-dependent manner. These data suggest that intracellular calcium and ER stress may be involved in permethrin-induced adipogenesis of 3T3-L1 cells. Copyright © 2017 Elsevier Ltd. All rights reserved.

[Over-expressed Bax inhibitor 1 (BI-1) inhibits apoptosis of hippocampal neurons via endoplasmic reticulum IRE1-JNK pathway in rats with subarachnoid hemorrhage].

PubMed

Liu, Jiaxin; Zhou, Shuai; Qian, Xiying; Zhang, Yueting; Zhao, Jianhua

2017-10-01

Objective To investigate the protective effect of lentivirus-mediated BI-1 overexpression on hippocampal neurons in rats with subarachnoid hemorrhage (SAH) and the relationship with endoplasmic reticulum IRE1-JNK signaling pathway. Methods The lentivirus solution of BI-1 over-expression was injected into the brain of rats 24 hours before SAH rat model was established by intravascular puncture method. At 24 hours after modeling, the brain water content and neurological score of the rats were measured. The apoptosis of hippocampal neurons was detected by TUNEL assay. Western blotting was used to detect the expressions of BI-1 protein and endoplasmic reticulum stress (ERS) marker proteins GRP78 and IRE1. ERS in hippocampal neurons of the rats with SAH was intervened by IRE1α-specific inhibitor KIRA6, and then the protein expressions of p-IRE1, p-JNK, Bax, Bcl2 and caspase-3 were detected by Western blotting. Results BI-1 over-expression improved neurobehavioral score, decreased brain water content and hippocampal neuron apoptosis rate, and also down-regulated GRP78 and IRE1 protein levels in the rats with SAH. Both the interference of KIRA6 and the over-expression of BI-1 inhibited the expressions of p-IRE1, p-JNK, Bax and caspase-3, and promoted the expression of anti-apoptotic protein Bcl2. Conclusion Over-expression of BI-1 can inhibit the apoptosis of hippocampal neurons in rats with SAH by inhibiting the activation of ERS-mediated IRE1-JNK signaling pathway, thus ultimately attenuating the early brain injury following SAH.

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

PubMed

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.

Endoplasmic reticulum stress inhibition reduces hypertension through the preservation of resistance blood vessel structure and function.

PubMed

Carlisle, Rachel E; Werner, Kaitlyn E; Yum, Victoria; Lu, Chao; Tat, Victor; Memon, Muzammil; No, Yejin; Ask, Kjetil; Dickhout, Jeffrey G

2016-08-01

Our purpose was to determine if endoplasmic reticulum stress inhibition lowers blood pressure (BP) in hypertension by correcting vascular dysfunction. The spontaneously hypertensive rat (SHR) was used as a model of human essential hypertension with its normotensive control, the Wistar Kyoto rat. Animals were subjected to endoplasmic reticulum stress inhibition with 4-phenylbutyric acid (4-PBA; 1 g/kg per day, orally) for 5 weeks from 12 weeks of age. BP was measured weekly noninvasively and at endpoint with carotid arterial cannulation. Small mesenteric arteries were removed for vascular studies. Function was assessed with a Mulvany-Halpern style myograph, and structure was assessed by measurement of medial-to-lumen ratio in perfusion fixed vessels as well as three-dimensional confocal reconstruction of vessel wall components. Endoplasmic reticulum stress was assessed by quantitative real time-PCR and western blotting; oxidative stress was assessed by 3-nitrotyrosine and dihydroethidium staining. 4-PBA significantly lowered BP in SHR (vehicle 206.1 ± 4.3 vs. 4-PBA 178.9 ± 3.1, systolic) but not Wistar Kyoto. 4-PBA diminished contractility and augmented endothelial-dependent vasodilation in SHR small mesenteric arteries, as well as reducing media-to-lumen ratio. 4-PBA significantly reduced endoplasmic reticulum stress in SHR resistance vessels. Normotensive resistance vessels, treated with the endoplasmic reticulum stress-inducing agent, tunicamycin, show decreased endothelial-dependent vasodilation; this was improved with 4-PBA treatment. 3-Nitrotyrosine and dihydroethidium staining indicated that endoplasmic reticulum stress leads to reactive oxygen species generation resolvable by 4-PBA treatment. Endoplasmic reticulum stress caused endothelial-mediated vascular dysfunction contributing to elevated BP in the SHR model of human essential hypertension.

Link between SSZ ophiolite formation, emplacement and arc inception, Northland, New Zealand: U Pb SHRIMP constraints; Cenozoic SW Pacific tectonic implications

NASA Astrophysics Data System (ADS)

Whattam, Scott A.; Malpas, John; Smith, Ian E. M.; Ali, Jason R.

2006-10-01

New U-Pb age-data from zircons separated from a Northland ophiolite gabbro yield a mean 206Pb/ 238U age of 31.6 ± 0.2 Ma, providing support for a recently determined 28.3 ± 0.2 Ma SHRIMP age of an associated plagiogranite and ˜ 29-26 Ma 40Ar/ 39Ar ages ( n = 9) of basalts of the ophiolite. Elsewhere, Miocene arc-related calc-alkaline andesite dikes which intrude the ophiolitic rocks contain zircons which yield mean 206Pb/ 238U ages of 20.1 ± 0.2 and 19.8 ± 0.2 Ma. The ophiolite gabbro and the andesites both contain rare inherited zircons ranging from 122-104 Ma. The Early Cretaceous zircons in the arc andesites are interpreted as xenocrysts from the Mt. Camel basement terrane through which magmas of the Northland Miocene arc lavas erupted. The inherited zircons in the ophiolite gabbros suggest that a small fraction of this basement was introduced into the suboceanic mantle by subduction and mixed with mantle melts during ophiolite formation. We postulate that the tholeiitic suite of the ophiolite represents the crustal segment of SSZ lithosphere (SSZL) generated in the southern South Fiji Basin (SFB) at a northeast-dipping subduction zone that was initiated at about 35 Ma. The subduction zone nucleated along a pre-existing transform boundary separating circa 45-20 Ma oceanic lithosphere to the north and west of the Northland Peninsula from nascent back arc basin lithosphere of the SFB. Construction of the SSZL propagated southward along the transform boundary as the SFB continued to unzip to the southeast. After subduction of a large portion of oceanic lithosphere by about 26 Ma and collision of the SSZL with New Zealand, compression between the Australian Plate and the Pacific Plate was taken up along a new southwest-dipping subduction zone behind the SSZL. Renewed volcanism began in the oceanic forearc at 25 Ma producing boninitic-like, SSZ and within-plate alkalic and calc-alkaline rocks. Rocks of these types temporally overlap ophiolite emplacement and

TANGO1 recruits ERGIC membranes to the endoplasmic reticulum for procollagen export

PubMed Central

Santos, António JM; Raote, Ishier; Scarpa, Margherita; Brouwers, Nathalie; Malhotra, Vivek

2015-01-01

Previously we showed that membrane fusion is required for TANGO1-dependent export of procollagen VII from the endoplasmic reticulum (ER) (Nogueira, et al., 2014). Along with the t-SNARE Syntaxin 18, we now reveal the complete complement of SNAREs required in this process, t-SNAREs BNIP1 and USE1, and v-SNARE YKT6. TANGO1 recruits YKT6-containing ER Golgi Intermediate Compartment (ERGIC) membranes to procollagen VII-enriched patches on the ER. Moreover residues 1214-1396, that include the first coiled coil of TANGO1, specifically recruit ERGIC membranes even when targeted to mitochondria. TANGO1 is thus pivotal in concentrating procollagen VII in the lumen and recruiting ERGIC membranes on the cytoplasmic surface of the ER. Our data reveal that growth of a mega transport carrier for collagen export from the ER is not by acquisition of a larger patch of ER membrane, but instead by addition of ERGIC membranes to procollagen-enriched domains of the ER by a TANGO1-mediated process. DOI: http://dx.doi.org/10.7554/eLife.10982.001 PMID:26568311

[Effect of endoplasmic reticulum stress in trophocytes on the pathogenesis of intrahepatic cholestasis of pregnancy].

PubMed

Yu, Y; Zhou, C L; Yu, T T; Han, X J; Shi, H Y; Wang, H Z; Shen, J J; He, J

2017-06-25

Objective: To evaluate the effect of endoplasmic reticulum stress in trophocytes, in patients with intrahepatic cholestasis of pregnancy (ICP). Methods: Sixty-one pregnant women who were hospitalized in Women's Hospital, School of Medicine, Zhejiang University from January to December 2015 were recruited. Thirty-one women who were diagnosed as ICP were defined as the ICP group and 30 healthy pregnant women were defined as the control group. The localization and expression intensity of glucose regulated protein 78 (GRP-78) in placental tissues were detected by immunohistochemistry technique. Electronic microscope was used to observe ultra-microstructure change of the endoplasmic reticulum in trophocytes and cell line Swan71. Reverse transcription (RT)-PCR and western blot were used to investigate the expression of GRP-78 mRNA and protein in Swan 71 cell. Results: (1) GRP-78 protein was mainly expressed in the cytoplasm of cytotrophoblasts and syncytiotrophoblasts. The protein expression of GRP-78 in placentas of the ICP group (13.2±2.4) was significantly higher than that in the control group (7.8±1.3, P< 0.01). (2) The volume of endoplasmie reticulum did not increase and the microvilli developed well, with no swelling and no expansion of endoplasmic reticulum in the control group.In the ICP group, microvilli injury, endoplasmic reticulum edema were found; the volume of endoplasmic reticulum increased, with dilation, vacuolation and significant degranulation. After treated with 100 μmol/L cholyglycine for 24 hours, universal dilatation of the endoplasmic reticulum were seen in the Swan71 cells. (3) In Swan71 cells, cholylglycine displayed a concentration-dependent up-regulation on the expression of GRP-78. The expressions of GRP-78 mRNA in 0, 25, 50, 100 μmol/L cholylglycine experimental group were 1.01±0.17, 2.17±0.16, 5.47±0.36, 5.65±0.82, respectively. The expression of GRP-78 protein in 0, 25, 50, 100 μmol/L cholylglycine experimental group were 1.01±0

Gating Behavior of Endoplasmic Reticulum Potassium Channels of Rat Hepatocytes in Diabetes

PubMed Central

Ghasemi, Maedeh; Khodaei, Naser; Salari, Sajjad; Eliassi, Afsaneh; Saghiri, Reza

2014-01-01

Background: Defects in endoplasmic reticulum homeostasis are common occurrences in different diseases, such as diabetes, in which the function of endoplasmic reticulum is disrupted. It is now well established that ion channels of endoplasmic reticulum membrane have a critical role in endoplasmic reticulum luminal homeostasis. Our previous studies showed the presence of an ATP-sensitive cationic channel in endoplasmic reticulum. Therefore, in this study, we examined and compared the activities of this channel in control and diabetic rats using single-channel recording techniques. Method: Male Wistar rats were made diabetic for 2 weeks with a single dose injection of streptozotocin (45 mg/kg). Ion channel incorporation of rough endoplasmic reticulum of diabetic hepatocytes into the bilayer lipid membrane allowed the characterization of K+ channel. Results: Ion channel incorporation of rough endoplasmic reticulum vesicles into the bilayer lipid revealed that the channel current-voltage (I-V) relation with a mean slope conductance of 520 ± 19 pS was unaffected in diabetes. Interestingly, the channel Po-voltage relation was significantly lower in diabetic rats at voltages above +30 mV. Conclusion: We concluded that the endoplasmic reticulum cationic channel is involved in diabetes. Also, this finding could be considered as a goal for further therapeutic plans. PMID:24842143

Gating behavior of endoplasmic reticulum potassium channels of rat hepatocytes in diabetes.

PubMed

Ghasemi, Maedeh; Khodaei, Naser; Salari, Sajjad; Eliassi, Afsaneh; Saghiri, Reza

2014-07-01

Defects in endoplasmic reticulum homeostasis are common occurrences in different diseases, such as diabetes, in which the function of endoplasmic reticulum is disrupted. It is now well established that ion channels of endoplasmic reticulum membrane have a critical role in endoplasmic reticulum luminal homeostasis. Our previous studies showed the presence of an ATP-sensitive cationic channel in endoplasmic reticulum. Therefore, in this study, we examined and compared the activities of this channel in control and diabetic rats using single-channel recording techniques. Male Wistar rats were made diabetic for 2 weeks with a single dose injection of streptozotocin (45 mg/kg). Ion channel incorporation of rough endoplasmic reticulum of diabetic hepatocytes into the bilayer lipid membrane allowed the characterization of K+ channel. Ion channel incorporation of rough endoplasmic reticulum vesicles into the bilayer lipid revealed that the channel current-voltage (I-V) relation with a mean slope conductance of 520 ± 19 pS was unaffected in diabetes. Interestingly, the channel Po-voltage relation was significantly lower in diabetic rats at voltages above +30 mV. We concluded that the endoplasmic reticulum cationic channel is involved in diabetes. Also, this finding could be considered as a goal for further therapeutic plans.

Unraveling the contribution of pancreatic beta-cell suicide in autoimmune type 1 diabetes✩

PubMed Central

Jaberi-Douraki, Majid; Schnell, Santiago; Pietropaolo, Massimo; Khadra, Anmar

2014-01-01

In type 1 diabetes, an autoimmune disease mediated by autoreactive T-cells that attack insulin-secreting pancreatic beta-cells, it has been suggested that disease progression may additionally require protective mechanisms in the target tissue to impede such auto-destructive mechanisms. We hypothesize that the autoimmune attack against beta-cells causes endoplasmic reticulum stress by forcing the remaining beta-cells to synthesize and secrete defective insulin. To rescue beta-cell from the endoplasmic reticulum stress, beta-cells activate the unfolded protein response to restore protein homeostasis and normal insulin synthesis. Here we investigate the compensatory role of unfolded protein response by developing a multi-state model of type 1 diabetes that takes into account beta-cell destruction caused by pathogenic autoreactive T-cells and apoptosis triggered by endoplasmic reticulum stress. We discuss the mechanism of unfolded protein response activation and how it counters beta-cell extinction caused by an autoimmune attack and/or irreversible damage by endoplasmic reticulum stress. Our results reveal important insights about the balance between beta-cell destruction by autoimmune attack (beta-cell homicide) and beta-cell apoptosis by endoplasmic reticulum stress (beta-cell suicide). It also provides an explanation as to why the unfolded protein response may not be a successful therapeutic target to treat type 1 diabetes. PMID:24831415

Ticagrelor protects against AngII-induced endothelial dysfunction by alleviating endoplasmic reticulum stress.

PubMed

Wang, Xiaoyu; Han, Xuejie; Li, Minghui; Han, Yu; Zhang, Yun; Zhao, Shiqi; Li, Yue

2018-05-16

Ticagrelor has been reported to decrease cardiovascular mortality compared with clopidogrel. This benefit cannot be fully explained by the more efficient platelet inhibition. Many studies demonstrated that ticagrelor improved endothelial function, leaving the mechanism elusive though. The present study aims to investigate whether ticagrelor protects against endothelial dysfunction induced by angiotensinII (AngII) through alleviating endoplasmic reticulum (ER) stress. Male Sprague Dawley rats were infused with AngII or vehicle and administrated with ticagrelor or vehicle for 14 days. Reactive oxygen species (ROS) was detected. Aortas from normal mice were incubated with endoplasmic reticulum stress inducer tunicamycin with or without ticagrelor. Vasorecactivity was measured on wire myography. Rat aortic endothelial cells (RAECs) were pretreated with ticagrelor followed by AngII or tunicamycin. Endothelial nitric oxide synthase (eNOS) phosphorylation and ER stress markers were determined by western blotting. Impaired endothelial function, induction of ER stress, reduced eNOS phosphorylation and elevated ROS generation was restored by ticagrelor treatment in vivo. In addition, tunicamycin induced endothelial dysfunction was improved by ticagrelor. In vitro, the induction of ER stress and inhibited eNOS phosphorylation in REACs exposed to AngII as well as tunicamycin was reversed by co-culturing with ticagrelor. In conclusion, ticagrelor protects against AngII-induced endothelial dysfunction via alleviating ER stress. Copyright © 2017. Published by Elsevier Inc.

26 CFR 1.412(c)(1)-3 - Applying the minimum funding requirements to restored plans.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 26 Internal Revenue 5 2013-04-01 2013-04-01 false Applying the minimum funding requirements to..., Stock Bonus Plans, Etc. § 1.412(c)(1)-3 Applying the minimum funding requirements to restored plans. (a) In general—(1) Restoration method. The restoration method is a funding method that adapts the...

Endoplasmic Reticulum Stress in Ischemic and Nephrotoxic Acute Kidney Injury.

PubMed

Yan, Mingjuan; Shu, Shaoqun; Guo, Chunyuan; Tang, Chengyuan; Dong, Zheng

2018-06-12

Acute kidney injury is a medical condition characterized by kidney damage with a rapid decline of renal function, which is associated with high mortality and morbidity. Recent research has further established an intimate relationship between acute kidney injury and chronic kidney disease. Perturbations of kidney cells in acute kidney injury result in the accumulation of unfolded and misfolded proteins in the endoplasmic reticulum, leading to unfolded protein response or endoplasmic reticulum stress. In this review, we analyze the role and regulation of endoplasmic reticulum stress in acute kidney injury triggered by renal ischemia-reperfusion and cisplatin nephrotoxicity. The balance between the two major components of unfolded protein response, the adaptive pathway and the apoptotic pathway, plays a critical role in determining the cell fate in endoplasmic reticulum stress. The adaptive pathway is evoked to attenuate translation, induce chaperones, maintain protein homeostasis, and promote cell survival. Prolonged endoplasmic reticulum stress activates the apoptotic pathway, resulting in the elimination of dysfunctional cells. Therefore, regulating ER stress in kidney cells may provide a therapeutic target in acute kidney injury.

Endoplasmic reticulum stress suppresses lipin-1 expression in 3T3-L1 adipocytes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Takahashi, Nobuhiko, E-mail: ntkhs@hoku-iryo-u.ac.jp; Division of Gastroenterology and Hematology/Oncology, Department of Medicine, Asahikawa Medical University, 2-1-1-1, Midorigaoka-Higashi, Asahikawa, Hokkaido 078-8510; Yoshizaki, Takayuki

Highlights: ► Lipin-1 involves lipid metabolism, adipocyte differentiation, and inflammation. ► Adipose lipin-1 expression is reduced in obesity. ► ER stress suppresses lipin-1 expression in 3T3-L1 adipocytes. ► Activation of PPAR-γ recovers ER stress-induced lipin-1 reduction. -- Abstract: Lipin-1 plays crucial roles in the regulation of lipid metabolism and cell differentiation in adipocytes. In obesity, adipose lipin-1 mRNA expression is decreased and positively correlated with systemic insulin sensitivity. Amelioration of the lipin-1 depletion might be improved dysmetabolism. Although some cytokines such as TNF-α and interleukin-1β reduces adipose lipin-1 expression, the mechanism of decreased adipose lipin-1 expression in obesity remains unclear.more » Recently, endoplasmic reticulum (ER) stress is implicated in the pathogenesis of obesity. Here we investigated the role of ER stress on the lipin-1 expression in 3T3-L1 adipocytes. We demonstrated that lipin-1 expression was suppressed by the treatment with ER stress inducers (tunicamycin and thapsigargin) at transcriptional level. We also showed that constitutive lipin-1 expression could be maintained by peroxisome proliferator-activated receptor-γ in 3T3-L1 adipocytes. Activation of peroxisome proliferator-activated receptor-γ recovered the ER stress-induced lipin-1 suppression. These results suggested that ER stress might be involved in the pathogenesis of obesity through lipin-1 depletion.« less

Isolation of the Copper Redox Steps in the Standard Selective Catalytic Reduction on Cu-SSZ-13

DOE Office of Scientific and Technical Information (OSTI.GOV)

Paolucci, Christopher; Verma, Anuj A.; Bates, Shane A.

2014-10-27

Operando X-ray absorption experiments and density functional theory (DFT) calculations are reported that elucidate the role of copper redox chemistry in the selective catalytic reduction (SCR) of NO over Cu-exchanged SSZ-13. Catalysts prepared to contain only isolated, exchanged CuII ions evidence both CuII and CuI ions under standard SCR conditions at 473 K. Reactant cutoff experiments show that NO and NH3 together are necessary for CuII reduction to CuI. DFT calculations show that NO-assisted NH3 dissociation is both energetically favorable and accounts for the observed CuII reduction. The calculations predict in situ generation of Brønsted sites proximal to CuI uponmore » reduction, which we quantify in separate titration experiments. Both NO and O2 are necessary for oxidation of CuI to CuII, which DFT suggests to occur by a NO2 intermediate. Reaction of Cu-bound NO2 with proximal NH4 + completes the catalytic cycle. N2 is produced in both reduction and oxidation half-cycles.« less

Isolation of the Copper Redox Steps in the Standard Selective Catalytic Reduction on Cu-SSZ-13

DOE Office of Scientific and Technical Information (OSTI.GOV)

Paolucci, Christopher; Verma, Anuj A.; Bates, Shane A.

2014-09-12

Operando X-ray absorption experiments and density functional theory (DFT) calculations are reported that elucidate the role of copper redox chemistry in the selective catalytic reduction (SCR) of NO over Cu-exchanged SSZ-13. Catalysts prepared to contain only isolated, exchanged CuII ions evidence both CuII and CuI ions under standard SCR conditions at 473 K. Reactant cutoff experiments show that NO and NH3 together are necessary for CuII reduction to CuI. DFT calculations show that NO-assisted NH3 dissociation is both energetically favorable and accounts for the observed CuII reduction. The calculations predict in situ generation of Brønsted sites proximal to CuI uponmore » reduction, which we quantify in separate titration experiments. Both NO and O2 are necessary for oxidation of CuI to CuII, which DFT suggests to occur by a NO2 intermediate. Reaction of Cu-bound NO2 with proximal NH4 + completes the catalytic cycle. N2 is produced in both reduction and oxidation half-cycles.« less

Experimental and Computational Interrogation of Fast SCR Mechanism and Active Sites on H-Form SSZ-13

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li, Sichi; Zheng, Yang; Gao, Feng

Experiment and density functional theory (DFT) models are combined to develop a unified, quantitative model of the mechanism and kinetics of fast selective catalytic reduction (SCR) of NO/NO2 mixtures over H-SSZ-13 zeolite. Rates, rate orders, and apparent activation energies collected under differential conditions reveal two distinct kinetic regimes. First-principles thermodynamics simulations are used to determine the relative coverages of free Brønsted sites, chemisorbed NH4+ and physisorbed NH3 as a function of reaction conditions. First-principles metadynamics calculations show that all three sites can contribute to the rate-limiting N-N bond forming step in fast SCR. The results are used to parameterize amore » kinetic model that encompasses the full range of reaction conditions and recovers observed rate orders and apparent activation energies. Observed kinetic regimes are related to changes in most-abundant surface intermediates. Financial support was provided by the National Science Foundation GAOLI program under award number 1258690-CBET. We thank the Center for Research Computing at Notre« less

Rkr1/Ltn1 Ubiquitin Ligase-mediated Degradation of Translationally Stalled Endoplasmic Reticulum Proteins*

PubMed Central

Crowder, Justin J.; Geigges, Marco; Gibson, Ryan T.; Fults, Eric S.; Buchanan, Bryce W.; Sachs, Nadine; Schink, Andrea; Kreft, Stefan G.; Rubenstein, Eric M.

2015-01-01

Aberrant nonstop proteins arise from translation of mRNA molecules beyond the coding sequence into the 3′-untranslated region. If a stop codon is not encountered, translation continues into the poly(A) tail, resulting in C-terminal appendage of a polylysine tract and a terminally stalled ribosome. In Saccharomyces cerevisiae, the ubiquitin ligase Rkr1/Ltn1 has been implicated in the proteasomal degradation of soluble cytosolic nonstop and translationally stalled proteins. Rkr1 is essential for cellular fitness under conditions associated with increased prevalence of nonstop proteins. Mutation of the mammalian homolog causes significant neurological pathology, suggesting broad physiological significance of ribosome-associated quality control. It is not known whether and how soluble or transmembrane nonstop and translationally stalled proteins targeted to the endoplasmic reticulum (ER) are detected and degraded. We generated and characterized model soluble and transmembrane ER-targeted nonstop and translationally stalled proteins. We found that these proteins are indeed subject to proteasomal degradation. We tested three candidate ubiquitin ligases (Rkr1 and ER-associated Doa10 and Hrd1) for roles in regulating abundance of these proteins. Our results indicate that Rkr1 plays the primary role in targeting the tested model ER-targeted nonstop and translationally stalled proteins for degradation. These data expand the catalog of Rkr1 substrates and highlight a previously unappreciated role for this ubiquitin ligase at the ER membrane. PMID:26055716

Compositional Heterogeneity and Spatial Segmentation of Suprasubduction (ssz-type) Ophiolites: Evidence From The Kamchatka Arc

NASA Astrophysics Data System (ADS)

Osipenko, A.; Krylov, K.

In ophiolite complexes from the Eastern Asian accretion belts the spatial heterogeneity of geochemical parameters for different components of an ophiolite sequence is estab- lished: restite mantle-derived peridotites, cumulative layered complex and volcanics. This heterogeneity is displayed as at a regional level (tens - hundred km), and at a level of local structures (hundred i - first tens km). As a rule, distinction is observed on a complex of geochemical parameters (concentration and form of REE spectra, EPG distribution, isotope characteristics, Cr-spinel and pyroxene composition etc.). Revealed at once in several suprasubduction-type ophiolite belts (Kamuikotan, Philip- pines New Guinea etc.), the spatial variations of geochemical parameters have not gradual, and discrete character. For an explanation of the reasons of ophiolite com- positional heterogeneity several mechanisms are offered: (1) tectonical overlapping of various fragments of lithosphere; (2) different specify of deep processes, resulting to compositional heterogeneity of rocks from the same lithosphere level; 3) hetero- geneity of the upper mantle and/or mantle metasomatism; 4) evolution of ophiolites (Shervais, 2001) and/or center of magma generation (mixture of continuous series of melt portions, separated during different stages of progressive mantle source melting (Bazylev et al., 2001)); 5) preservations of relict blocks of low lithosphere and upper mantle from the previous stage in suprasubduction conditions. The authors consider regional geochemical heterogeneity and segmentation of suprasubduction ophiolites (SSZ-type) on an example of peridotites from the Eastern Kamchatka ophiolite belt (EKOB), where sublongitude zones, crossed the basic geological structures of a penin- sula (including EROB) were allocated earlier. For each of zones the complex of geo- chemical attributes, steady is established within the limits of a zone, but distinct from of the characteristics of other zones

TNF/TNFR{sub 1} pathway and endoplasmic reticulum stress are involved in ofloxacin-induced apoptosis of juvenile canine chondrocytes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhang, Fu-Tao; Ding, Yi; Shah, Zahir

Background and purpose: Quinolones cause obvious cartilaginous lesions in juvenile animals by chondrocyte apoptosis, which results in the restriction of their use in pediatric and adolescent patients. Studies showed that chondrocytes can be induced to produce TNFα, and the cisternae of the endoplasmic reticulum in quinolone-treated chondrocytes become dilated. We investigated whether TNF/TNFR{sub 1} pathway and endoplasmic reticulum stress (ERs) are involved in ofloxacin (a typical quinolone)-induced apoptosis of juvenile canine chondrocytes. Experimental approach: Canine juvenile chondrocytes were treated with ofloxacin. Cell survival and apoptosis rates were determined with MTT method and flow cytometry, respectively. The gene expression levels ofmore » the related signaling molecules (TNFα, TNFR{sub 1}, TRADD, FADD and caspase-8) in death receptor pathways and main apoptosis-related molecules (calpain, caspase-12, GADD153 and GRP78) in ERs were measured by qRT-PCR. The gene expression of TNFR{sub 1} was suppressed with its siRNA. The protein levels of TNFα, TNFR{sub 1} and caspase-12 were assayed using Western blotting. Key results: The survival rates decreased while apoptosis rates increased after the chondrocytes were treated with ofloxacin. The mRNA levels of the measured apoptosis-related molecules in death receptor pathways and ERs, and the protein levels of TNFα, TNFR{sub 1} and caspase-12 increased after the chondrocytes were exposed to ofloxacin. The downregulated mRNA expressions of TNFR{sub 1}, Caspase-8 and TRADD, and the decreased apoptosis rates of the ofloxacin-treated chondrocytes occurred after TNFR{sub 1}–siRNA interference. Conclusions and implications: Ofloxacin-induced chondrocyte apoptosis in a time- and concentration-dependent fashion. TNF/TNFR{sub 1} pathway and ERs are involved in ofloxacin-induced apoptosis of juvenile canine chondrocytes in the early stage. - Highlights: • Chondrocyte apoptosis is induced by ofloxacin in a

Unfolded protein response activation compensates endoplasmic reticulum-associated degradation deficiency in Arabidopsis.

PubMed

Li, Qingliang; Wei, Hai; Liu, Lijing; Yang, Xiaoyuan; Zhang, Xiansheng; Xie, Qi

2017-07-01

Abiotic stresses often disrupt protein folding and induce endoplasmic reticulum (ER) stress. There is a sophisticated ER quality control (ERQC) system to mitigate the effects of malfunctioning proteins and maintain ER homeostasis. The accumulation of misfolded proteins in the ER activates the unfolded protein response (UPR) to enhance ER protein folding and the degradation of misfolded proteins mediate by ER-associated degradation (ERAD). That ERQC reduces abiotic stress damage has been well studied in mammals and yeast. However, in plants, both ERAD and UPR have been studied separately and found to be critical for plant abiotic stress tolerance. In this study, we discovered that UPR-associated transcription factors AtbZIP17, AtbZIP28 and AtbZIP60 responded to tunicamycin (TM) and NaCl induced ER stress and subsequently enhanced Arabidopsis thaliana abiotic stress tolerance. They regulated the expression level of ER chaperones and the HRD1-complex components. Moreover, overexpression of AtbZIP17, AtbZIP28 and AtbZIP60 could restore stress tolerance via ERAD in the HRD1-complex mutant hrd3a-2, which suggested that UPR and ERAD have an interactive mechanism in Arabidopsis. © 2017 Institute of Botany, Chinese Academy of Sciences.

Endoplasmic Reticulum Stress in the Diabetic Kidney, the Good, the Bad and the Ugly.

PubMed

Cunard, Robyn

2015-04-20

Diabetic kidney disease is the leading worldwide cause of end stage kidney disease and a growing public health challenge. The diabetic kidney is exposed to many environmental stressors and each cell type has developed intricate signaling systems designed to restore optimal cellular function. The unfolded protein response (UPR) is a homeostatic pathway that regulates endoplasmic reticulum (ER) membrane structure and secretory function. Studies suggest that the UPR is activated in the diabetic kidney to restore normal ER function and viability. However, when the cell is continuously stressed in an environment that lies outside of its normal physiological range, then the UPR is known as the ER stress response. The UPR reduces protein synthesis, augments the ER folding capacity and downregulates mRNA expression of genes by multiple pathways. Aberrant activation of ER stress can also induce inflammation and cellular apoptosis, and modify signaling of protective processes such as autophagy and mTORC activation. The following review will discuss our current understanding of ER stress in the diabetic kidney and explore novel means of modulating ER stress and its interacting signaling cascades with the overall goal of identifying therapeutic strategies that will improve outcomes in diabetic nephropathy.

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

PubMed

Kumar, Ravindra; Kumari, Bandana; Kumar, Manish

2017-01-01

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. 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. 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 83.69%. We have also annotated six different

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

PubMed

Zhang, Jintao; Yi, Man; Zha, Longying; Chen, Siqiang; Li, Zhijia; Li, Cheng; Gong, Mingxing; Deng, Hong; Chu, Xinwei; Chen, Jiehua; Zhang, Zheqing; Mao, Limei; Sun, Suxia

2016-01-01

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 (Atg)3. 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 (si

Endoplasmic reticulum stress increases AT1R mRNA expression via TIA-1-dependent mechanism

PubMed Central

Backlund, Michael; Paukku, Kirsi; Kontula, Kimmo K.; Lehtonen, Jukka Y.A.

2016-01-01

As the formation of ribonucleoprotein complexes is a major mechanism of angiotensin II type 1 receptor (AT1R) regulation, we sought to identify novel AT1R mRNA binding proteins. By affinity purification and mass spectroscopy, we identified TIA-1. This interaction was confirmed by colocalization of AT1R mRNA and TIA-1 by FISH and immunofluorescence microscopy. In immunoprecipitates of endogenous TIA- 1, reverse transcription-PCR amplified AT1R mRNA. TIA-1 has two binding sites within AT1R 3′-UTR. The binding site proximal to the coding region is glyceraldehyde-3-phosphate dehydrogenase (GAPDH)-dependent whereas the distal binding site is not. TIA-1 functions as a part of endoplasmic reticulum (ER) stress response leading to stress granule (SG) formation and translational silencing. We and others have shown that AT1R expression is increased by ER stress-inducing factors. In unstressed cells, TIA-1 binds to AT1R mRNA and decreases AT1R protein expression. Fluorescence microscopy shows that ER stress induced by thapsigargin leads to the transfer of TIA-1 to SGs. In FISH analysis AT1R mRNA remains in the cytoplasm and no longer colocalizes with TIA-1. Thus, release of TIA-1-mediated suppression by ER stress increases AT1R protein expression. In conclusion, AT1R mRNA is regulated by TIA-1 in a ER stress-dependent manner. PMID:26681690

Endoplasmic reticulum stress in spinal and bulbar muscular atrophy: a potential target for therapy

PubMed Central

Montague, Karli; Malik, Bilal; Gray, Anna L.; La Spada, Albert R.; Hanna, Michael G.; Szabadkai, Gyorgy

2014-01-01

Spinal and bulbar muscular atrophy is an X-linked degenerative motor neuron disease caused by an abnormal expansion in the polyglutamine encoding CAG repeat of the androgen receptor gene. There is evidence implicating endoplasmic reticulum stress in the development and progression of neurodegenerative disease, including polyglutamine disorders such as Huntington’s disease and in motor neuron disease, where cellular stress disrupts functioning of the endoplasmic reticulum, leading to induction of the unfolded protein response. We examined whether endoplasmic reticulum stress is also involved in the pathogenesis of spinal and bulbar muscular atrophy. Spinal and bulbar muscular atrophy mice that carry 100 pathogenic polyglutamine repeats in the androgen receptor, and develop a late-onset neuromuscular phenotype with motor neuron degeneration, were studied. We observed a disturbance in endoplasmic reticulum-associated calcium homeostasis in cultured embryonic motor neurons from spinal and bulbar muscular atrophy mice, which was accompanied by increased endoplasmic reticulum stress. Furthermore, pharmacological inhibition of endoplasmic reticulum stress reduced the endoplasmic reticulum-associated cell death pathway. Examination of spinal cord motor neurons of pathogenic mice at different disease stages revealed elevated expression of markers for endoplasmic reticulum stress, confirming an increase in this stress response in vivo. Importantly, the most significant increase was detected presymptomatically, suggesting that endoplasmic reticulum stress may play an early and possibly causal role in disease pathogenesis. Our results therefore indicate that the endoplasmic reticulum stress pathway could potentially be a therapeutic target for spinal and bulbar muscular atrophy and related polyglutamine diseases. PMID:24898351

Selective Catalytic Reduction over Cu/SSZ-13: Linking Homo- and Heterogeneous Catalysis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gao, Feng; Mei, Donghai; Wang, Yilin

Active centers in Cu/SSZ-13 selective catalytic reduction (SCR) catalysts have been recently identified as isolated Cu2+ and [CuII(OH)]+ ions. A redox reaction mechanism has also been established, where Cu-ions cycle between CuI and CuII oxidation states during SCR reaction. While the mechanism for the reduction half-cycle (CuII CuI) is reasonably well understood, that for the oxidation half-cycle (CuI CuII) remains an unsettled debate. Herein we report detailed reaction kinetics on low-temperature standard NH3-SCR, supplemented by DFT calculations, as strong evidence that the low-temperature oxidation half-cycle occurs with the participation of two isolated CuI ions, via formation of a transient [CuI(NH3)2]+-O2-[CuI(NH3)2]+more » intermediate. The feasibility of this reaction mechanism is confirmed from DFT calculations, and the simulated energy barrier and rate constants are consistent with experimental findings. Significantly, the low-temperature standard SCR mechanism proposed here provides full consistency with low-temperature SCR kinetics. The authors gratefully acknowledge the US Department of Energy (DOE), Energy Efficiency and Renewable Energy, Vehicle Technologies Office for the support of this work. The research described in this paper was performed in the Environmental Molecular Sciences Laboratory (EMSL), a national scientific user facility sponsored by the DOE’s Office of Biological and Environmental Research and located at Pacific Northwest National Laboratory (PNNL). PNNL is operated for the US DOE by Battelle.« less

Autophagy modulates endoplasmic reticulum stress-induced cell death in podocytes: A protective role

PubMed Central

Cheng, Yu-Chi; Chang, Jer-Ming; Chen, Chien-An

2015-01-01

Endoplasmic reticulum stress occurs in a variety of patho-physiological mechanisms and there has been great interest in managing this pathway for the treatment of clinical diseases. Autophagy is closely interconnected with endoplasmic reticulum stress to counteract the possible injurious effects related with the impairment of protein folding. Studies have shown that glomerular podocytes exhibit high rate of autophagy to maintain as terminally differentiated cells. In this study, podocytes were exposed to tunicamycin and thapsigargin to induce endoplasmic reticulum stress. Thapsigargin/tunicamycin treatment induced a significant increase in endoplasmic reticulum stress and of cell death, represented by higher GADD153 and GRP78 expression and propidium iodide flow cytometry, respectively. However, thapsigargin/tunicamycin stimulation also enhanced autophagy development, demonstrated by monodansylcadaverine assay and LC3 conversion. To evaluate the regulatory effects of autophagy on endoplasmic reticulum stress-induced cell death, rapamycin (Rap) or 3-methyladenine (3-MA) was added to enhance or inhibit autophagosome formation. Endoplasmic reticulum stress-induced cell death was decreased at 6 h, but was not reduced at 24 h after Rap+TG or Rap+TM treatment. In contrast, endoplasmic reticulum stress-induced cell death increased at 6 and 24 h after 3-MA+TG or 3-MA+TM treatment. Our study demonstrated that thapsigargin/tunicamycin treatment induced endoplasmic reticulum stress which resulted in podocytes death. Autophagy, which counteracted the induced endoplasmic reticulum stress, was simultaneously enhanced. The salvational role of autophagy was supported by adding Rap/3-MA to mechanistically regulate the expression of autophagy and autophagosome formation. In summary, autophagy helps the podocytes from cell death and may contribute to sustain the longevity as a highly differentiated cell lineage. PMID:25322957

Survey of molecular chaperone requirement for the biosynthesis of hamster polyomavirus VP1 protein in Saccharomyces cerevisiae.

PubMed

Valaviciute, Monika; Norkiene, Milda; Goda, Karolis; Slibinskas, Rimantas; Gedvilaite, Alma

2016-07-01

A number of viruses utilize molecular chaperones during various stages of their life cycle. It has been shown that members of the heat-shock protein 70 (Hsp70) chaperone family assist polyomavirus capsids during infection. However, the molecular chaperones that assist the formation of recombinant capsid viral protein 1 (VP1)-derived virus-like particles (VLPs) in yeast remain unclear. A panel of yeast strains with single chaperone gene deletions were used to evaluate the chaperones required for biosynthesis of recombinant hamster polyomavirus capsid protein VP1. The impact of deletion or mild overexpression of chaperone genes was determined in live cells by flow cytometry using enhanced green fluorescent protein (EGFP) fused with VP1. Targeted genetic analysis demonstrated that VP1-EGFP fusion protein levels were significantly higher in yeast strains in which the SSZ1 or ZUO1 genes encoding ribosome-associated complex components were deleted. The results confirmed the participation of cytosolic Hsp70 chaperones and suggested the potential involvement of the Ydj1 and Caj1 co-chaperones and the endoplasmic reticulum chaperones in the biosynthesis of VP1 VLPs in yeast. Likewise, the markedly reduced levels of VP1-EGFP in Δhsc82 and Δhsp82 yeast strains indicated that both Hsp70 and Hsp90 chaperones might assist VP1 VLPs during protein biosynthesis.

Sigma Receptor 1 Modulates Endoplasmic Reticulum Stress in Retinal Neurons

PubMed Central

Ha, Yonju; Dun, Ying; Thangaraju, Muthusamy; Duplantier, Jennifer; Dong, Zheng; Liu, Kebin; Ganapathy, Vadivel

2011-01-01

Purpose. To investigate the mechanism of σ receptor 1 (σR1) neuroprotection in retinal neurons. Methods. Oxidative stress, which is implicated in diabetic retinopathy, was induced in mouse primary ganglion cells (GCs) and RGC-5 cells, and the effect of the σR1 ligand (+)-pentazocine on pro- and anti-apoptotic and endoplasmic reticulum (ER) stress gene expression was examined. Binding of σR1 to BiP, an ER chaperone protein, and σR1 phosphorylation status were examined by immunoprecipitation. Retinas were harvested from Ins2Akita/+ diabetic mice treated with (+)-pentazocine, and the expression of ER stress genes and of the retinal transcriptome was evaluated. Results. Oxidative stress induced the death of primary GCs and RGC-5 cells. The effect was decreased by the application of (+)-pentazocine. Stress increased σR1 binding to BiP and enhanced σR1 phosphorylation in RGC-5 cells. BiP binding was prevented, and σR1 phosphorylation decreased in the presence of (+)-pentazocine. The ER stress proteins PERK, ATF4, ATF6, IRE1α, and CHOP were upregulated in RGC-5 cells during oxidative stress, but decreased in the presence of (+)-pentazocine. A similar phenomenon was observed in retinas of Ins2Akita/+ diabetic mice. Retinal transcriptome analysis of Ins2Akita/+ mice compared with wild-type revealed differential expression of the genes critically involved in oxidative stress, differentiation, and cell death. The expression profile of those genes was reversed when the Ins2Akita/+ mice were treated with (+)-pentazocine. Conclusions. In retinal neurons, the molecular chaperone σR1 binds BiP under stressful conditions; (+)-pentazocine may exert its effects by dissociating σR1 from BiP. As stress in retinal cells increases, phosphorylation of σR1 is increased, which is attenuated when agonists bind to the receptor. PMID:20811050

Sigma receptor 1 modulates endoplasmic reticulum stress in retinal neurons.

PubMed

Ha, Yonju; Dun, Ying; Thangaraju, Muthusamy; Duplantier, Jennifer; Dong, Zheng; Liu, Kebin; Ganapathy, Vadivel; Smith, Sylvia B

2011-01-01

To investigate the mechanism of σ receptor 1 (σR1) neuroprotection in retinal neurons. Oxidative stress, which is implicated in diabetic retinopathy, was induced in mouse primary ganglion cells (GCs) and RGC-5 cells, and the effect of the σR1 ligand (+)-pentazocine on pro- and anti-apoptotic and endoplasmic reticulum (ER) stress gene expression was examined. Binding of σR1 to BiP, an ER chaperone protein, and σR1 phosphorylation status were examined by immunoprecipitation. Retinas were harvested from Ins2Akita/+ diabetic mice treated with (+)-pentazocine, and the expression of ER stress genes and of the retinal transcriptome was evaluated. Oxidative stress induced the death of primary GCs and RGC-5 cells. The effect was decreased by the application of (+)-pentazocine. Stress increased σR1 binding to BiP and enhanced σR1 phosphorylation in RGC-5 cells. BiP binding was prevented, and σR1 phosphorylation decreased in the presence of (+)-pentazocine. The ER stress proteins PERK, ATF4, ATF6, IRE1α, and CHOP were upregulated in RGC-5 cells during oxidative stress, but decreased in the presence of (+)-pentazocine. A similar phenomenon was observed in retinas of Ins2Akita/+ diabetic mice. Retinal transcriptome analysis of Ins2Akita/+ mice compared with wild-type revealed differential expression of the genes critically involved in oxidative stress, differentiation, and cell death. The expression profile of those genes was reversed when the Ins2Akita/+ mice were treated with (+)-pentazocine. In retinal neurons, the molecular chaperone σR1 binds BiP under stressful conditions; (+)-pentazocine may exert its effects by dissociating σR1 from BiP. As stress in retinal cells increases, phosphorylation of σR1 is increased, which is attenuated when agonists bind to the receptor.

VCP and ATL1 regulate endoplasmic reticulum and protein synthesis for dendritic spine formation

PubMed Central

Shih, Yu-Tzu; Hsueh, Yi-Ping

2016-01-01

Imbalanced protein homeostasis, such as excessive protein synthesis and protein aggregation, is a pathogenic hallmark of a range of neurological disorders. Here, using expression of mutant proteins, a knockdown approach and disease mutation knockin mice, we show that VCP (valosin-containing protein), together with its cofactor P47 and the endoplasmic reticulum (ER) morphology regulator ATL1 (Atlastin-1), regulates tubular ER formation and influences the efficiency of protein synthesis to control dendritic spine formation in neurons. Strengthening the significance of protein synthesis in dendritic spinogenesis, the translation blocker cyclohexamide and the mTOR inhibitor rapamycin reduce dendritic spine density, while a leucine supplement that increases protein synthesis ameliorates the dendritic spine defects caused by Vcp and Atl1 deficiencies. Because VCP and ATL1 are the causative genes of several neurodegenerative and neurodevelopmental disorders, we suggest that impaired ER formation and inefficient protein synthesis are significant in the pathogenesis of multiple neurological disorders. PMID:26984393

VCP and ATL1 regulate endoplasmic reticulum and protein synthesis for dendritic spine formation.

PubMed

Shih, Yu-Tzu; Hsueh, Yi-Ping

2016-03-17

Imbalanced protein homeostasis, such as excessive protein synthesis and protein aggregation, is a pathogenic hallmark of a range of neurological disorders. Here, using expression of mutant proteins, a knockdown approach and disease mutation knockin mice, we show that VCP (valosin-containing protein), together with its cofactor P47 and the endoplasmic reticulum (ER) morphology regulator ATL1 (Atlastin-1), regulates tubular ER formation and influences the efficiency of protein synthesis to control dendritic spine formation in neurons. Strengthening the significance of protein synthesis in dendritic spinogenesis, the translation blocker cyclohexamide and the mTOR inhibitor rapamycin reduce dendritic spine density, while a leucine supplement that increases protein synthesis ameliorates the dendritic spine defects caused by Vcp and Atl1 deficiencies. Because VCP and ATL1 are the causative genes of several neurodegenerative and neurodevelopmental disorders, we suggest that impaired ER formation and inefficient protein synthesis are significant in the pathogenesis of multiple neurological disorders.

Lactobacillus acidophilus suppresses intestinal inflammation by inhibiting endoplasmic reticulum stress.

PubMed

Kim, Da Hye; Kim, Soochan; Lee, Jin Ha; Kim, Jae Hyeon; Che, Xiumei; Ma, Hyun Woo; Seo, Dong Hyuk; Kim, Tae Il; Kim, Won Ho; Kim, Seung Won; Cheon, Jae Hee

2018-06-22

Nuclear factor kappa B (NF-κB) activation and endoplasmic reticulum (ER) stress signaling play significant roles in the pathogenesis of inflammatory bowel disease (IBD). Thus, we evaluated whether new therapeutic probiotics have anti-colitic effects and we investigated their mechanisms related to NF-κB and ER-stress pathways. Luciferase, nitric oxide (NO), and cytokine assays using HT-29 or RAW264.7 cells were conducted. Mouse colitis was induced using dextran sulfate sodium (DSS) and confirmed by disease activity index and histology. Macrophages and T-cell subsets in isolated peritoneal cavity cells (PCCs) and splenocytes were analyzed by flow cytometry. Gene and cytokine expression profiles were determined using RT-PCR. Lactobacillus acidophilus (LA1) and Pediococcus pentosaceus inhibited NO production in RAW264.7 cells, but only LA1 inhibited Tnfa and induced Il10 expression. LA1 increased the life span of DSS-treated mice and attenuated the severity of colitis by inducing M2 macrophages in PCCs and Th2 and Treg cells in splenocytes. The restoration of goblet cells in the colon was accompanied by the induction of Il10 expression and the suppression of proinflammatory cytokines. Additionally, we found that LA1 exerts an anti-colitic effect by improving ER stress in HT-29 cells as well as in vivo. We showed that LA1 significantly interferes with ER stress and suppresses NF-κB activation. Our findings suggest that LA1 can be used as a potent immunomodulator in IBD treatment and the regulation of ER stress may have significant implications in treating IBD. This article is protected by copyright. All rights reserved.

Nuclear pore complex integrity requires Lnp1, a regulator of cortical endoplasmic reticulum

PubMed Central

Casey, Amanda K.; Chen, Shuliang; Novick, Peter; Ferro-Novick, Susan; Wente, Susan R.

2015-01-01

The nuclear envelope (NE) and endoplasmic reticulum (ER) are components of the same contiguous membrane system and yet have distinct cellular functions. Mounting evidence suggests roles for some ER proteins in the NE for proper nuclear pore complex (NPC) structure and function. In this study, we identify a NE role in Saccharomyces cerevisiae for Lnp1 and Sey1, proteins required for proper cortical ER formation. Both lnp1Δ and sey1Δ mutants exhibit synthetic genetic interactions with mutants in genes encoding key NPC structural components. Both Lnp1 and Sey1 physically associate with other ER components that have established NPC roles, including Rtn1, Yop1, Pom33, and Per33. Of interest, lnp1Δ rtn1Δ mutants but not rtn1Δ sey1Δ mutants exhibit defects in NPC distribution. Furthermore, the essential NPC assembly factor Ndc1 has altered interactions in the absence of Sey1. Lnp1 dimerizes in vitro via its C-terminal zinc finger motif, a property that is required for proper ER structure but not NPC integrity. These findings suggest that Lnp1's role in NPC integrity is separable from functions in the ER and is linked to Ndc1 and Rtn1 interactions. PMID:26041935

26 CFR 1.412(c)(1)-3T - Applying the minimum funding requirements to restored plans (temporary).

Code of Federal Regulations, 2013 CFR

2013-04-01

... 26 Internal Revenue 5 2013-04-01 2013-04-01 false Applying the minimum funding requirements to...-Sharing, Stock Bonus Plans, Etc. § 1.412(c)(1)-3T Applying the minimum funding requirements to restored plans (temporary). (a) In general—(1) Restoration method. The restoration method is a funding method...

HLA-B35 Upregulates Endothelin-1 and Downregulates Endothelial Nitric Oxide Synthase via Endoplasmic Reticulum Stress Response in Endothelial Cells

PubMed Central

Lenna, Stefania; Townsend, Danyelle M.; Tan, Filemon K.; Kapanadze, Bagrat; Markiewicz, Malgorzata; Trojanowska, Maria; Scorza, Raffaella

2013-01-01

The presence of the HLA-B35 allele has emerged as an important risk factor for the development of isolated pulmonary hypertension in patients with scleroderma, however the mechanisms underlying this association have not been fully elucidated. The goal of our study was to determine the molecular mechanisms that mediate the biological effects of HLA-B35 in endothelial cells (ECs). Our data demonstrate that HLA-B35 expression at physiological levels via adenoviral vector resulted in significantly increased endothelin-1 (ET-1) and a significantly decreased endothelial NO synthase (eNOS), mRNA, and protein levels. Furthermore, HLA-B35 greatly upregulated expression of chaperones, including heat shock proteins (HSPs) HSP70 (HSPA1A and HSPA1B) and HSP40 (DNAJB1 and DNAJB9), suggesting that HLA-B35 induces the endoplasmic reticulum (ER) stress and unfolded protein response in ECs. Examination of selected mediators of the unfolded protein response, including H chain binding protein (BiP; GRP78), C/Ebp homologous protein (CHOP; GADD153), endoplasmic reticulum oxidase, and protein disulfide isomerase has revealed a consistent increase of BiP expression levels. Accordingly, thapsigargin, a known ER stress inducer, stimulated ET-1mRNAand protein levels in ECs. This study suggests that HLA-B35 could contribute to EC dysfunction via ER stress-mediated induction of ET-1 in patients with pulmonary hypertension. PMID:20335527

HLA-B35 upregulates endothelin-1 and downregulates endothelial nitric oxide synthase via endoplasmic reticulum stress response in endothelial cells.

PubMed

Lenna, Stefania; Townsend, Danyelle M; Tan, Filemon K; Kapanadze, Bagrat; Markiewicz, Malgorzata; Trojanowska, Maria; Scorza, Raffaella

2010-05-01

The presence of the HLA-B35 allele has emerged as an important risk factor for the development of isolated pulmonary hypertension in patients with scleroderma, however the mechanisms underlying this association have not been fully elucidated. The goal of our study was to determine the molecular mechanisms that mediate the biological effects of HLA-B35 in endothelial cells (ECs). Our data demonstrate that HLA-B35 expression at physiological levels via adenoviral vector resulted in significantly increased endothelin-1 (ET-1) and a significantly decreased endothelial NO synthase (eNOS), mRNA, and protein levels. Furthermore, HLA-B35 greatly upregulated expression of chaperones, including heat shock proteins (HSPs) HSP70 (HSPA1A and HSPA1B) and HSP40 (DNAJB1 and DNAJB9), suggesting that HLA-B35 induces the endoplasmic reticulum (ER) stress and unfolded protein response in ECs. Examination of selected mediators of the unfolded protein response, including H chain binding protein (BiP; GRP78), C/Ebp homologous protein (CHOP; GADD153), endoplasmic reticulum oxidase, and protein disulfide isomerase has revealed a consistent increase of BiP expression levels. Accordingly, thapsigargin, a known ER stress inducer, stimulated ET-1 mRNA and protein levels in ECs. This study suggests that HLA-B35 could contribute to EC dysfunction via ER stress-mediated induction of ET-1 in patients with pulmonary hypertension.

Characterization of wheat endoplasmic reticulum oxidoreductin 1 and its application in Chinese steamed bread.

PubMed

Liu, Guang; Wang, JingJing; Hou, Yi; Huang, Yan-Bo; Wang, JiaJia; Li, Cunzhi; Guo, ShiJun; Li, Lin; Hu, Song-Qing

2018-08-01

This study investigated characteristics of recombinant wheat Endoplasmic Reticulum Oxidoreductin 1 (wEro1) and its influence on Chinese steamed bread (CSB) qualities. The purified wEro1 monomer, which contained two conserved redox active motif sites, bound to flavin adenine dinucleotide (FAD) cofactor with a molecular weight of ∼47 kDa. wEro1 catalyzed the reduction of both bound and free FAD, and its reduction activity of free FAD reached 7.8 U/mg. Moreover, wEro1 catalyzed the oxidation of dithiothreitol and wheat protein disulfide isomerase (wPDI). Both glutathione and the reduced ribonuclease could work as electron donors for wEro1 in catalyzing the oxidation of wPDI. Additionally, wEro1 supplementation improved the CSB qualities with an increased specific volume of CSB and decreased crumb hardness, which was attributed to water-insoluble wheat proteins increasing and gluten network strengthening. The results give an understanding of the properties and function of wEro1 to facilitate its application especially in the flour-processing industry. Copyright © 2018 Elsevier Ltd. All rights reserved.

Endoplasmic reticulum stress increases AT1R mRNA expression via TIA-1-dependent mechanism.

PubMed

Backlund, Michael; Paukku, Kirsi; Kontula, Kimmo K; Lehtonen, Jukka Y A

2016-04-20

As the formation of ribonucleoprotein complexes is a major mechanism of angiotensin II type 1 receptor (AT1R) regulation, we sought to identify novel AT1R mRNA binding proteins. By affinity purification and mass spectroscopy, we identified TIA-1. This interaction was confirmed by colocalization of AT1R mRNA and TIA-1 by FISH and immunofluorescence microscopy. In immunoprecipitates of endogenous TIA- 1, reverse transcription-PCR amplified AT1R mRNA. TIA-1 has two binding sites within AT1R 3'-UTR. The binding site proximal to the coding region is glyceraldehyde-3-phosphate dehydrogenase (GAPDH)-dependent whereas the distal binding site is not. TIA-1 functions as a part of endoplasmic reticulum (ER) stress response leading to stress granule (SG) formation and translational silencing. We and others have shown that AT1R expression is increased by ER stress-inducing factors. In unstressed cells, TIA-1 binds to AT1R mRNA and decreases AT1R protein expression. Fluorescence microscopy shows that ER stress induced by thapsigargin leads to the transfer of TIA-1 to SGs. In FISH analysis AT1R mRNA remains in the cytoplasm and no longer colocalizes with TIA-1. Thus, release of TIA-1-mediated suppression by ER stress increases AT1R protein expression. In conclusion, AT1R mRNA is regulated by TIA-1 in a ER stress-dependent manner. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

Isolation of Endoplasmic Reticulum Fractions from Mammary Epithelial Tissue.

PubMed

Chanat, Eric; Le Parc, Annabelle; Lahouassa, Hichem; Badaoui, Bouabid

2016-06-01

In the mammary glands of lactating animals, the mammary epithelial cells that surround the lumen of the acini produce and secrete copious amounts of milk. Functional differentiation of these mammary epithelial cells depends on the development of high-efficiency secretory pathways, notably for protein and lipid secretion. Protein secretion is a fundamental process common to all animal cells that involves a subset of cellular organelles, including the endoplasmic reticulum and the Golgi apparatus. In contrast, en masse secretion of triglycerides and cholesterol esters in the form of milk fat globules is a unique feature of the mammary epithelial cell. Cytoplasmic lipid droplets, the intracellular precursors of milk fat globules, originate from the endoplasmic reticulum, as do most milk-specific proteins. This organelle is therefore pivotal in the biogenesis of milk components. Fractionation of the cell into its subcellular parts is an approach that has proven very powerful for understanding organelle function and for studying the specific role of an organelle in a given cell activity. Here we describe a method for the purification of both smooth and rough microsomes, the membrane-bound endoplasmic reticulum fragments that form from endoplasmic reticulum domains when cells are broken up, from mammary gland tissue at lactation.

Unfolded Protein Response of the Endoplasmic Reticulum in Tumor Progression and Immunogenicity

PubMed Central

Yoo, Yoon Seon; Han, Hye Gyeong

2017-01-01

The endoplasmic reticulum (ER) is a pivotal regulator of folding, quality control, trafficking, and targeting of secreted and transmembrane proteins, and accordingly, eukaryotic cells have evolved specialized machinery to ensure that the ER enables these proteins to acquire adequate folding and maturation in the presence of intrinsic and extrinsic insults. This adaptive capacity of the ER to intrinsic and extrinsic perturbations is important for maintaining protein homeostasis, which is termed proteostasis. Failure in adaptation to these perturbations leads to accumulation of misfolded or unassembled proteins in the ER, which is termed ER stress, resulting in the activation of unfolded protein response (UPR) of the ER and the execution of ER-associated degradation (ERAD) to restore homeostasis. Furthermore, both of the two axes play key roles in the control of tumor progression, inflammation, immunity, and aging. Therefore, understanding UPR of the ER and subsequent ERAD will provide new insights into the pathogenesis of many human diseases and contribute to therapeutic intervention in these diseases. PMID:29430279

Multivesicular body formation enhancement and exosome release during endoplasmic reticulum stress

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kanemoto, Soshi; Nitani, Ryota; Murakami, Tatsuhiko

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 ormore » PERK deficient cells, exosome release was not upregulated, indicating that IRE1- and PERK-mediated pathways are involved in ER stress-dependent exosome release. - Highlights: • Endoplasmic reticulum (ER) stress induces multivesicular body (MVB) formation. • ER stress transducers IRE1 and PERK mediate MVB formation. • Exosome release is enhanced after ER stress. • IRE1 or PERK deficiency blocks upregulation of ER stress-dependent exosome release.« less

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

PubMed

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.

A meta-analysis of the efficacy of sulfasalazine in comparison with 5-aminosalicylates in the induction of improvement and maintenance of remission in patients with ulcerative colitis.

PubMed

Nikfar, Shekoufeh; Rahimi, Roja; Rezaie, Ali; Abdollahi, Mohammad

2009-06-01

Historically, sulfasalazine (SSZ) and 5-aminosalicylates (5-ASAs) have been a mainstay of mild-to-moderate ulcerative colitis (UC) remission induction and maintenance therapy. Considering the pivotal role of intestinal microbial flora in pathophysiology of UC and antimicrobial activity of sulfapyridine, we hypothesized that SSZ might be more effective than 5-ASAs in the management of UC. To compare the efficacy and tolerability of SSZ with each of the 5-ASAs (mesalamine, olsalazine, and balsalazide) by a meta-analysis technique. Pubmed, Embase, Scopus, Web of Science, and Cochrane Central Register of Controlled Trials were searched for studies compared efficacy and/or tolerability of SSZ with 5-ASAs in the management of UC. The search terms were: "sulfasalazine" or "sulfasalazine" and "5-aminosalicylic acid," "mesalazine," "mesalamine," "olsalazine" or "balsalazide" and "ulcerative colitis." Data were collected from 1966 to April 2008. There was no language restriction. "Overall improvement," "relapse rate," "total adverse events," and "withdrawals because of adverse events" were the key outcomes of interest. Twenty randomized placebo controlled trials met our criteria and were included in the meta-analysis. Comparison of SSZ with mesalamine yielded a nonsignificant relative risk (RR) of 1.04 (95% confidence interval of 0.89-1.21, P = 0.63) for overall improvement, a nonsignificant RR of 0.98 (95% CI 0.78-1.23, P = 0.85) for relapse, a nonsignificant RR of 0.76 (95% CI 0.54-1.07, P = 0.11) for any adverse events, and a nonsignificant RR of 0.78 (95% CI 0.46-1.3, P = 0.33) for withdrawals due to adverse events. Comparison of SSZ with olsalazine yielded a nonsignificant RR of 1.14 (95% CI 0.91-1.43, P = 0.25) for overall improvement, a nonsignificant RR of 0.93 (95% CI 0.77-1.12, P = 0.42) for relapse, a nonsignificant RR of 1.21 (95% CI 0.9-1.61, P = 0.20) for any adverse events, and a nonsignificant RR of 1.53 (95% CI 0.93-2.52, P = 0.09) for withdrawals due to

The mitochondrial translocator protein, TSPO, inhibits HIV-1 envelope glycoprotein biosynthesis via the endoplasmic reticulum-associated protein degradation pathway.

PubMed

Zhou, Tao; Dang, Ying; Zheng, Yong-Hui

2014-03-01

The HIV-1 Env glycoprotein is folded in the endoplasmic reticulum (ER), which is necessary for viral entry and replication. Currently, it is still unclear how this process is regulated. The glycoprotein folding in the ER is controlled by the ER-associated protein degradation (ERAD) pathway, which specifically targets misfolded proteins for degradation. Previously, we reported that HIV-1 replication is restricted in the human CD4(+) T cell line CEM.NKR (NKR). To understand this mechanism, we first analyzed cellular protein expression in NKR cells and discovered that levels of the mitochondrial translocator protein TSPO were upregulated by ∼64-fold. Notably, when NKR cells were treated with TSPO antagonist PK-11195, Ro5-4864, or diazepam, HIV restriction was completely disrupted, and TSPO knockdown by short hairpin RNAs (shRNAs) achieved a similar effect. We next analyzed viral protein expression, and, interestingly, we discovered that Env expression was specifically inhibited. Both TSPO knockdown and treatment with TSPO antagonist could restore Env expression in NKR cells. We further discovered that Env proteins were rapidly degraded and that kifunensine, an ERAD pathway inhibitor, could restore Env expression and viral replication, indicating that Env proteins were misfolded and degraded through the ERAD pathway in NKR cells. We also knocked out the TSPO gene in 293T cells using CRISPR/Cas9 (clustered, regularly interspaced, short palindromic repeat [CRISPR]/CRISPR-associated-9) technology and found that TSPO could similarly inhibit Env expression in these cells. Taken together, these results demonstrate that TSPO inhibits Env protein expression through the ERAD pathway and suggest that mitochondria play an important role in regulating the Env folding process. The HIV-1 Env glycoprotein is absolutely required for viral infection, and an understanding of its expression pathway in infected cells will identify new targets for antiretroviral therapies. Env proteins

ERManI (Endoplasmic Reticulum Class I α-Mannosidase) Is Required for HIV-1 Envelope Glycoprotein Degradation via Endoplasmic Reticulum-associated Protein Degradation Pathway.

PubMed

Zhou, Tao; Frabutt, Dylan A; Moremen, Kelley W; Zheng, Yong-Hui

2015-09-04

Previously, we reported that the mitochondrial translocator protein (TSPO) induces HIV-1 envelope (Env) degradation via the endoplasmic reticulum (ER)-associated protein degradation (ERAD) pathway, but the mechanism was not clear. Here we investigated how the four ER-associated glycoside hydrolase family 47 (GH47) α-mannosidases, ERManI, and ER-degradation enhancing α-mannosidase-like (EDEM) proteins 1, 2, and 3, are involved in the Env degradation process. Ectopic expression of these four α-mannosidases uncovers that only ERManI inhibits HIV-1 Env expression in a dose-dependent manner. In addition, genetic knock-out of the ERManI gene MAN1B1 using CRISPR/Cas9 technology disrupts the TSPO-mediated Env degradation. Biochemical studies show that HIV-1 Env interacts with ERManI, and between the ERManI cytoplasmic, transmembrane, lumenal stem, and lumenal catalytic domains, the catalytic domain plays a critical role in the Env-ERManI interaction. In addition, functional studies show that inactivation of the catalytic sites by site-directed mutagenesis disrupts the ERManI activity. These studies identify ERManI as a critical GH47 α-mannosidase in the ER-associated protein degradation pathway that initiates the Env degradation and suggests that its catalytic domain and enzymatic activity play an important role in this process. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

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

PubMed

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.

26 CFR 1.412(c)(1)-3 - Applying the minimum funding requirements to restored plans.

Code of Federal Regulations, 2011 CFR

2011-04-01

...) In general—(1) Restoration method. The restoration method is a funding method that adapts the... spread gain method that maintains an unfunded liability. A plan may adopt any cost method that satisfies...

Endoplasmic Reticulum Stress in Sepsis

PubMed Central

Khan, Mohammad Moshahid; Yang, Weng-Lang; Wang, Ping

2015-01-01

Sepsis is an enormous public health issue and the leading cause of death in critically ill patients in intensive care units (ICU). Overwhelming inflammation, characterized by cytokine storm, oxidative threats, and neutrophil sequestration is an underlying component of sepsis-associated organ failure. Despite recent advances in sepsis research, there is still no effective treatment available beyond the standard of care and supportive therapy. To reduce sepsis-related mortality, a better understanding of the biological mechanism associated with the sepsis is essential. Endoplasmic reticulum (ER), a subcellular organelle is responsible for the facilitation of protein folding and assembly and involved in several other physiological activities. Under the stress and inflammation condition, ER loses the homeostasis in its function, which is termed as ER stress. During ER stress, unfolded protein response (UPR) is activated to restore ER function to its normal balance. However, once the stress is beyond the compensatory capacity of UPR or protracted, the apoptosis would be initiated by triggering cell injuries, even to cell death. As such, ER stress and UPR are reported to be implicated in several pathological and inflammatory conditions. Although the detrimental role of ER stress during infections has been demonstrated, there is growing evidences that ER stress participate in the pathogenesis of sepsis. In this review, we summarize the current research in the context of ER stress and UPR signaling associated with sepsis and its related clinical conditions, such as trauma- hemorrhage, and ischemia/reperfusion (I/R) injury. We also discuss the potential implication of ER stress as a novel therapeutic target and prognostic marker in patients with sepsis. PMID:26125088

Endoplasmic reticulum degradation-enhancing α-mannosidase-like protein 1 targets misfolded HLA-B27 dimers for endoplasmic reticulum-associated degradation.

PubMed

Guiliano, David B; Fussell, Helen; Lenart, Izabela; Tsao, Edward; Nesbeth, Darren; Fletcher, Adam J; Campbell, Elaine C; Yousaf, Nasim; Williams, Sarah; Santos, Susana; Cameron, Amy; Towers, Greg J; Kellam, Paul; Hebert, Daniel N; Gould, Keith G; Powis, Simon J; Antoniou, Antony N

2014-11-01

HLA-B27 forms misfolded heavy chain dimers, which may predispose individuals to inflammatory arthritis by inducing endoplasmic reticulum (ER) stress and the unfolded protein response (UPR). This study was undertaken to define the role of the UPR-induced ER-associated degradation (ERAD) pathway in the disposal of HLA-B27 dimeric conformers. HeLa cell lines expressing only 2 copies of a carboxy-terminally Sv5-tagged HLA-B27 were generated. The ER stress-induced protein ER degradation-enhancing α-mannosidase-like protein 1 (EDEM1) was overexpressed by transfection, and dimer levels were monitored by immunoblotting. EDEM1, the UPR-associated transcription factor X-box binding protein 1 (XBP-1), the E3 ubiquitin ligase hydroxymethylglutaryl-coenzyme A reductase degradation 1 (HRD1), and the degradation-associated proteins derlin 1 and derlin 2 were inhibited using either short hairpin RNA or dominant-negative mutants. The UPR-associated ERAD of HLA-B27 was confirmed using ER stress-inducing pharamacologic agents in kinetic and pulse chase assays. We demonstrated that UPR-induced machinery can target HLA-B27 dimers and that dimer formation can be controlled by alterations to expression levels of components of the UPR-induced ERAD pathway. HLA-B27 dimers and misfolded major histocompatibility complex class I monomeric molecules bound to EDEM1 were detected, and overexpression of EDEM1 led to inhibition of HLA-B27 dimer formation. EDEM1 inhibition resulted in up-regulation of HLA-B27 dimers, while UPR-induced ERAD of dimers was prevented in the absence of EDEM1. HLA-B27 dimer formation was also enhanced in the absence of XBP-1, HRD1, and derlins 1 and 2. The present findings indicate that the UPR ERAD pathway can dispose of HLA-B27 dimers, thus presenting a potential novel therapeutic target for modulation of HLA-B27-associated inflammatory disease. Copyright © 2014 by the American College of Rheumatology.

Endoplasmic reticulum stress and eIF2α phosphorylation: The Achilles heel of pancreatic β cells.

PubMed

Cnop, Miriam; Toivonen, Sanna; Igoillo-Esteve, Mariana; Salpea, Paraskevi

2017-09-01

Pancreatic β cell dysfunction and death are central in the pathogenesis of most if not all forms of diabetes. Understanding the molecular mechanisms underlying β cell failure is important to develop β cell protective approaches. Here we review the role of endoplasmic reticulum stress and dysregulated endoplasmic reticulum stress signaling in β cell failure in monogenic and polygenic forms of diabetes. There is substantial evidence for the presence of endoplasmic reticulum stress in β cells in type 1 and type 2 diabetes. Direct evidence for the importance of this stress response is provided by an increasing number of monogenic forms of diabetes. In particular, mutations in the PERK branch of the unfolded protein response provide insight into its importance for human β cell function and survival. The knowledge gained from different rodent models is reviewed. More disease- and patient-relevant models, using human induced pluripotent stem cells differentiated into β cells, will further advance our understanding of pathogenic mechanisms. Finally, we review the therapeutic modulation of endoplasmic reticulum stress and signaling in β cells. Pancreatic β cells are sensitive to excessive endoplasmic reticulum stress and dysregulated eIF2α phosphorylation, as indicated by transcriptome data, monogenic forms of diabetes and pharmacological studies. This should be taken into consideration when devising new therapeutic approaches for diabetes.

Chronic Sleep Fragmentation During the Sleep Period Induces Hypothalamic Endoplasmic Reticulum Stress and PTP1b-Mediated Leptin Resistance in Male Mice

PubMed Central

Hakim, Fahed; Wang, Yang; Carreras, Alba; Hirotsu, Camila; Zhang, Jing; Peris, Eduard; Gozal, David

2015-01-01

Background: Sleep fragmentation (SF) is highly prevalent and may constitute an important contributing factor to excessive weight gain and the metabolic syndrome. Increased endoplasmic reticulum (ER) stress and activation of the unfolded protein response (UPR) leading to the attenuation of leptin receptor signaling in the hypothalamus leads to obesity and metabolic dysfunction. Methods: Mice were exposed to SF and sleep control (SC) for varying periods of time during which ingestive behaviors were monitored. UPR pathways and leptin receptor signaling were assessed in hypothalami. To further examine the mechanistic role of ER stress, changes in leptin receptor (ObR) signaling were also examined in wild-type mice treated with the ER chaperone tauroursodeoxycholic acid (TUDCA), as well as in CHOP −/+ transgenic mice. Results: Fragmented sleep in male mice induced increased food intake starting day 3 and thereafter, which was preceded by increases in ER stress and activation of all three UPR pathways in the hypothalamus. Although ObR expression was unchanged, signal transducer and activator of transcription 3 (STAT3) phosphorylation was decreased, suggesting reduced ObR signaling. Unchanged suppressor of cytokine signaling-3 (SOCS3) expression and increases in protein-tyrosine phosphatase 1B (PTP1B) expression and activity emerged with SF, along with reduced p-STAT3 responses to exogenous leptin. SF-induced effects were reversed following TUDCA treatment and were absent in CHOP −/+ mice. Conclusions: Sleep fragmentation (SF) induces hyperphagic behaviors and reduced leptin signaling in hypothalamus that are mediated by activation of endoplasmic reticulum (ER) stress, and ultimately lead to increased PTP1B activity. ER stress pathways are therefore potentially implicated in SF-induced weight gain and metabolic dysfunction, and may represent a viable therapeutic target. Citation: Hakim F, Wang Y, Carreras A, Hirotsu C, Zhang J, Peris E, Gozal D. Chronic sleep

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

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pan, Mu-Yun; Shen, Yuh-Chiang; National Research Institute of Chinese Medicine, Taipei, Taiwan

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 ERmore » 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

Preconditioning with Endoplasmic Reticulum Stress Ameliorates Endothelial Cell Inflammation

PubMed Central

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. PMID:25356743

Preconditioning with endoplasmic reticulum stress ameliorates endothelial cell inflammation.

PubMed

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.

Efficient IgM assembly and secretion require the plasma cell induced endoplasmic reticulum protein pERp1

PubMed Central

van Anken, Eelco; Pena, Florentina; Hafkemeijer, Nicole; Christis, Chantal; Romijn, Edwin P.; Grauschopf, Ulla; Oorschot, Viola M. J.; Pertel, Thomas; Engels, Sander; Ora, Ari; Lástun, Viorica; Glockshuber, Rudi; Klumperman, Judith; Heck, Albert J. R.; Luban, Jeremy; Braakman, Ineke

2009-01-01

Plasma cells daily secrete their own mass in antibodies, which fold and assemble in the endoplasmic reticulum (ER). To reach these levels, cells require pERp1, a novel lymphocyte-specific small ER-resident protein, which attains expression levels as high as BiP when B cells differentiate into plasma cells. Although pERp1 has no homology with known ER proteins, it does contain a CXXC motif typical for oxidoreductases. In steady state, the CXXC cysteines are locked by two parallel disulfide bonds with a downstream C(X)6C motif, and pERp1 displays only modest oxidoreductase activity. pERp1 emerged as a dedicated folding factor for IgM, associating with both heavy and light chains and promoting assembly and secretion of mature IgM. PMID:19805154

Fenofibrate Does Not Affect Burn-Induced Hepatic Endoplasmic Reticulum Stress

PubMed Central

Hiyama, Yaeko; Marshall, Alexandra H.; Kraft, Robert; Arno, Anna; Jeschke, Marc G.

2013-01-01

Background Burn injury causes major metabolic derangements such as hypermetabolism, hyperlipidemia, and insulin resistance and is associated with liver damage, hepatomegaly, and hepatic endoplasmic reticulum (ER) stress. Although the physiological consequences of such derangements have been delineated, the underlying molecular mechanisms remain unknown. Previously, it was shown that fenofibrate improves patient outcome by attenuating post-burn stress responses. Methods Fenofibrate, a peroxisome proliferator-activated receptor (PPAR)-α agonist, regulates liver lipid metabolism and has been used to treat hypertriglyceridemia and hypercholesterolemia for many years. The aim of the present study is to determine the effects of fenofibrate on burn-induced hepatic morphologic and metabolic changes. We randomized rats to sham, burn injury, and burn injury plus fenofibrate. Animals were sacrificed and livers were assessed at 24 or 48 hours post-burn. Results Burn injury decreased albumin and increased alanine transaminase (p = 0.1 vs. sham), indicating liver injury. Fenofibrate administration did not restore albumin or decrease alanine transaminase. In addition, ER stress was significantly increased after burn injury both with and without fenofibrate (p < 0.05 vs. sham). Burn injury increased fatty acid metabolism gene expression (p < 0.05 vs. sham), downstream of PPARα. Fenofibrate treatment increased fatty acid metabolism further, which reduced post-burn hepatic steatosis (burn vs. sham p < 0.05, burn+fenofibrate vs. sham not significant). Conclusions Fenofibrate did not alleviate thermal injury induced hepatic ER stress and dysfunction but reduced hepatic steatosis by modulating hepatic genes related to fat metabolism. PMID:23866789

Endoplasmic reticulum stress sensor protein kinase R-like endoplasmic reticulum kinase (PERK) protects against pressure overload-induced heart failure and lung remodeling.

PubMed

Liu, Xiaoyu; Kwak, Dongmin; Lu, Zhongbing; Xu, Xin; Fassett, John; Wang, Huan; Wei, Yidong; Cavener, Douglas R; Hu, Xinli; Hall, Jennifer; Bache, Robert J; Chen, Yingjie

2014-10-01

Studies have reported that development of congestive heart failure is associated with increased endoplasmic reticulum stress. Double stranded RNA-activated protein kinase R-like endoplasmic reticulum kinase (PERK) is a major transducer of the endoplasmic reticulum stress response and directly phosphorylates eukaryotic initiation factor 2α, resulting in translational attenuation. However, the physiological effect of PERK on congestive heart failure development is unknown. To study the effect of PERK on ventricular structure and function, we generated inducible cardiac-specific PERK knockout mice. Under unstressed conditions, cardiac PERK knockout had no effect on left ventricular mass, or its ratio to body weight, cardiomyocyte size, fibrosis, or left ventricular function. However, in response to chronic transverse aortic constriction, PERK knockout mice exhibited decreased ejection fraction, increased left ventricular fibrosis, enhanced cardiomyocyte apoptosis, and exacerbated lung remodeling in comparison with wild-type mice. PERK knockout also dramatically attenuated cardiac sarcoplasmic reticulum Ca(2+)-ATPase expression in response to aortic constriction. Our findings suggest that PERK is required to protect the heart from pressure overload-induced congestive heart failure. © 2014 American Heart Association, Inc.

Mizoribine corrects defective nephrin biogenesis by restoring intracellular energy balance.

PubMed

Nakajo, Aya; Khoshnoodi, Jamshid; Takenaka, Hitoshi; Hagiwara, Emi; Watanabe, Takashi; Kawakami, Hayato; Kurayama, Ryota; Sekine, Yuji; Bessho, Fumio; Takahashi, Shori; Swiatecka-Urban, Agnieszka; Tryggvason, Karl; Yan, Kunimasa

2007-09-01

Proteins are modified and folded within the endoplasmic reticulum (ER). When the influx of proteins exceeds the capacity of the ER to handle the load, the ER is "stressed" and protein biogenesis is affected. We have previously shown that the induction of ER stress by ATP depletion in podocytes leads to mislocalization of nephrin and subsequent injury of podocytes. The aim of the present study was to determine whether ER stress is associated with proteinuria in vivo and whether the immunosuppressant mizoribine may exert its antiproteinuric effect by restoring normal nephrin biogenesis. Induction of nephrotic-range proteinuria with puromycin aminonucleoside in mice increased expression of the ER stress marker GRP78 in podocytes, and led to the mislocalization of nephrin to the cytoplasm. In vitro, mizoribine, through a mechanism likely dependent on the inhibition of inosine 5'-monophosphate dehydrogenase (IMPDH) activity in podocytes, restored the intracellular energy balance by increasing levels of ATP and corrected the posttranslational processing of nephrin. Therefore, we speculate that mizoribine may induce remission of proteinuria, at least in part, by restoring the biogenesis of slit diaphragm proteins in injured podocytes. Further understanding of the ER microenvironment may lead to novel approaches to treat diseases in which abnormal handling of proteins plays a role in pathogenesis.

Influence of resveratrol on endoplasmic reticulum stress and expression of adipokines in adipose tissues/adipocytes induced by high-calorie diet or palmitic acid.

PubMed

Chen, Li; Wang, Ting; Chen, Guanjun; Wang, Nuojin; Gui, Li; Dai, Fang; Fang, Zhaohui; Zhang, Qiu; Lu, Yunxia

2017-03-01

This study aimed to determine whether resveratrol treatment alleviates endoplasmic reticulum stress and changes the expression of adipokines in adipose tissues and cells. 8-week-old male C57BL/6 mice were fed a high-calorie diet (HCD group) or high-calorie diet supplemented with resveratrol (high-calorie diet  + resveratrol group) for 3 months. Insulin resistance, serum lipids and proinflammatory indices, the size and inflammatory cell infiltration in subcutaneous and visceral adipose tissues were analyzed. The gene expressions of endoplasmic reticulum stress, adipokines, and inflammatory cytokines were determined. The induced mature 3T3-L1 cells were pretreated with resveratrol and then palmitic acid, and the gene expressions of endoplasmic reticulum stress, adipokines, and inflammatory cytokines were determined. Subcutaneous and visceral adipose tissues in the high-calorie diet-fed mice exhibited adipocyte hypertrophy, inflammatory activation, and endoplasmic reticulum stress. Resveratrol alleviated high-calorie diet-induced insulin resistance and endoplasmic reticulum stress, increased expression of SIRT1, and reversed expression of adipokines in varying degrees in both subcutaneous and visceral adipose tissues. The effects of resveratrol on palmitic acid-treated adipocytes were similar to those shown in the tissues. Resveratrol treatment obviously reversed adipocyte hypertrophy and insulin resistance by attenuating endoplasmic reticulum stress and inflammation, thus increasing the expression of SIRT1 and inverting the expression of adipokines in vivo and in vitro.

The shape of things to come: regulation of shape changes in endoplasmic reticulum.

PubMed

Paiement, J; Bergeron, J

2001-01-01

Shape changes in the endoplasmic reticulum control fundamental cell processes including nuclear envelope assembly in mitotic cells, calcium homeostasis in cytoplasmic domains of secreting and motile cells, and membrane traffic in the early secretion apparatus between the endoplasmic reticulum and Golgi. Opposing forces of assembly (membrane fusion) and disassembly (membrane fragmentation) ultimately determine the size and shape of this organelle. This review examines some of the regulatory mechanisms involved in these processes and how they occur at specific sites or subcompartments of the endoplasmic reticulum.

UDP-glucose:glycoprotein glucosyltransferase (UGGT1) promotes substrate solubility in the endoplasmic reticulum

PubMed Central

Ferris, Sean P.; Jaber, Nikita S.; Molinari, Maurizio; Arvan, Peter; Kaufman, Randal J.

2013-01-01

Protein folding in the endoplasmic reticulum (ER) is error prone, and ER quality control (ERQC) processes ensure that only correctly folded proteins are exported from the ER. Glycoproteins can be retained in the ER by ERQC, and this retention contributes to multiple human diseases, termed ER storage diseases. UDP-glucose:glycoprotein glucosyltransferase (UGGT1) acts as a central component of glycoprotein ERQC, monoglucosylating deglucosylated N-glycans of incompletely folded glycoproteins and promoting subsequent reassociation with the lectin-like chaperones calreticulin and calnexin. The extent to which UGGT1 influences glycoprotein folding, however, has only been investigated for a few selected substrates. Using mouse embryonic fibroblasts lacking UGGT1 or those with UGGT1 complementation, we investigated the effect of monoglucosylation on the soluble/insoluble distribution of two misfolded α1-antitrypsin (AAT) variants responsible for AAT deficiency disease: null Hong Kong (NHK) and Z allele. Whereas substrate solubility increases directly with the number of N-linked glycosylation sites, our results indicate that additional solubility is conferred by UGGT1 enzymatic activity. Monoglucosylation-dependent solubility decreases both BiP association with NHK and unfolded protein response activation, and the solubility increase is blocked in cells deficient for calreticulin. These results suggest that UGGT1-dependent monoglucosylation of N-linked glycoproteins promotes substrate solubility in the ER. PMID:23864712

Endoplasmic Reticulum Stress and Oxidative Stress: A Vicious Nexus Implicated in Bowel Disease Pathophysiology

PubMed Central

Chong, Wai Chin; Shastri, Madhur D.; Eri, Rajaraman

2017-01-01

The endoplasmic reticulum (ER) is a complex protein folding and trafficking organelle. Alteration and discrepancy in the endoplasmic reticulum environment can affect the protein folding process and hence, can result in the production of misfolded proteins. The accumulation of misfolded proteins causes cellular damage and elicits endoplasmic reticulum stress. Under such stress conditions, cells exhibit reduced functional synthesis, and will undergo apoptosis if the stress is prolonged. To resolve the ER stress, cells trigger an intrinsic mechanism called an unfolded protein response (UPR). UPR is an adaptive signaling process that triggers multiple pathways through the endoplasmic reticulum transmembrane transducers, to reduce and remove misfolded proteins and improve the protein folding mechanism, in order to improve and maintain endoplasmic reticulum homeostasis. An increasing number of studies support the view that oxidative stress has a strong connection with ER stress. During the protein folding process, reactive oxygen species are produced as by-products, leading to impaired reduction-oxidation (redox) balance conferring oxidative stress. As the protein folding process is dependent on redox homeostasis, the oxidative stress can disrupt the protein folding mechanism and enhance the production of misfolded proteins, causing further ER stress. It is proposed that endoplasmic reticulum stress and oxidative stress together play significant roles in the pathophysiology of bowel diseases. PMID:28379196

Endoplasmic Reticulum Stress and Oxidative Stress: A Vicious Nexus Implicated in Bowel Disease Pathophysiology.

PubMed

Chong, Wai Chin; Shastri, Madhur D; Eri, Rajaraman

2017-04-05

The endoplasmic reticulum (ER) is a complex protein folding and trafficking organelle. Alteration and discrepancy in the endoplasmic reticulum environment can affect the protein folding process and hence, can result in the production of misfolded proteins. The accumulation of misfolded proteins causes cellular damage and elicits endoplasmic reticulum stress. Under such stress conditions, cells exhibit reduced functional synthesis, and will undergo apoptosis if the stress is prolonged. To resolve the ER stress, cells trigger an intrinsic mechanism called an unfolded protein response (UPR). UPR is an adaptive signaling process that triggers multiple pathways through the endoplasmic reticulum transmembrane transducers, to reduce and remove misfolded proteins and improve the protein folding mechanism, in order to improve and maintain endoplasmic reticulum homeostasis. An increasing number of studies support the view that oxidative stress has a strong connection with ER stress. During the protein folding process, reactive oxygen species are produced as by-products, leading to impaired reduction-oxidation (redox) balance conferring oxidative stress. As the protein folding process is dependent on redox homeostasis, the oxidative stress can disrupt the protein folding mechanism and enhance the production of misfolded proteins, causing further ER stress. It is proposed that endoplasmic reticulum stress and oxidative stress together play significant roles in the pathophysiology of bowel diseases.

Ero1-α and PDIs constitute a hierarchical electron transfer network of endoplasmic reticulum oxidoreductases

PubMed Central

Araki, Kazutaka; Iemura, Shun-ichiro; Kamiya, Yukiko; Ron, David; Kato, Koichi; Natsume, Tohru

2013-01-01

Ero1-α and endoplasmic reticulum (ER) oxidoreductases of the protein disulfide isomerase (PDI) family promote the efficient introduction of disulfide bonds into nascent polypeptides in the ER. However, the hierarchy of electron transfer among these oxidoreductases is poorly understood. In this paper, Ero1-α–associated oxidoreductases were identified by proteomic analysis and further confirmed by surface plasmon resonance. Ero1-α and PDI were found to constitute a regulatory hub, whereby PDI induced conformational flexibility in an Ero1-α shuttle cysteine (Cys99) facilitated intramolecular electron transfer to the active site. In isolation, Ero1-α also oxidized ERp46, ERp57, and P5; however, kinetic measurements and redox equilibrium analysis revealed that PDI preferentially oxidized other oxidoreductases. PDI accepted electrons from the other oxidoreductases via its a′ domain, bypassing the a domain, which serves as the electron acceptor from reduced glutathione. These observations provide an integrated picture of the hierarchy of cooperative redox interactions among ER oxidoreductases in mammalian cells. PMID:24043701

Endoplasmic Reticulum Stress Mediates Methamphetamine-Induced Blood–Brain Barrier Damage

PubMed Central

Qie, Xiaojuan; Wen, Di; Guo, Hongyan; Xu, Guanjie; Liu, Shuai; Shen, Qianchao; Liu, Yi; Zhang, Wenfang; Cong, Bin; Ma, Chunling

2017-01-01

Methamphetamine (METH) abuse causes serious health problems worldwide, and long-term use of METH disrupts the blood–brain barrier (BBB). Herein, we explored the potential mechanism of endoplasmic reticulum (ER) stress in METH-induced BBB endothelial cell damage in vitro and the therapeutic potential of endoplasmic reticulum stress inhibitors for METH-induced BBB disruption in C57BL/6J mice. Exposure of immortalized BMVEC (bEnd.3) cells to METH significantly decreased cell viability, induced apoptosis, and diminished the tightness of cell monolayers. METH activated ER stress sensor proteins, including PERK, ATF6, and IRE1, and upregulated the pro-apoptotic protein CHOP. The ER stress inhibitors significantly blocked the upregulation of CHOP. Knockdown of CHOP protected bEnd.3 cells from METH-induced cytotoxicity. Furthermore, METH elevated the production of reactive oxygen species (ROS) and induced the dysfunction of mitochondrial characterized by a Bcl2/Bax ratio decrease, mitochondrial membrane potential collapse, and cytochrome c. ER stress release was partially reversed by ROS inhibition, and cytochrome c release was partially blocked by knockdown of CHOP. Finally, PBA significantly attenuated METH-induced sodium fluorescein (NaFluo) and Evans Blue leakage, as well as tight junction protein loss, in C57BL/6J mice. These data suggest that BBB endothelial cell damage was caused by METH-induced endoplasmic reticulum stress, which further induced mitochondrial dysfunction, and that PBA was an effective treatment for METH-induced BBB disruption. PMID:28959203

Sirtuin1 protects endothelial Caveolin-1 expression and preserves endothelial function via suppressing miR-204 and endoplasmic reticulum stress.

PubMed

Kassan, M; Vikram, A; Kim, Y R; Li, Q; Kassan, A; Patel, H H; Kumar, S; Gabani, M; Liu, J; Jacobs, J S; Irani, K

2017-02-09

Sirtuin1 (Sirt1) is a class III histone deacetylase that regulates a variety of physiological processes, including endothelial function. Caveolin1 (Cav1) is also an important determinant of endothelial function. We asked if Sirt1 governs endothelial Cav1 and endothelial function by regulating miR-204 expression and endoplasmic reticulum (ER) stress. Knockdown of Sirt1 in endothelial cells, and in vivo deletion of endothelial Sirt1, induced endothelial ER stress and miR-204 expression, reduced Cav1, and impaired endothelium-dependent vasorelaxation. All of these effects were reversed by a miR-204 inhibitor (miR-204 I) or with overexpression of Cav1. A miR-204 mimic (miR-204 M) decreased Cav1 in endothelial cells. In addition, high-fat diet (HFD) feeding induced vascular miR-204 and reduced endothelial Cav1. MiR-204-I protected against HFD-induced downregulation of endothelial Cav1. Moreover, pharmacologic induction of ER stress with tunicamycin downregulated endothelial Cav1 and impaired endothelium-dependent vasorelaxation that was rescued by overexpressing Cav1. In conclusion, Sirt1 preserves Cav1-dependent endothelial function by mitigating miR-204-mediated vascular ER stress.

Endoplasmic reticulum stress in chondrodysplasias caused by mutations in collagen types II and X.

PubMed

Gawron, Katarzyna

2016-11-01

The endoplasmic reticulum is primarily recognized as the site of synthesis and folding of secreted, membrane-bound, and some organelle-targeted proteins. An imbalance between the load of unfolded proteins and the processing capacity in endoplasmic reticulum leads to the accumulation of unfolded or misfolded proteins and endoplasmic reticulum stress, which is a hallmark of a number of storage diseases, including neurodegenerative diseases, a number of metabolic diseases, and cancer. Moreover, its contribution as a novel mechanistic paradigm in genetic skeletal diseases associated with abnormalities of the growth plates and dwarfism is considered. In this review, I discuss the mechanistic significance of endoplasmic reticulum stress, abnormal folding, and intracellular retention of mutant collagen types II and X in certain variants of skeletal chondrodysplasia.

Propofol restores TRPV1 sensitivity via a TRPA1-, nitric oxide synthase-dependent activation of PKCε

PubMed Central

Sinharoy, Pritam; Zhang, Hongyu; Sinha, Sayantani; Prudner, Bethany C; Bratz, Ian N; Damron, Derek S

2015-01-01

We previously demonstrated that the intravenous anesthetic, propofol, restores the sensitivity of transient receptor potential vanilloid channel subtype-1 (TRPV1) receptors via a protein kinase C epsilon (PKCε)-dependent and transient receptor potential ankyrin channel subtype-1 (TRPA1)-dependent pathway in sensory neurons. The extent to which the two pathways are directly linked or operating in parallel has not been determined. Using a molecular approach, our objectives of the current study were to confirm that TRPA1 activation directly results in PKCε activation and to elucidate the cellular mechanism by which this occurs. F-11 cells were transfected with complimentary DNA (cDNA) for TRPV1 only or both TRPV1 and TRPA1. Intracellular Ca2+ concentration was measured in individual cells via fluorescence microscopy. An immunoblot analysis of the total and phosphorylated forms of PKCε, nitric oxide synthase (nNOS), and TRPV1 was also performed. In F-11 cells containing both channels, PKCε inhibition prevented the propofol- and allyl isothiocyanate (AITC)-induced restoration of TRPV1 sensitivity to agonist stimulation as well as increased phosphorylation of PKCε and TRPV1. In cells containing TRPV1 only, neither agonist induced PKCε or TRPV1 phosphorylation. Moreover, NOS inhibition blocked propofol-and AITC-induced restoration of TRPV1 sensitivity and PKCε phosphorylation, and PKCε inhibition prevented the nitric oxide donor, SNAP, from restoring TRPV1 sensitivity. Also, propofol-and AITC-induced phosphorylation of nNOS and nitric oxide (NO) production were blocked with the TRPA1-antagonist, HC-030031. These data indicate that the AITC- and propofol-induced restoration of TRPV1 sensitivity is mediated by a TRPA1-dependent, nitric oxide synthase-dependent activation of PKCε. PMID:26171233

Functional Characterization of Ice Plant SKD1, an AAA-Type ATPase Associated with the Endoplasmic Reticulum-Golgi Network, and Its Role in Adaptation to Salt Stress1[W

PubMed Central

Jou, Yingtzy; Chiang, Chih-Pin; Jauh, Guang-Yuh; Yen, Hungchen Emilie

2006-01-01

A salt-induced gene mcSKD1 (suppressor of K+ transport growth defect) able to facilitate K+ uptake has previously been identified from the halophyte ice plant (Mesembryanthemum crystallinum). The sequence of mcSKD1 is homologous to vacuolar protein sorting 4, an ATPase associated with a variety of cellular activities-type ATPase that participates in the sorting of vacuolar proteins into multivesicular bodies in yeast (Saccharomyces cerevisiae). Recombinant mcSKD1 exhibited ATP hydrolytic activities in vitro with a half-maximal rate at an ATP concentration of 1.25 mm. Point mutations on active site residues abolished its ATPase activity. ADP is both a product and a strong inhibitor of the reaction. ADP-binding form of mcSDK1 greatly reduced its catalytic activity. The mcSKD1 protein accumulated ubiquitously in both vegetative and reproductive parts of plants. Highest accumulation was observed in cells actively engaging in the secretory processes, such as bladder cells of leaf epidermis. Membrane fractionation and double-labeling immunofluorescence showed the predominant localization of mcSKD1 in the endoplasmic reticulum-Golgi network. Immunoelectron microscopy identified the formation of mcSKD1 proteins into small aggregates in the cytosol and associated with membrane continuum within the endomembrane compartments. These results indicated that this ATPase participates in the endoplasmic reticulum-Golgi mediated protein sorting machinery for both housekeeping function and compartmentalization of excess Na+ under high salinity. PMID:16581876

Endoplasmic Reticulum Transport of Glutathione by Sec61 Is Regulated by Ero1 and Bip.

PubMed

Ponsero, Alise J; Igbaria, Aeid; Darch, Maxwell A; Miled, Samia; Outten, Caryn E; Winther, Jakob R; Palais, Gael; D'Autréaux, Benoit; Delaunay-Moisan, Agnès; Toledano, Michel B

2017-09-21

In the endoplasmic reticulum (ER), Ero1 catalyzes disulfide bond formation and promotes glutathione (GSH) oxidation to GSSG. Since GSSG cannot be reduced in the ER, maintenance of the ER glutathione redox state and levels likely depends on ER glutathione import and GSSG export. We used quantitative GSH and GSSG biosensors to monitor glutathione import into the ER of yeast cells. We found that glutathione enters the ER by facilitated diffusion through the Sec61 protein-conducting channel, while oxidized Bip (Kar2) inhibits transport. Increased ER glutathione import triggers H 2 O 2 -dependent Bip oxidation through Ero1 reductive activation, which inhibits glutathione import in a negative regulatory loop. During ER stress, transport is activated by UPR-dependent Ero1 induction, and cytosolic glutathione levels increase. Thus, the ER redox poise is tuned by reciprocal control of glutathione import and Ero1 activation. The ER protein-conducting channel is permeable to small molecules, provided the driving force of a concentration gradient. Copyright © 2017 Elsevier Inc. All rights reserved.

Following the movement of Cu ions in a SSZ-13 zeolite during dehydration, reduction and adsorption: a combined in situ TP-XRD, XANES/DRIFTS study

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kwak, Ja Hun; Varga, Tamas; Peden, Charles HF

2014-05-05

Cu-SSZ-13 has been shown to possess high activity and superior N2 formation selectivity in the selective catalytic reduction of NOx under oxygen rich conditions. Here, a combination of synchrotron-based (XRD and XANES) and vibrational (DRIFTS) spectroscopy tools have been used to follow the changes in the location and coordination environment of copper ions in a Cu-SSZ-13 zeolite during calcinations, reduction with CO, and adsorption of CO and H2O. XANES spectra collected during these procedures provides critical information not only on the variation in the oxidation state of the copper species in the zeolite structure, but also on the changes inmore » the coordination environment around these ions as they interact with the framework, and with different adsorbates (H2O and CO). Time-resolved XRD data indicate the movement of copper ions and the consequent variation of the unit cell parameters during dehydration. DRIFT spectra provide information about the adsorbed species present in the zeolite, as well as the oxidation states of and coordination environment around the copper ions. A careful analysis of the asymmetric T-O-T vibrations of the CHA framework perturbed by copper ions in different coordination environments proved to be especially informative. The results of this study will aid the identification of the location, coordination and oxidation states of copper ions obtained during in operando catalytic studies. Financial support was provided by the US Department of Energy (DOE), Office of Energy Efficiency and Renewable Energy, Vehicle Technologies Program. Part of this work (sample preparation) was performed in the Environmental Molecular Sciences Laboratory (EMSL) at Pacific Northwest National Laboratory (PNNL). The EMSL is a national scientific user facility supported by the US DOE, Office of Biological and Environmental Research. PNNL is a multi-program national laboratory operated for the US DOE by Battelle. All of the spectroscopy work reported here

15 CFR 990.53 - Restoration selection-developing restoration alternatives.

Code of Federal Regulations, 2011 CFR

2011-01-01

... 15 Commerce and Foreign Trade 3 2011-01-01 2011-01-01 false Restoration selection-developing... OIL POLLUTION ACT REGULATIONS NATURAL RESOURCE DAMAGE ASSESSMENTS Restoration Planning Phase § 990.53 Restoration selection—developing restoration alternatives. (a) General. (1) If the information on injury...

15 CFR 990.53 - Restoration selection-developing restoration alternatives.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 15 Commerce and Foreign Trade 3 2010-01-01 2010-01-01 false Restoration selection-developing... OIL POLLUTION ACT REGULATIONS NATURAL RESOURCE DAMAGE ASSESSMENTS Restoration Planning Phase § 990.53 Restoration selection—developing restoration alternatives. (a) General. (1) If the information on injury...

Loss of Oca2 disrupts the unfolded protein response and increases resistance to endoplasmic reticulum stress in melanocytes

PubMed Central

Cheng, Tsing; Orlow, Seth J.; Manga, Prashiela

2013-01-01

Summary Accumulation of proteins in the endoplasmic reticulum (ER) typically induces stress and initiates the unfolded protein response (UPR) to facilitate recovery. If homeostasis is not restored, apoptosis is induced. However, adaptation to chronic UPR activation can increase resistance to subsequent acute ER stress. We therefore investigated adaptive mechanisms in Oculocutaneous albinism type 2 (Oca2)-null melanocytes where UPR signaling is arrested despite continued tyrosinase accumulation leading to resistance to the chemical ER stressor thapsigargin. Although thapsigargin triggers UPR activation, instead of Perk-mediated phosphorylation of eIF2α, in Oca2-null melanocytes, eIF2α was rapidly dephosphorylated upon treatment. Dephosphorylation was mediated by the Gadd34-PP1α phosphatase complex. Gadd34-complex inhibition blocked eIF2α dephosphorylation and significantly increased Oca2-null melanocyte sensitivity to thapsigargin. Thus, Oca2-null melanocytes adapt to acute ER stress by disruption of proapoptotic Perk signaling, which promotes cell survival. This is the first study to demonstrate rapid eIF2α dephosphorylation as an adaptive mechanism to ER stress. PMID:23962237

Ibutilide protects against cardiomyocytes injury via inhibiting endoplasmic reticulum and mitochondrial stress pathways.

PubMed

Wang, Yu; Wang, Yi-Li; Huang, Xia; Yang, Yang; Zhao, Ya-Jun; Wei, Cheng-Xi; Zhao, Ming

2017-02-01

Atrial fibrillation (AF) is a complex disease with multiple inter-relating causes culminating in rapid atrial activation and atrial structural remodeling. The contribution of endoplasmic reticulum and mitochondria stress to AF has been highlighted. As the class III antiarrhythmic agent, ibutilide are widely used to AF. This study was designed to explore whether ibutilide could treat AF by inhibiting endoplasmic reticulum stress pathways and mitochondria stress. The neonatal rat cardiomyocytes were isolated and exposed to H 2 O 2 , ibutilide was add to the culture medium 12 h. Then the cell viability, oxidative stress levels and apoptotic rate were analyzed. In addition, endoplasmic reticulum stress related protein (GRP78, GRP94, CHOP), mitochondria-dependent protein (Bax, Bcl-2) and caspase-3/9/12 were identified by real-time PCR and western blot analysis. In our results, remarkable decreased cell viability and oxidative stress levels were detected in cardiomyocytes after treating with H 2 O 2 . The apoptotic rate and the expression of proteins involved in mitochondrial stress and endoplasmic reticulum stress pathways increased. While ibutilide significantly inhibited these changes. These data suggested that ibutilide serves a protective role against H 2 O 2 -induced apoptosis of neonatal rat cardiomyocytes, and the mechanism is related to suppression of mitochondrial stress and endoplasmic reticulum stress.

Endoplasmic Reticulum-Plasma Membrane Contacts Regulate Cellular Excitability.

PubMed

Dickson, Eamonn J

2017-01-01

Cells that have intrinsic electrical excitability utilize changes in membrane potential to communicate with neighboring cells and initiate cellular cascades. Excitable cells like neurons and myocytes have evolved highly specialized subcellular architectures to translate these electrical signals into cellular events. One such structural specialization is sarco-/endoplasmic reticulum-plasma membrane contact sites. These membrane contact sites are positioned by specific membrane-membrane tethering proteins and contain an ever-expanding list of additional proteins that organize information transfer across the junctional space (~ 15-25 nm distance) to shape membrane identity and control cellular excitability. In this chapter we discuss how contacts between the sarco-/endoplasmic reticulum and plasma membrane are essential for regulated excitation-contraction coupling in striated muscle and control of lipid-dependent ion channels.

Adenovirus Modulates Toll-Like Receptor 4 Signaling by Reprogramming ORP1L-VAP Protein Contacts for Cholesterol Transport from Endosomes to the Endoplasmic Reticulum.

PubMed

Cianciola, Nicholas L; Chung, Stacey; Manor, Danny; Carlin, Cathleen R

2017-03-15

Human adenoviruses (Ads) generally cause mild self-limiting infections but can lead to serious disease and even be fatal in high-risk individuals, underscoring the importance of understanding how the virus counteracts host defense mechanisms. This study had two goals. First, we wished to determine the molecular basis of cholesterol homeostatic responses induced by the early region 3 membrane protein RIDα via its direct interaction with the sterol-binding protein ORP1L, a member of the evolutionarily conserved family of oxysterol-binding protein (OSBP)-related proteins (ORPs). Second, we wished to determine how this interaction regulates innate immunity to adenovirus. ORP1L is known to form highly dynamic contacts with endoplasmic reticulum-resident VAP proteins that regulate late endosome function under regulation of Rab7-GTP. Our studies have demonstrated that ORP1L-VAP complexes also support transport of LDL-derived cholesterol from endosomes to the endoplasmic reticulum, where it was converted to cholesteryl esters stored in lipid droplets when ORP1L was bound to RIDα. The virally induced mechanism counteracted defects in the predominant cholesterol transport pathway regulated by the late endosomal membrane protein Niemann-Pick disease type C protein 1 (NPC1) arising during early stages of viral infection. However, unlike NPC1, RIDα did not reconstitute transport to endoplasmic reticulum pools that regulate SREBP transcription factors. RIDα-induced lipid trafficking also attenuated proinflammatory signaling by Toll-like receptor 4, which has a central role in Ad pathogenesis and is known to be tightly regulated by cholesterol-rich "lipid rafts." Collectively, these data show that RIDα utilizes ORP1L in a way that is distinct from its normal function in uninfected cells to fine-tune lipid raft cholesterol that regulates innate immunity to adenovirus in endosomes. IMPORTANCE Early region 3 proteins encoded by human adenoviruses that attenuate immune

Photodegradation of sulfasalazine and its human metabolites in water by UV and UV/peroxydisulfate processes.

PubMed

Ji, Yuefei; Yang, Yan; Zhou, Lei; Wang, Lu; Lu, Junhe; Ferronato, Corinne; Chovelon, Jean-Marc

2018-04-15

The widespread occurrence of pharmaceuticals and their metabolites in natural waters has raised great concerns about their potential risks on human health and ecological systems. This study systematically investigates the degradation of sulfasalazine (SSZ) and its two human metabolites, sulfapyridine (SPD) and 5-aminosalicylic acid (5-ASA), by UV and UV/peroxydisulfate (UV/PDS) processes. Experimental results show that SPD and 5-ASA were readily degraded upon UV 254 nm direct photolysis, with quantum yields measured to be (8.6 ± 0.8) × 10 -3 and (2.4 ± 0.1) × 10 -2  mol Einstein -1 , respectively. Although SSZ was resistant to direct UV photolysis, it could be effectively removed by both UV/H 2 O 2 and UV/PDS processes, with fluence-based pseudo-first-order rate constants determined to be 0.0030 and 0.0038 cm 2  mJ -1 , respectively. Second-order rate constant between SO 4 •- and SSZ was measured as (1.33 ± 0.01) × 10 9  M -1 s -1 by competition kinetic method. A kinetic model was established for predicting the degradation rate of SSZ in the UV/PDS process. Increasing the dosage of PDS significantly enhanced the degradation of SSZ in the UV/PDS process, which can be well predicted by the developed kinetic model. Natural water constituents, such as natural organic matter (NOM) and bicarbonate (HCO 3 - ), influenced the degradation of SSZ differently. The azo functional group of SSZ molecule was predicted as the reactive site susceptible to electrophilic attack by SO 4 •- by frontier electron densities (FEDs) calculations. Four intermediate products arising from azo bond cleavage and SO 2 extrusion were identified by solid phase extraction-liquid chromatography-triple quadrupole mass spectrometry (SPE-LC-MS/MS). Based on the products identified, detailed transformation pathways for SSZ degradation in the UV/PDS system were proposed. Results reveal that UV/PDS could be an efficient approach for remediation of water

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

PubMed

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.

Mangiferin inhibits endoplasmic reticulum stress-associated thioredoxin-interacting protein/NLRP3 inflammasome activation with regulation of AMPK in endothelial cells.

PubMed

Song, Junna; Li, Jia; Hou, Fangjie; Wang, Xiaona; Liu, Baolin

2015-03-01

Endothelial dysfunction is tightly associated with cardiovascular complications in diabetic patients. This study aims to investigate the effects of mangiferin on the regulation of endothelial homeostasis under endoplasmic reticulum stress (ER stress) conditions. High glucose (25 mmol/L) exposure induced ER stress and promoted ROS production in endothelial cells. Mangiferin effectively inhibited ER stress-associated oxidative stress by attenuating IRE1α phosphorylation and reducing ROS production. In response to ER stress, thioredoxin-interacting protein (TXNIP) expression increased, followed by NLRP3 inflammasome activation and increased IL-1β secretion. Mangiferin treatment attenuated the expressions of TXNIP and NLRP3 and reduced IL-1β and IL-6 production, demonstrating its inhibitory effects on TXNIP/NLRP3 inflammasome activation. NLRP3 inflammasome activation is responsible for mitochondrial cell death. Mangiferin restored the loss of the mitochondrial membrane potential (Δψm) and inhibited caspase-3 activity, and thereby protected cells from high glucose-induced apoptosis. Moreover, mangiferin inhibited ET-1 secretion and restored the loss of NO production when cells were exposed to high glucose. Mangiferin enhanced AMPK phosphorylation and AMPK inhibitor compound C diminished its beneficial effects, indicating the potential role of AMPK in its action. Our work showed the beneficial effects of mangiferin on the improvement of endothelial homeostasis and elucidated the molecular pathway through which mangiferin ameliorated endothelial dysfunction by inhibition of ER stress-associated TXNIP/NLRP3 inflammasome activation in endothelial cells. These findings demonstrated the beneficial effects of mangiferin on the regulation of endothelial homeostasis and indicated its potential application in the management of diabetic cardiovascular complications. Copyright © 2015 Elsevier Inc. All rights reserved.

26 CFR 1.412(c)(1)-3T - Applying the minimum funding requirements to restored plans (temporary).

Code of Federal Regulations, 2010 CFR

2010-04-01

... (temporary). (a) In general—(1) Restoration method. The restoration method is a funding method that adapts... spread gain method that maintains an unfunded liability. A plan may adopt any cost method that satisfies...

37 CFR 1.452 - Restoration of right of priority.

Code of Federal Regulations, 2010 CFR

2010-07-01

... priority. 1.452 Section 1.452 Patents, Trademarks, and Copyrights UNITED STATES PATENT AND TRADEMARK OFFICE, DEPARTMENT OF COMMERCE GENERAL RULES OF PRACTICE IN PATENT CASES International Processing Provisions Priority § 1.452 Restoration of right of priority. (a) If the international application has an international...

From vein precipitates to deformation and fluid rock interaction within a SSZ: Insights from the Izu-Bonin-Mariana fore arc

NASA Astrophysics Data System (ADS)

Micheuz, Peter; Quandt, Dennis; Kurz, Walter

2017-04-01

International Ocean Discovery Program (IODP) expeditions 352 and 351 drilled through oceanic crust of the Philippine Sea plate. The two study areas are located near the outer Izu-Bonin-Mariana (IBM) fore arc and in the Amami Sankaku Basin. The primary objective was to improve our understanding of supra-subduction zones (SSZ) and the process of subduction initiation. The recovered drill cores during IODP expedition 352 represent approximately 50 Ma old fore arc basalts (FAB) and boninites revealing an entire volcanic sequence of a SSZ. Expedition 351 drilled FAB like oceanic crust similar in age to the FABs of expedition 352. In this study we present data on vein microstructures, geochemical data and isotopic signatures of vein precipitates to give new insights into fluid flow and precipitation processes and deformation within the Izu-Bonin fore arc. Veins formed predominantly as a consequence of hydrofracturing resulting in the occurrence of branched vein systems and brecciated samples. Along these hydrofractures the amount of altered host rock fragments varies and locally alters the host rock completely to zeolites and carbonates. Subordinately extensional veins released after the formation of the host rocks. Cross-cutting relationships of different vein types point to multiple fracturing events subsequently filled with minerals originating from a fluid with isotopic seawater signature. Based on vein precipitates, their morphology and their growth patterns four vein types have been defined. Major vein components are (Mg-) calcite and various zeolites determined by Raman spectra and electron microprobe analyses. Zeolites result from alteration of volcanic glass during interaction with a seawaterlike fluid. Type I veins which are characterized by micritic infill represent neptunian dykes. They predominantly occur in the upper levels of drill cores being the result of an initial volume change subsequently to crystallization of the host rocks. Type II veins are

Endoplasmic Reticulum Stress and Obesity.

PubMed

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.

Coordination of Endoplasmic Reticulum (ER) Signaling During Maize Seed Development

DOE Office of Scientific and Technical Information (OSTI.GOV)

Boston, Rebecca S.

2010-11-20

Seed storage reserves represent one of the most important sources of renewable fixed carbon and nitrogen found in nature. Seeds are well-adapted for diverting metabolic resources to synthesize storage proteins as well as enzymes and structural proteins needed for their transport and packaging into membrane bound storage protein bodies. Our underlying hypothesis is that the endoplasmic reticulum (ER) stress response provides the critical cellular control of metabolic flux required for optimal accumulation of storage reserves in seeds. This highly conserved response is a cellular mechanism to monitor the protein folding environment of the ER and restore homeostasis in the presencemore » of unfolded or misfolded proteins. In seeds, deposition of storage proteins in protein bodies is a highly specialized process that takes place even in the presence of mutant proteins that no longer fold and package properly. The capacity of the ER to deposit these aberrant proteins in protein bodies during a period that extends several weeks provides an excellent model for deconvoluting the ER stress response of plants. We have focused in this project on the means by which the ER senses and responds to functional perturbations and the underlying intracellular communication that occurs among biosynthetic, trafficking and degradative pathways for proteins during seed development.« less

Single-molecule analysis of diffusion and trapping of STIM1 and Orai1 at endoplasmic reticulum–plasma membrane junctions

PubMed Central

Wu, Minnie M.; Covington, Elizabeth D.; Lewis, Richard S.

2014-01-01

Following endoplasmic reticulum (ER) Ca2+ depletion, STIM1 and Orai1 complexes assemble autonomously at ER–plasma membrane (PM) junctions to trigger store-operated Ca2+ influx. One hypothesis to explain this process is a diffusion trap in which activated STIM1 diffusing in the ER becomes trapped at junctions through interactions with the PM, and STIM1 then traps Orai1 in the PM through binding of its calcium release-activated calcium activation domain. We tested this model by analyzing STIM1 and Orai1 diffusion using single-particle tracking, photoactivation of protein ensembles, and Monte Carlo simulations. In resting cells, STIM1 diffusion is Brownian, while Orai1 is slightly subdiffusive. After store depletion, both proteins slow to the same speeds, consistent with complex formation, and are confined to a corral similar in size to ER–PM junctions. While the escape probability at high STIM:Orai expression ratios is <1%, it is significantly increased by reducing the affinity of STIM1 for Orai1 or by expressing the two proteins at comparable levels. Our results provide direct evidence that STIM-Orai complexes are trapped by their physical connections across the junctional gap, but also reveal that the complexes are surprisingly dynamic, suggesting that readily reversible binding reactions generate free STIM1 and Orai1, which engage in constant diffusional exchange with extrajunctional pools. PMID:25057023

Mangiferin suppresses endoplasmic reticulum stress in perivascular adipose tissue and prevents insulin resistance in the endothelium.

PubMed

Xu, Xiaoshan; Chen, Yupeng; Song, Junna; Hou, Fangjie; Ma, Xuelian; Liu, Baolin; Huang, Fang

2018-06-01

Mangiferin is a naturally occurring glucosylxanthone with beneficial effects on glucose and lipid homeostasis. This study investigates the potential therapeutic effect of Mangiferin in perivascular adipose tissue (PVAT) and whether it contributes to regulating insulin action in the endothelium. Palmitate challenge evoked ROS-associated endoplasmic reticulum stress (ER stress) and NLRP3 inflammasome activation in PVAT. The conditioned medium from PA-stimulated PVAT was prepared to induce endothelial insulin resistance, and improved endothelium-dependent vasodilation in response to insulin was detected in vitro and in vivo. Mangiferin treatment enhanced LKB1-dependent AMPK activity and suppressed ER stress with downregulation of TXNIP induction, leading to the inhibition of NLRP3 inflammasome activation evidenced by attenuated NLRP3 and cleaved caspase-1 expression as well as reduced IL-1β secretion. Moreover, Mangiferin restored insulin-mediated Akt and eNOS phosphorylations with increased NO production, immunohistochemistry examination of adipocytes, and endothelial tissue in high-fat diet-fed mice also showed that oral administration of Mangiferin inhibited ER stress and NLRP3 induction in PVAT, and then effectively prevented insulin resistance in the vessel endothelium. Taken together, these results revealed that Mangiferin suppressed ER stress-associated NLRP3 inflammasome activation in PVAT through regulation of AMPK activity, which prevented endothelial insulin resistance. These findings suggested that the amelioration of PVAT dysfunction may be a therapeutic strategy for the prevention of endothelial insulin resistance.

Restoration handbook for sagebrush steppe ecosystems with emphasis on greater sage-grouse habitat—Part 1. Concepts for understanding and applying restoration

USGS Publications Warehouse

Pyke, David A.; Chambers, Jeanne C.; Pellant, Mike; Knick, Steven T.; Miller, Richard F.; Beck, Jeffrey L.; Doescher, Paul S.; Schupp, Eugene W.; Roundy, Bruce A.; Brunson, Mark; McIver, James D.

2015-10-26

Sagebrush steppe ecosystems in the United States currently occur on only about one-half of their historical land area because of changes in land use, urban growth, and degradation of land, including invasions of non-native plants. The existence of many animal species depends on the existence of sagebrush steppe habitat. The greater sage-grouse (Centrocercus urophasianus) is a landscape-dependent bird that requires intact habitat and combinations of sagebrush and perennial grasses to exist. In addition, other sagebrush-obligate animals also have similar requirements and restoration of landscapes for greater sage-grouse also will benefit these animals. Once sagebrush lands are degraded, they may require restoration actions to make those lands viable habitat for supporting sagebrushobligate animals. This restoration handbook is the first in a three-part series on restoration of sagebrush ecosystems. In Part 1, we discuss concepts surrounding landscape and restoration ecology of sagebrush ecosystems and greater sage-grouse that habitat managers and restoration practitioners need to know to make informed decisions regarding where and how to restore specific areas. We will describe the plant dynamics of sagebrush steppe ecosystems and their responses to major disturbances, fire, and defoliation. We will introduce the concepts of ecosystem resilience to disturbances and resistance to invasions of annual grasses within sagebrush steppe. An introduction to soils and ecological site information will provide insights into the specific plants that can be restored in a location. Soil temperature and moisture regimes are described as a tool for determining resilience and resistance and the potential for various restoration actions. Greater sage-grouse are considered landscape birds that require large areas of intact sagebrush steppe; therefore, we describe concepts of landscape ecology that aid our decisions regarding habitat restoration. We provide a brief overview of

Endoplasmic reticulum stress contributes to acetylcholine receptor degradation by promoting endocytosis in skeletal muscle cells.

PubMed

Du, Ailian; Huang, Shiqian; Zhao, Xiaonan; Zhang, Yun; Zhu, Lixun; Ding, Ji; Xu, Congfeng

2016-01-15

After binding by acetylcholine released from a motor neuron, a nicotinic acetylcholine receptor at the neuromuscular junction produces a localized end-plate potential, which leads to muscle contraction. Improper turnover and renewal of acetylcholine receptors contributes to the pathogenesis of myasthenia gravis. In the present study, we demonstrate that endoplasmic reticulum (ER) stress contributes to acetylcholine receptor degradation in C2C12 myocytes. We further show that ER stress promotes acetylcholine receptor endocytosis and lysosomal degradation, which was dampened by blocking endocytosis or treating with lysosome inhibitor. Knockdown of ER stress proteins inhibited acetylcholine receptor endocytosis and degradation, while rescue assay restored its endocytosis and degradation, confirming the effects of ER stress on promoting endocytosis-mediated degradation of junction acetylcholine receptors. Thus, our studies identify ER stress as a factor promoting acetylcholine receptor degradation through accelerating endocytosis in muscle cells. Blocking ER stress and/or endocytosis might provide a novel therapeutic approach for myasthenia gravis. Copyright © 2015 Elsevier B.V. All rights reserved.

The integrated endoplasmic reticulum stress response in Leishmania amazonensis macrophage infection: the role of X-box binding protein 1 transcription factor.

PubMed

Dias-Teixeira, Karina Luiza; Calegari-Silva, Teresa Cristina; dos Santos, Guilherme R R M; Vitorino Dos Santos, José; Lima, Carolina; Medina, Jorge Mansur; Aktas, Bertal Huseyin; Lopes, Ulisses G

2016-04-01

Endoplasmic reticulum (ER) stress triggers the integrated ER-stress response (IERSR) that ensures cellular survival of ER stress and represents a primordial form of innate immunity. We investigated the role of IERSR duringLeishmania amazonensisinfection. Treatment of RAW 264.7 infected macrophages with the ER stress-inducing agent thapsigargin (TG; 1 μM) increasedL. amazonensisinfectivity in an IFN1-α receptor (IFNAR)-dependent manner. In Western blot assays, we showed thatL. amazonensisactivates the inositol-requiring enzyme (IRE1)/ X-box binding protein (XBP)-1-splicing arms of the IERSR in host cells. In chromatin immunoprecipitation (ChIP) assays, we showed an increased occupancy of enhancer and promoter sequences for theIfnbgene by XBP1 in infected RAW 264.7 cells. Knocking down XBP1 expression by transducing RAW 264.7 cells with the short hairpin XBP1 lentiviral vector significantly reduced the parasite proliferation associated with impaired translocation of phosphorylated IFN regulatory transcription factor (IRF)-3 to the nucleus and a decrease in IFN1-β expression. Knocking down XBP1 expression also increased NO concentration, as determined by Griess reaction and reduced the expression of antioxidant genes, such as heme oxygenase (HO)-1, that protect parasites from oxidative stress. We conclude thatL. amazonensisactivation of XBP1 plays a critical role in infection by protecting the parasites from oxidative stress and increasing IFN1-β expression.-Dias-Teixeira, K. L., Calegari-Silva, T. C., Dos Santos, G. R. R. M., Vitorino dos Santos, J., Lima, C., Medina, J. M., Aktas, B. H., Lopes, U. G. The integrated endoplasmic reticulum stress response inLeishmania amazonensismacrophage infection: the role of X-box binding protein 1 transcription factor. © FASEB.

Endoplasmic reticulum stress in wake-active neurons progresses with aging.

PubMed

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.

Type 2 diabetes mellitus induces congenital heart defects in murine embryos by increasing oxidative stress, endoplasmic reticulum stress, and apoptosis.

PubMed

Wu, Yanqing; Reece, E Albert; Zhong, Jianxiang; Dong, Daoyin; Shen, Wei-Bin; Harman, Christopher R; Yang, Peixin

2016-09-01

Maternal type 1 and 2 diabetes mellitus are strongly associated with high rates of severe structural birth defects, including congenital heart defects. Studies in type 1 diabetic embryopathy animal models have demonstrated that cellular stress-induced apoptosis mediates the teratogenicity of maternal diabetes leading to congenital heart defect formation. However, the mechanisms underlying maternal type 2 diabetes mellitus-induced congenital heart defects remain largely unknown. We aim to determine whether oxidative stress, endoplasmic reticulum stress, and excessive apoptosis are the intracellular molecular mechanisms underlying maternal type 2 diabetes mellitus-induced congenital heart defects. A mouse model of maternal type 2 diabetes mellitus was established by feeding female mice a high-fat diet (60% fat). After 15 weeks on the high-fat diet, the mice showed characteristics of maternal type 2 diabetes mellitus. Control dams were either fed a normal diet (10% fat) or the high-fat diet during pregnancy only. Female mice from the high-fat diet group and the 2 control groups were mated with male mice that were fed a normal diet. At E12.5, embryonic hearts were harvested to determine the levels of lipid peroxides and superoxide, endoplasmic reticulum stress markers, cleaved caspase 3 and 8, and apoptosis. E17.5 embryonic hearts were harvested for the detection of congenital heart defect formation using India ink vessel patterning and histological examination. Maternal type 2 diabetes mellitus significantly induced ventricular septal defects and persistent truncus arteriosus in the developing heart, along with increasing oxidative stress markers, including superoxide and lipid peroxidation; endoplasmic reticulum stress markers, including protein levels of phosphorylated-protein kinase RNA-like endoplasmic reticulum kinase, phosphorylated-IRE1α, phosphorylated-eIF2α, C/EBP homologous protein, and binding immunoglobulin protein; endoplasmic reticulum chaperone gene

HSPB1 facilitates ERK-mediated phosphorylation and degradation of BIM to attenuate endoplasmic reticulum stress-induced apoptosis

PubMed Central

Kennedy, Donna; Mnich, Katarzyna; Oommen, Deepu; Chakravarthy, Reka; Almeida-Souza, Leonardo; Krols, Michiel; Saveljeva, Svetlana; Doyle, Karen; Gupta, Sanjeev; Timmerman, Vincent; Janssens, Sophie; Gorman, Adrienne M; Samali, Afshin

2017-01-01

BIM, a pro-apoptotic BH3-only protein, is a key regulator of the intrinsic (or mitochondrial) apoptosis pathway. Here, we show that BIM induction by endoplasmic reticulum (ER) stress is suppressed in rat PC12 cells overexpressing heat shock protein B1 (HSPB1 or HSP27) and that this is due to enhanced proteasomal degradation of BIM. HSPB1 and BIM form a complex that immunoprecipitates with p-ERK1/2. We found that HSPB1-mediated proteasomal degradation of BIM is dependent on MEK-ERK signaling. Other studies have shown that several missense mutations in HSPB1 cause the peripheral neuropathy, Charcot-Marie-Tooth (CMT) disease, which is associated with nerve degeneration. Here we show that cells overexpressing CMT-related HSPB1 mutants exhibited increased susceptibility to ER stress-induced cell death and high levels of BIM. These findings identify a novel function for HSPB1 as a negative regulator of BIM protein stability leading to protection against ER stress-induced apoptosis, a function that is absent in CMT-associated HSPB1 mutants. PMID:29048431

HSPB1 facilitates ERK-mediated phosphorylation and degradation of BIM to attenuate endoplasmic reticulum stress-induced apoptosis.

PubMed

Kennedy, Donna; Mnich, Katarzyna; Oommen, Deepu; Chakravarthy, Reka; Almeida-Souza, Leonardo; Krols, Michiel; Saveljeva, Svetlana; Doyle, Karen; Gupta, Sanjeev; Timmerman, Vincent; Janssens, Sophie; Gorman, Adrienne M; Samali, Afshin

2017-08-31

BIM, a pro-apoptotic BH3-only protein, is a key regulator of the intrinsic (or mitochondrial) apoptosis pathway. Here, we show that BIM induction by endoplasmic reticulum (ER) stress is suppressed in rat PC12 cells overexpressing heat shock protein B1 (HSPB1 or HSP27) and that this is due to enhanced proteasomal degradation of BIM. HSPB1 and BIM form a complex that immunoprecipitates with p-ERK1/2. We found that HSPB1-mediated proteasomal degradation of BIM is dependent on MEK-ERK signaling. Other studies have shown that several missense mutations in HSPB1 cause the peripheral neuropathy, Charcot-Marie-Tooth (CMT) disease, which is associated with nerve degeneration. Here we show that cells overexpressing CMT-related HSPB1 mutants exhibited increased susceptibility to ER stress-induced cell death and high levels of BIM. These findings identify a novel function for HSPB1 as a negative regulator of BIM protein stability leading to protection against ER stress-induced apoptosis, a function that is absent in CMT-associated HSPB1 mutants.

SLY1 and Syntaxin 18 specify a distinct pathway for procollagen VII export from the endoplasmic reticulum

PubMed Central

Nogueira, Cristina; Erlmann, Patrik; Villeneuve, Julien; Santos, António JM; Martínez-Alonso, Emma; Martínez-Menárguez, José Ángel; Malhotra, Vivek

2014-01-01

TANGO1 binds and exports Procollagen VII from the endoplasmic reticulum (ER). In this study, we report a connection between the cytoplasmic domain of TANGO1 and SLY1, a protein that is required for membrane fusion. Knockdown of SLY1 by siRNA arrested Procollagen VII in the ER without affecting the recruitment of COPII components, general protein secretion, and retrograde transport of the KDEL-containing protein BIP, and ERGIC53. SLY1 is known to interact with the ER-specific SNARE proteins Syntaxin 17 and 18, however only Syntaxin 18 was required for Procollagen VII export. Neither SLY1 nor Syntaxin 18 was required for the export of the equally bulky Procollagen I from the ER. Altogether, these findings reveal the sorting of bulky collagen family members by TANGO1 at the ER and highlight the existence of different export pathways for secretory cargoes one of which is mediated by the specific SNARE complex containing SLY1 and Syntaxin 18. DOI: http://dx.doi.org/10.7554/eLife.02784.001 PMID:24842878

Ablation of PGC1 beta prevents mTOR dependent endoplasmic reticulum stress response

PubMed Central

Camacho, Alberto; Rodriguez-Cuenca, Sergio; Blount, Margaret; Prieur, Xavier; Barbarroja, Nuria; Fuller, Maria; Hardingham, Giles E.; Vidal-Puig, Antonio

2012-01-01

Mitochondria dysfunction contributes to the pathophysiology of obesity, diabetes, neurodegeneration and ageing. The peroxisome proliferator-activated receptor-gamma coactivator-1β (PGC-1β) coordinates mitochondrial biogenesis and function as well as fatty acid metabolism. It has been suggested that endoplasmic reticulum (ER) stress may be one of the mechanisms linking mitochondrial dysfunction and these pathologies. Here we investigate whether PGC-1β ablation affects the ER stress response induced by specific nutritional and pharmacological challenges in the CNS. By using flow cytometry, western blot, real time PCR and several pharmacological and nutritional interventions in PGC-1β knock out and WT mice, we confirmed that PGC-1β coordinates mitochondria function in brain and reported for the first time that a) ablation of PGC-1β is associated with constitutive activation of mTORC1 pathway associated with increased basal GRP78 protein levels in hypothalamus and cortex of animals fed chow diet; and b) in animals fed chronically with high fat diet (HFD) or high protein diet (HPD), we observed a failure to appropriately induce ER stress response in the absence of PGC-1β, associated with an increase in mTOR pathway phosphorylation. This contrasted with the appropriate upregulation of ER stress response observed in wild type littermates. Additionally, inefficient in vitro induction of ER stress by thapsigargin seems result in apoptotic neuronal cell death in PGC-1β KO. Our data indicate that PGC-1β is required for a neuronal ER response to nutritional stress imposed by HFD and HPD diets and that genetic ablation of PGC-1β might increase the susceptibility to neuronal damage and cell death. PMID:22771762

Effects of Si/Al Ratio on Cu/SSZ-13 NH3-SCR Catalysts: Implications for the active Cu species and the Roles of Brønsted Acidity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gao, Feng; Washton, Nancy M.; Wang, Yilin

2015-09-03

Cu/SSZ-13 catalysts with three Si/Al ratios of 6, 12 and 35 were synthesized with Cu incorporation via solution ion exchange. The implications of varying Si/Al ratios on the nature of the multiple Cu species that can be present in the SSZ-13 zeolite are a major focus of this work, as highlighted by the results of a variety of catalyst characterization and reaction kinetics measurements. Specifically, catalysts were characterized with surface area/pore volume measurements, temperature programmed reduction by H2 (H2-TPR), NH3 temperature programmed desorption (NH3-TPD), and DRIFTS and solid-state nuclear magnetic resonance (NMR) spectroscopies. Catalytic properties were examined using NO oxidation,more » ammonia oxidation, and standard ammonia selective catalytic reduction (NH3-SCR) reactions on selected catalysts under differential conditions. Besides indicating possible variably active multiple Cu species for these reactions, the measurements are also used to untangle some of the complexities caused by the interplay between redox of Cu ion centers and Brønsted acidity. All three reactions appear to follow a redox reaction mechanism, yet the roles of Brønsted acidity are quite different. For NO oxidation, increasing Si/Al ratio lowers Cu redox barriers, thus enhancing reaction rates. Brønsted acidity appears to play essentially no role for this reaction. For standard NH3-SCR, residual Brønsted acidity plays a significant beneficial role at both low- and high-temperature regimes. For NH3 oxidation, no clear trend is observed suggesting both Cu ion center redox and Brønsted acidity play important and perhaps competing roles. The authors gratefully acknowledge the US Department of Energy (DOE), Energy Efficiency and Renewable Energy, Vehicle Technologies Office for the support of this work. The research described in this paper was performed in the Environmental Molecular Sciences Laboratory (EMSL), a national scientific user facility sponsored by the DOE�

XBP1 (X-Box-Binding Protein-1)-Dependent O-GlcNAcylation Is Neuroprotective in Ischemic Stroke in Young Mice and Its Impairment in Aged Mice Is Rescued by Thiamet-G.

PubMed

Jiang, Meng; Yu, Shu; Yu, Zhui; Sheng, Huaxin; Li, Ying; Liu, Shuai; Warner, David S; Paschen, Wulf; Yang, Wei

2017-06-01

Impaired protein homeostasis induced by endoplasmic reticulum dysfunction is a key feature of a variety of age-related brain diseases including stroke. To restore endoplasmic reticulum function impaired by stress, the unfolded protein response is activated. A key unfolded protein response prosurvival pathway is controlled by the endoplasmic reticulum stress sensor (inositol-requiring enzyme-1), XBP1 (downstream X-box-binding protein-1), and O-GlcNAc (O-linked β-N-acetylglucosamine) modification of proteins (O-GlcNAcylation). Stroke impairs endoplasmic reticulum function, which activates unfolded protein response. The rationale of this study was to explore the potentials of the IRE1/XBP1/O-GlcNAc axis as a target for neuroprotection in ischemic stroke. Mice with Xbp1 loss and gain of function in neurons were generated. Stroke was induced by transient or permanent occlusion of the middle cerebral artery in young and aged mice. Thiamet-G was used to increase O-GlcNAcylation. Deletion of Xbp1 worsened outcome after transient and permanent middle cerebral artery occlusion. After stroke, O-GlcNAcylation was activated in neurons of the stroke penumbra in young mice, which was largely Xbp1 dependent. This activation of O-GlcNAcylation was impaired in aged mice. Pharmacological increase of O-GlcNAcylation before or after stroke improved outcome in both young and aged mice. Our study indicates a critical role for the IRE1/XBP1 unfolded protein response branch in stroke outcome. O-GlcNAcylation is a prosurvival pathway that is activated in the stroke penumbra in young mice but impaired in aged mice. Boosting prosurvival pathways to counterbalance the age-related decline in the brain's self-healing capacity could be a promising strategy to improve ischemic stroke outcome in aged brains. © 2017 American Heart Association, Inc.

Endoplasmic reticulum stress related molecular mechanisms in nonalcoholic fatty liver disease (NAFLD).

PubMed

Wang, Lifeng; Chen, J; Ning, C; Lei, D; Ren, Jun

2018-05-16

Non-alcoholic fatty liver disease (NAFLD) has emerged as a common public health problem and a common cause of chronic liver diseases. However, the underlying mechanisms leading to the development and progression of NAFLD remain elusive. Accumulating evidence has depicted an essential role for endoplasmic reticulum (ER) stress in the development of steatosis and later progression into nonalcoholic steatohepatitis and hepatocarcinoma. With the accumulation of unfolded and misfolded proteins in the ER lumen, ER stress is provoked to turn on the unfolded protein response (UPR). ER stress triggers a cascade reaction of transcriptional and translational events that restore ER homeostasis, promoting cell survival and adaptation. However, prolonged ER stress may be transit physiological mechanisms to pathological consequences, including insulin resistance, fat accumulation, inflammation, apoptosis, and autophagy, all of which with important roles in the development of NAFLD. Therefore, understanding the role of ER stress in the onset and pathogenesis of NAFLD is pertinent to the management of this devastating metabolic disease. Here we will summarize available information on recent findings linking ER stress to the pathogenesis of NAFLD. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Resveratrol prevents doxorubicin-induced cardiotoxicity in H9c2 cells through the inhibition of endoplasmic reticulum stress and the activation of the Sirt1 pathway.

PubMed

Lou, Yu; Wang, Zhen; Xu, Yi; Zhou, Ping; Cao, Junxian; Li, Yuanshi; Chen, Yeping; Sun, Junfeng; Fu, Lu

2015-09-01

Treatment with doxorubicin (DOX) is one of the major causes of chemotherapy-induced cardiotoxicity and is therefore, the principal limiting factor in the effectiveness of chemotherapy for cancer patients. DOX‑induced heart failure is thought to result from endoplasmic reticulum (ER) stress and cardiomyocyte apoptosis. Resveratrol (RV), a polyphenol antioxidant found in red wine, has been shown to play a cardioprotective role. The aim of the present study was to examine the effects of RV on DOX‑induced cardiotoxicity in H9c2 cells. We hypothesized that RV would protect H9c2 cells against DOX‑induced ER stress and subsequent cell death through the activation of the Sirt1 pathway. Our results demonstrated that the decrease observed in the viability of the H9c2 cells following exposure to DOX was accompanied by a significant increase in the expression of the ER stress‑related proteins, glucose‑regulated protein 78 (GRP78) and C/EBP homologous protein (CHOP). However, we found that RV downregulated the expression of ER stress marker protein in the presence of DOX and restored the viability of the H9c2 cells. Exposure to RV or DOX alone only slightly increased the protein expression of Sirt1, whereas a significant increase in Sirt1 protein levels was observed in the cells treated with both RV and DOX. The Sirt1 inhibitor, nicotinamide (NIC), partially neutralized the effects of RV on the expression of Sirt1 in the DOX‑treated cells and completely abolished the effects of RV on the expression of GRP78 and CHOP. The findings of our study suggest that RV protects H9c2 cells against DOX‑induced ER stress through ER stabilization, and more specifically through the activation of the Sirt1 pathway, thereby leading to cardiac cell survival.

Gemcitabine treatment induces endoplasmic reticular (ER) stress and subsequently upregulates urokinase plasminogen activator (uPA) to block mitochondrial-dependent apoptosis in Panc-1 cancer stem-like cells (CSCs).

PubMed

Wang, Li; Zhang, Yi; Wang, Weiguo; Zhu, Yunjie; Chen, Yang; Tian, Bole

2017-01-01

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive cancer with poor survival rates. The presence of cancer stem-like cells (CSCs) is believed to be among the underlying reasons for the aggressiveness of PDAC, which contributes to chemoresistance and recurrence. However, the mechanisms that induce chemoresistance and inhibit apoptosis remain largely unknown. We used serum-free medium to enrich CSCs from panc-1 human pancreatic cancer cells and performed sphere formation testing, flow cytometry, quantitative reverse transcription polymerase chain reaction (RT-qPCR) and semi-quantitative western blotting to confirm the stemness of panc-1 CSCs. Hallmarks of endoplasmic reticulum (ER) stress, including IRE1, PERK, ATF4, ATF6α, GRP78 and uPA expression, were detected after gemcitabine treatment. Effects of gemcitabine-induced uPA expression on cell invasion, sphere formation, colony formation and gemcitabine sensitivity were detected. Electrophoretic mobility shift assays (EMSAs) and RNA-immunoprecipitation (RIP) were performed to detect interaction between the uPA mRNA 3'-UTR and mutant p53-R273H expressed by panc-1 CSCs. The effects of upregulated uPA by gemcitabine on apoptosis were detected by Annexin V-FITC/PI staining, and the impact of uPA on small molecule CP-31398-restored mutant p53 transcriptional activity was measured by a luciferase reporter assay. Enriched panc-1 CSCs expressing high levels of CD44 and CD133 also produced significantly higher amounts of Oct4 and Nanog. Compared with panc-1 cells, panc-1 CSCs presented chemoresistance to gemcitabine. ER stress gene detections demonstrated effects of gemcitabine-induced ER stress on both the pro-apoptotic and pro-survival branches. ER stress-induced ATF6α upregulated level of uPA by transcriptionally activating GRP78. Gemcitabine-induced uPA promoted invasion, sphere formation and colony formation and attenuated apoptosis induced by gemcitabine in panc-1 CSCs, depending on interaction with mutant p53

Gemcitabine treatment induces endoplasmic reticular (ER) stress and subsequently upregulates urokinase plasminogen activator (uPA) to block mitochondrial-dependent apoptosis in Panc-1 cancer stem-like cells (CSCs)

PubMed Central

Wang, Weiguo; Zhu, Yunjie; Chen, Yang

2017-01-01

Background Pancreatic ductal adenocarcinoma (PDAC) is an aggressive cancer with poor survival rates. The presence of cancer stem-like cells (CSCs) is believed to be among the underlying reasons for the aggressiveness of PDAC, which contributes to chemoresistance and recurrence. However, the mechanisms that induce chemoresistance and inhibit apoptosis remain largely unknown. Methods We used serum-free medium to enrich CSCs from panc-1 human pancreatic cancer cells and performed sphere formation testing, flow cytometry, quantitative reverse transcription polymerase chain reaction (RT-qPCR) and semi-quantitative western blotting to confirm the stemness of panc-1 CSCs. Hallmarks of endoplasmic reticulum (ER) stress, including IRE1, PERK, ATF4, ATF6α, GRP78 and uPA expression, were detected after gemcitabine treatment. Effects of gemcitabine-induced uPA expression on cell invasion, sphere formation, colony formation and gemcitabine sensitivity were detected. Electrophoretic mobility shift assays (EMSAs) and RNA-immunoprecipitation (RIP) were performed to detect interaction between the uPA mRNA 3’-UTR and mutant p53-R273H expressed by panc-1 CSCs. The effects of upregulated uPA by gemcitabine on apoptosis were detected by Annexin V-FITC/PI staining, and the impact of uPA on small molecule CP-31398-restored mutant p53 transcriptional activity was measured by a luciferase reporter assay. Results Enriched panc-1 CSCs expressing high levels of CD44 and CD133 also produced significantly higher amounts of Oct4 and Nanog. Compared with panc-1 cells, panc-1 CSCs presented chemoresistance to gemcitabine. ER stress gene detections demonstrated effects of gemcitabine-induced ER stress on both the pro-apoptotic and pro-survival branches. ER stress-induced ATF6α upregulated level of uPA by transcriptionally activating GRP78. Gemcitabine-induced uPA promoted invasion, sphere formation and colony formation and attenuated apoptosis induced by gemcitabine in panc-1 CSCs, depending on

Revisiting PC1/3 Mutants: Dominant-Negative Effect of Endoplasmic Reticulum-Retained Mutants.

PubMed

Blanco, Elias H; Ramos-Molina, Bruno; Lindberg, Iris

2015-10-01

Prohormone convertase 1/3 (PC1/3), encoded by the gene PCSK1, is critical for peptide hormone synthesis. An increasing number of studies have shown that inactivating mutations in PCSK1 are correlated with endocrine pathologies ranging from intestinal dysfunction to morbid obesity, whereas the common nonsynonymous polymorphisms rs6232 (N221D) and rs6234-rs6235 (Q665E-S690T) are highly associated with obesity risk. In this report, we revisited the biochemical and cellular properties of PC1/3 variants in the context of a wild-type PC1/3 background instead of the S357G hypermorph background used for all previous studies. In the wild-type background the PC1/3 N221D variant exhibited 30% lower enzymatic activity in a fluorogenic assay than wild-type PC1/3; this inhibition was greater than that detected in an equivalent experiment using the PC1/3 S357G background. A PC1/3 variant with the linked carboxyl-terminal polymorphisms Q665E-S690T did not show this difference. We also analyzed the biochemical properties of 2 PC1/3 mutants, G209R and G593R, which are retained in the endoplasmic reticulum (ER), and studied their effects on wild-type PC1/3. The expression of ER-retained mutants induced ER stress markers and also resulted in dominant-negative blockade of wild-type PC1/3 prodomain cleavage and decreased expression of wild-type PC1/3, suggesting facilitation of the entry of wild-type protein to a degradative proteasomal pathway. Dominant-negative effects of PC1/3 mutations on the expression and maturation of wild-type protein, with consequential effects on PC1/3 availability, add a new element which must be considered in population and clinical studies of this gene.

EBS7 is a plant-specific component of a highly conserved endoplasmic reticulum-associated degradation system in Arabidopsis

PubMed Central

Liu, Yidan; Zhang, Congcong; Wang, Dinghe; Su, Wei; Liu, Linchuan; Wang, Muyang; Li, Jianming

2015-01-01

Endoplasmic reticulum (ER)-associated degradation (ERAD) is an essential part of an ER-localized protein quality-control system for eliminating terminally misfolded proteins. Recent studies have demonstrated that the ERAD machinery is conserved among yeast, animals, and plants; however, it remains unknown if the plant ERAD system involves plant-specific components. Here we report that the Arabidopsis ethyl methanesulfonate-mutagenized brassinosteroid-insensitive 1 suppressor 7 (EBS7) gene encodes an ER membrane-localized ERAD component that is highly conserved in land plants. Loss-of-function ebs7 mutations prevent ERAD of brassinosteroid insensitive 1-9 (bri1-9) and bri1-5, two ER-retained mutant variants of the cell-surface receptor for brassinosteroids (BRs). As a result, the two mutant receptors accumulate in the ER and consequently leak to the plasma membrane, resulting in the restoration of BR sensitivity and phenotypic suppression of the bri1-9 and bri1-5 mutants. EBS7 accumulates under ER stress, and its mutations lead to hypersensitivity to ER and salt stresses. EBS7 interacts with the ER membrane-anchored ubiquitin ligase Arabidopsis thaliana HMG-CoA reductase degradation 1a (AtHrd1a), one of the central components of the Arabidopsis ERAD machinery, and an ebs7 mutation destabilizes AtHrd1a to reduce polyubiquitination of bri1-9. Taken together, our results uncover a plant-specific component of a plant ERAD pathway and also suggest its likely biochemical function. PMID:26371323

Arresting a Torsin ATPase Reshapes the Endoplasmic Reticulum*

PubMed Central

Rose, April E.; Zhao, Chenguang; Turner, Elizabeth M.; Steyer, Anna M.; Schlieker, Christian

2014-01-01

Torsins are membrane-tethered AAA+ ATPases residing in the nuclear envelope (NE) and endoplasmic reticulum (ER). Here, we show that the induction of a conditional, dominant-negative TorsinB variant provokes a profound reorganization of the endomembrane system into foci containing double membrane structures that are derived from the ER. These double-membrane sinusoidal structures are formed by compressing the ER lumen to a constant width of 15 nm, and are highly enriched in the ATPase activator LULL1. Further, we define an important role for a highly conserved aromatic motif at the C terminus of Torsins. Mutations in this motif perturb LULL1 binding, reduce ATPase activity, and profoundly limit the induction of sinusoidal structures. PMID:24275647

EDEM1 targets misfolded HLA-B27 dimers for endoplasmic reticulum associated degradation

PubMed Central

Guiliano, David B.; Fussell, Helen; Lenart, Izabela; Tsao, Edward; Nesbeth, Darren; Fletcher, Adam J.; Campbell, Elaine C.; Yousaf, Nasim; Williams, Sarah; Santos, Susana; Cameron, Amy; Towers, Greg J.; Kellam, Paul; Hebert, Daniel N.; Gould, Keith; Powis, Simon J.; Antoniou, Antony N.

2015-01-01

Objective HLA-B27 forms misfolded heavy chain dimers, which may predispose individuals to inflammatory arthritis by inducing endoplasmic reticulum (ER) stress and the unfolded protein response (UPR). We wanted to define the role of the UPR induced ER associated degradation (ERAD) pathway in the disposal of HLA-B27 dimeric conformers. Methods HeLa cell lines expressing only two copies of a carboxy terminally Sv5 tagged HLA-B27 were generated. The ER stress induced EDEM1 protein was over expressed by transfection and dimer levels monitored by immunoblotting. EDEM1, the UPR associated transcription factor XBP-1, the E3 ubiquitin ligase HRD1, the degradation associated derlin 1 and 2 proteins were inhibited by either short hairpin RNA or dominant negative mutants. The UPR associated ERAD of HLA-B27 was confirmed using ER stress inducing pharamacological agents in kinetic and pulse chase assays. Results We demonstrate that UPR induced machinery can target HLA-B27 dimers, and that dimer formation can be controlled by alterations to expression levels of components of the UPR induced ERAD pathway. HLA-B27 dimers and misfolded MHC class I monomeric molecules were detected bound to EDEM1, with overexpression of EDEM1 inhibiting HLA-B27 dimer formation. EDEM1 inhibition resulted in upregulation of HLA-B27 dimers, whilst UPR induced ERAD of dimers was prevented in the absence of EDEM1. HLA-B27 dimer formation was also enhanced in the absence of XBP-1, HRD1 and derlin1/2. Conclusion The UPR ERAD pathway as described here can dispose of HLA-B27 dimers and presents a potential novel therapeutic target for the modulation of HLA-B27 associated inflammatory disease. PMID:25132672

Systemic restoration of UBA1 ameliorates disease in spinal muscular atrophy

PubMed Central

Powis, Rachael A.; Karyka, Evangelia; Boyd, Penelope; Côme, Julien; Jones, Ross A.; Zheng, Yinan; Szunyogova, Eva; Groen, Ewout J.N.; Hunter, Gillian; Thomson, Derek; Wishart, Thomas M.; Becker, Catherina G.; Parson, Simon H.; Martinat, Cécile; Azzouz, Mimoun; Gillingwater, Thomas H.

2016-01-01

The autosomal recessive neuromuscular disease spinal muscular atrophy (SMA) is caused by loss of survival motor neuron (SMN) protein. Molecular pathways that are disrupted downstream of SMN therefore represent potentially attractive therapeutic targets for SMA. Here, we demonstrate that therapeutic targeting of ubiquitin pathways disrupted as a consequence of SMN depletion, by increasing levels of one key ubiquitination enzyme (ubiquitin-like modifier activating enzyme 1 [UBA1]), represents a viable approach for treating SMA. Loss of UBA1 was a conserved response across mouse and zebrafish models of SMA as well as in patient induced pluripotent stem cell–derive motor neurons. Restoration of UBA1 was sufficient to rescue motor axon pathology and restore motor performance in SMA zebrafish. Adeno-associated virus serotype 9–UBA1 (AAV9-UBA1) gene therapy delivered systemic increases in UBA1 protein levels that were well tolerated over a prolonged period in healthy control mice. Systemic restoration of UBA1 in SMA mice ameliorated weight loss, increased survival and motor performance, and improved neuromuscular and organ pathology. AAV9-UBA1 therapy was also sufficient to reverse the widespread molecular perturbations in ubiquitin homeostasis that occur during SMA. We conclude that UBA1 represents a safe and effective therapeutic target for the treatment of both neuromuscular and systemic aspects of SMA. PMID:27699224

Loss of Oca2 disrupts the unfolded protein response and increases resistance to endoplasmic reticulum stress in melanocytes.

PubMed

Cheng, Tsing; Orlow, Seth J; Manga, Prashiela

2013-11-01

Accumulation of proteins in the endoplasmic reticulum (ER) typically induces stress and initiates the unfolded protein response (UPR) to facilitate recovery. If homeostasis is not restored, apoptosis is induced. However, adaptation to chronic UPR activation can increase resistance to subsequent acute ER stress. We therefore investigated adaptive mechanisms in Oculocutaneous albinism type 2 (Oca2)-null melanocytes where UPR signaling is arrested despite continued tyrosinase accumulation leading to resistance to the chemical ER stressor thapsigargin. Although thapsigargin triggers UPR activation, instead of Perk-mediated phosphorylation of eIF2α, in Oca2-null melanocytes, eIF2α was rapidly dephosphorylated upon treatment. Dephosphorylation was mediated by the Gadd34-PP1α phosphatase complex. Gadd34-complex inhibition blocked eIF2α dephosphorylation and significantly increased Oca2-null melanocyte sensitivity to thapsigargin. Thus, Oca2-null melanocytes adapt to acute ER stress by disruption of pro-apoptotic Perk signaling, which promotes cell survival. This is the first study to demonstrate rapid eIF2α dephosphorylation as an adaptive mechanism to ER stress. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

The Specificity of Trimming of MHC Class I-Presented Peptides in the Endoplasmic Reticulum1

PubMed Central

Hearn, Arron; York, Ian A.; Rock, Kenneth L.

2010-01-01

Aminopeptidases in the endoplasmic reticulum (ER) can cleave antigenic peptides and in so doing either create or destroy MHC class I-presented epitopes. However the specificity of this trimming process overall and of the major ER aminopeptidase ERAP1 in particular is not well understood. This issue is important because peptide trimming influences the magnitude and specificity of CD8 T cell responses. By systematically varying the N-terminal flanking sequences of peptides in a cell free biochemical system and in intact cells, we elucidated the specificity of ERAP1 and of ER trimming overall. ERAP1 can cleave after many amino acids on the N-terminus of epitope precursors but does so at markedly different rates. The specificity seen with purified ERAP1 is similar to that observed for trimming and presentation of epitopes in the ER of intact cells. We define N-terminal sequences that are favorable or unfavorable for antigen presentation in ways that are independent from the epitopes core sequence. When databases of known presented peptides were analyzed, the residues that were preferred for the trimming of model peptide precursors were found to be overrepresented in N-terminal flanking sequences of epitopes generally. These data define key determinants in the specificity of antigen processing. PMID:19828632

Sigmar1 regulates endoplasmic reticulum stress-induced C/EBP-homologous protein expression in cardiomyocytes.

PubMed

Alam, Shafiul; Abdullah, Chowdhury S; Aishwarya, Richa; Orr, A Wayne; Traylor, James; Miriyala, Sumitra; Panchatcharam, Manikandan; Pattillo, Christopher B; Bhuiyan, Md Shenuarin

2017-08-31

C/EBP-homologous protein (CHOP) is a ubiquitously expressed stress-inducible transcription factor robustly induced by maladaptive endoplasmic reticulum (ER) stresses in a wide variety of cells. Here, we examined a novel function of Sigma 1 receptor (Sigmar1) in regulating CHOP expression under ER stress in cardiomyocytes. We also defined Sigmar1-dependent activation of the adaptive ER-stress pathway in regulating CHOP expression. We used adenovirus-mediated Sigmar1 overexpression as well as Sigmar1 knockdown by siRNA in neonatal rat ventricular cardiomyocytes (NRCs); to induce ER stress, cardiomyocytes were treated with tunicamycin. Sigmar1-siRNA knockdown significantly increased the expression of CHOP and significantly induced cellular toxicity by sustained activation of ER stress in cardiomyocytes. Sigmar1 overexpression decreased the expression of CHOP and significantly decreased cellular toxicity in cells. Using biochemical and immunocytochemical experiments, we also defined the specific ER-stress pathway associated with Sigmar1-dependent regulation of CHOP expression and cellular toxicity. We found that Sigmar1 overexpression significantly increased inositol requiring kinase 1α (IRE1α) phosphorylation and increased spliced X-box-binding proteins (XBP1s) expression as well as nuclear localization. In contrast, Sigmar1 knockdown significantly decreased IRE1α phosphorylation and decreased XBP1s expression as well as nuclear transport. Taken together, these results indicate that Sigmar1-dependent activation of IRE1α-XBP1s ER-stress response pathways are associated with inhibition of CHOP expression and suppression of cellular toxicity. Hence, Sigmar1 is an essential component of the adaptive ER-stress response pathways eliciting cellular protection in cardiomyocytes. © 2017 The Author(s).

Sigmar1 regulates endoplasmic reticulum stress-induced C/EBP-homologous protein expression in cardiomyocytes

PubMed Central

Alam, Shafiul; Abdullah, Chowdhury S.; Aishwarya, Richa; Orr, A. Wayne; Traylor, James; Miriyala, Sumitra; Panchatcharam, Manikandan; Pattillo, Christopher B.

2017-01-01

C/EBP-homologous protein (CHOP) is a ubiquitously expressed stress-inducible transcription factor robustly induced by maladaptive endoplasmic reticulum (ER) stresses in a wide variety of cells. Here, we examined a novel function of Sigma 1 receptor (Sigmar1) in regulating CHOP expression under ER stress in cardiomyocytes. We also defined Sigmar1-dependent activation of the adaptive ER-stress pathway in regulating CHOP expression. We used adenovirus-mediated Sigmar1 overexpression as well as Sigmar1 knockdown by siRNA in neonatal rat ventricular cardiomyocytes (NRCs); to induce ER stress, cardiomyocytes were treated with tunicamycin. Sigmar1-siRNA knockdown significantly increased the expression of CHOP and significantly induced cellular toxicity by sustained activation of ER stress in cardiomyocytes. Sigmar1 overexpression decreased the expression of CHOP and significantly decreased cellular toxicity in cells. Using biochemical and immunocytochemical experiments, we also defined the specific ER-stress pathway associated with Sigmar1-dependent regulation of CHOP expression and cellular toxicity. We found that Sigmar1 overexpression significantly increased inositol requiring kinase 1α (IRE1α) phosphorylation and increased spliced X-box-binding proteins (XBP1s) expression as well as nuclear localization. In contrast, Sigmar1 knockdown significantly decreased IRE1α phosphorylation and decreased XBP1s expression as well as nuclear transport. Taken together, these results indicate that Sigmar1-dependent activation of IRE1α-XBP1s ER-stress response pathways are associated with inhibition of CHOP expression and suppression of cellular toxicity. Hence, Sigmar1 is an essential component of the adaptive ER-stress response pathways eliciting cellular protection in cardiomyocytes. PMID:28667101

Coastal Prairie Restoration Information System: Version 1 (Louisiana)

USGS Publications Warehouse

Allain, Larry

2007-01-01

The Coastal Prairie Restoration Information System (CPR) is a Microsoft Access database that allows users to query and view data about Louisiana coastal prairie species. Less than 0.1% of Louisiana's coastal prairie vegetation remains in a relatively undisturbed condition. Encompassing as much as 1 million hectares of land, coastal prairie is a hybrid of coastal wetlands and tall grass prairie. Over 550 plant species have been identified in Louisiana's coastal prairies to date. Efforts to conserve and restore this endangered ecosystem are limited by the ability of workers to identify and access knowledge about this diverse group of plants. In this database, a variety of data are provided for each of 650 coastal prairie species in Louisiana. The database was developed at the U.S. Geological Survey National Wetlands Research Center by Larry Allain, with software development by Myra Silva. Additional funding was provided by the biology department of the University of Louisiana at Lafayette (ULL), the ULL Center for Environmental and Ecological Technology, and the National Science Foundation.

Cytoprotective role of the fatty acid binding protein 4 against oxidative and endoplasmic reticulum stress in 3T3-L1 adipocytes

PubMed Central

Kajimoto, Kazuaki; Minami, Yoshitaka; Harashima, Hideyoshi

2014-01-01

The fatty acid binding protein 4 (FABP4), one of the most abundant proteins in adipocytes, has been reported to have a proinflammatory function in macrophages. However, the physiological role of FABP4, which is constitutively expressed in adipocytes, has not been fully elucidated. Previously, we demonstrated that FABP4 was involved in the regulation of interleukin-6 (IL-6) and vascular endothelial growth factor (VEGF) production in 3T3-L1 adipocytes. In this study, we examined the effects of FABP4 silencing on the oxidative and endoplasmic reticulum (ER) stress in 3T3-L1 adipocytes. We found that the cellular reactive oxygen species (ROS) and 8-nitro-cyclic GMP levels were significantly elevated in the differentiated 3T3-L1 adipocytes transfected with a small interfering RNA (siRNA) against Fabp4, although the intracellular levels or enzyme activities of antioxidants including reduced glutathione (GSH), superoxide dismutase (SOD) and glutathione S-transferase A4 (GSTA4) were not altered. An in vitro evaluation using the recombinant protein revealed that FABP4 itself functions as a scavenger protein against hydrogen peroxide (H2O2). FABP4-knockdown resulted in a significant lowering of cell viability of 3T3-L1 adipocytes against H2O2 treatment. Moreover, four kinds of markers related to the ER stress response including the endoplasmic reticulum to nucleus signaling 1 (Ern1), the signal sequence receptor α (Ssr1), the ORM1-like 3 (Ormdl3), and the spliced X-box binding protein 1 (Xbp1s), were all elevated as the result of the knockdown of FABP4. Consequently, FABP4 might have a new role as an antioxidant protein against H2O2 and contribute to cytoprotection against oxidative and ER stress in adipocytes. PMID:25161868

Quantification of apolipoprotein B-48 and B-100 in rat liver endoplasmic reticulum and Golgi fractions.

PubMed Central

Cartwright, I J; Higgins, J A

1992-01-01

We have developed a method for measurement of apolipoprotein (apo) B-48 and apo B-100 in blood and subcellular fractions of rat liver based on SDS/PAGE followed by quantitative immunoblotting using 125I-Protein A. Standard curves were prepared in each assay using apo B prepared from total rat lipoproteins by extraction with tetramethylurea. Subcellular fractions (rough and smooth endoplasmic reticulum and Golgi fractions) were prepared from rat liver and separated into membrane and cisternal-content fractions. For quantification, membrane fractions were solubilized in Triton X-100, and the apo B was immunoprecipitated before separation by SDS/PAGE and immunoblotting. Content fractions were concentrated by ultrafiltration and separated by SDS/PAGE without immunoprecipitation. Quantification of apo B in subcellular fractions and detection of apo B by immunoblotting yielded consistent results. In all fractions apo B-48 was the major form, accounting for approximately three-quarters of the total apo B. By using marker enzymes as internal standards, it was calculated that all of the apo B was recovered in the endoplasmic reticulum and Golgi fractions, with approximately 80% of each form of apo B in the endoplasmic reticulum. More than 90% of the apo B of the rough- and smooth-endoplasmic-reticulum fractions was membrane-bound, whereas approx. 33 and 15% of the apo B of the cis-enriched Golgi fractions and trans-enriched Golgi fractions respectively were membrane-bound. Images Fig. 1. Fig. 3. Fig. 4. PMID:1637294

Determinant for Endoplasmic Reticulum Retention in the Luminal Domain of the Human Cytomegalovirus US3 Glycoprotein

PubMed Central

Lee, Sungwook; Park, Boyoun; Ahn, Kwangseog

2003-01-01

US3 of human cytomegalovirus is an endoplasmic reticulum resident transmembrane glycoprotein that binds to major histocompatibility complex class I molecules and prevents their departure. The endoplasmic reticulum retention signal of the US3 protein is contained in the luminal domain of the protein. To define the endoplasmic reticulum retention sequence in more detail, we have generated a series of deletion and point mutants of the US3 protein. By analyzing the rate of intracellular transport and immunolocalization of the mutants, we have identified Ser58, Glu63, and Lys64 as crucial for retention, suggesting that the retention signal of the US3 protein has a complex spatial arrangement and does not comprise a contiguous sequence of amino acids. We also show that a modified US3 protein with a mutation in any of these amino acids maintains its ability to bind class I molecules; however, such mutated proteins are no longer retained in the endoplasmic reticulum and are not able to block the cell surface expression of class I molecules. These findings indicate that the properties that allow the US3 glycoprotein to be localized in the endoplasmic reticulum and bind major histocompatibility complex class I molecules are located in different parts of the molecule and that the ability of US3 to block antigen presentation is due solely to its ability to retain class I molecules in the endoplasmic reticulum. PMID:12525649

4-Phenylbutyric Acid Reveals Good Beneficial Effects on Vital Organ Function via Anti-Endoplasmic Reticulum Stress in Septic Rats.

PubMed

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-endoplasmic

Peroxisomal Ascorbate Peroxidase Resides within a Subdomain of Rough Endoplasmic Reticulum in Wild-Type Arabidopsis Cells1

PubMed Central

Lisenbee, Cayle S.; Heinze, Michael; Trelease, Richard N.

2003-01-01

Previously we reported (R.T. Mullen, C.S. Lisenbee, J.A. Miernyk, R.N. Trelease [1999] Plant Cell 11: 2167–2185) that overexpressed ascorbate peroxidase (APX), a peroxisomal membrane protein, sorted indirectly to Bright Yellow-2 cell peroxisomes via a subdomain of the endoplasmic reticulum (ER; peroxisomal endoplasmic reticulum [pER]). More recently, a pER-like compartment also was identified in pumpkin (Cucurbita pepo) and transformed Arabidopsis cells (K. Nito, K. Yamaguchi, M. Kondo, M. Hayashi, M. Nishimura [2001] Plant Cell Physiol 42: 20–27). Here, we characterize more extensively the localization of endogenous Arabidopsis peroxisomal APX (AtAPX) in cultured wild-type Arabidopsis cells (Arabidopsis var. Landsberg erecta). AtAPX was detected in peroxisomes, but not in an ER subcompartment, using immunofluorescence microscopy. However, AtAPX was detected readily with immunoblots in both peroxisomal and ER fractions recovered from sucrose (Suc) density gradients. Most AtAPX in microsomes (200,000g, 1 h pellet) applied to gradients exhibited a Mg2+-induced shift from a distribution throughout gradients (approximately 18%–40% [w/w] Suc) to ≥42% (w/w) Suc regions of gradients, including pellets, indicative of localization in rough ER vesicles. Immunogold electron microscopy of the latter fractions verified these findings. Further analyses of peroxisomal and rough ER vesicle fractions revealed that AtAPX in both fractions was similarly associated with and located mostly on the cytosolic face of the membranes. Thus, at the steady state, endogenous peroxisomal AtAPX resides at different levels in rough ER and peroxisomes. Collectively, these findings show that rather than being a transiently induced sorting compartment formed in response to overexpressed peroxisomal APX, portions of rough ER (pER) in wild-type cells serve as a constitutive sorting compartment likely involved in posttranslational routing of constitutively synthesized peroxisomal APX. PMID

1. RESTORED GLOVER LOCOMOTIVE #81421 JUST WEST OF MARIETTA SQUARE; ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

1. RESTORED GLOVER LOCOMOTIVE #81421 JUST WEST OF MARIETTA SQUARE; 36 INCH GAUGE 2-6-0, BUILT 1-3-17. RETURNED FOR REPAIR AND RESALE IN 1923, LAST OFFERED FOR SALE IN 1931 FOR $750. - Glover Machine Works, 651 Butler Street, Marietta, Cobb County, GA

NAD(P)H quinone oxidoreductase 1 inhibits the proteasomal degradation of homocysteine-induced endoplasmic reticulum protein

DOE Office of Scientific and Technical Information (OSTI.GOV)

Maeda, Tomoji, E-mail: t-maeda@nichiyaku.ac.jp; Tanabe-Fujimura, Chiaki; Fujita, Yu

2016-05-13

Homocysteine-induced endoplasmic reticulum (ER) protein (Herp) is an ER stress-inducible key regulatory component of ER-associated degradation (ERAD) that has been implicated in insulin hypersecretion in diabetic mouse models. Herp expression is tightly regulated. Additionally, Herp is a highly labile protein and interacts with various proteins, which are characteristic features of ubiquitinated protein. Previously, we reported that ubiquitination is not required for Herp degradation. In addition, we found that the lysine residues of Herp (which are ubiquitinated by E3 ubiquitin ligase) are not sufficient for regulation of Herp degradation. In this study, we found that NAD(P)H quinone oxidoreductase 1 (NQO1)-mediated targetingmore » of Herp to the proteasome was involved in Herp degradation. In addition, we found that Herp protein levels were markedly elevated in synoviolin-null cells. The E3 ubiquitin ligase synoviolin is a central component of ERAD and is involved in the degradation of nuclear factor E2-related factor-2 (Nrf2), which regulates cellular reactive oxygen species. Additionally, NQO1 is a target of Nrf2. Thus, our findings indicated that NQO1 could stabilize Herp protein expression via indirect regulation of synoviolin. -- Highlights: •Herp interacts with NQO1. •NQO1 regulates Herp degradation.« less

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

PubMed

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, P<0.001), inhibited beta cell apoptosis as demonstrated by lower amounts of TUNEL staining nuclei (0.37±0.03 vs. 0.55±0.03, P<0.01) as well as decreased caspase3 activity (1.48±0.11 vs. 2.67±0.13, P<0.01) in islets of diabetic mice compared with untreated diabetic group. Further, vildagliptin 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, P<0.001), ATF-4(activating transcription factor 4) (0.83±0.06 vs. 1.42±0.02, P<0.001) and CHOP(C/EBP homologous protein) (0.07±0.01 vs. 0.16±0.01, P<0.001). Vildagliptin 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.

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

DOE Office of Scientific and Technical Information (OSTI.GOV)

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

2012-10-15

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

HIV-1-associated inflammation and antiretroviral therapy regulate astrocyte endoplasmic reticulum stress responses.

PubMed

Nooka, Shruthi; Ghorpade, Anuja

2017-01-01

Antiretroviral (ARV) therapy (ART) has effectively suppressed the incidence of human immunodeficiency virus (HIV)-associated dementia in HIV-1 positive individuals. However, the prevalence of more subtle forms of neurocognitive dysfunction continues to escalate. Recently, endoplasmic reticulum (ER) stress has been linked to many neurological diseases; yet, its role in HIV/neuroAIDS remains largely unexplored. Furthermore, upregulation of astrocyte elevated gene-1 ( AEG-1 ), a novel HIV-1 inducible gene, along with ER stress markers in a Huntington's disease model, suggests a possible role in HIV-associated ER stress. The current study is focused on unfolded protein responses (UPRs) and AEG-1 regulation in primary human astrocytes exposed to HIV-associated neurocognitive disorders (HAND)-relevant stimuli (HIV-1 virions, inflammation and ARV drugs). Interleukin (IL)-1 β and the nucleoside reverse transcriptase inhibitor abacavir upregulated expression of ER stress markers in human astrocytes, including binding immunoglobulin protein (BiP), C/EBP homologous protein (CHOP), and calnexin. In addition, IL-1 β activated all three well-known UPR pathways: protein kinase RNA-like ER kinase (PERK); activating transcription factor 6 (ATF-6); and inositol-requiring enzyme 1 α (IRE1 α ). AEG-1 upregulation correlated to ER stress and demonstrated astrocyte AEG-1 interaction with the calcium-binding chaperone, calnexin. IL-1 β and abacavir enhanced intracellular calcium signaling in astrocytes in the absence of extracellular calcium, illustrating ER-associated calcium release. Alternatively, calcium evoked in response to HAND-relevant stimuli led to mitochondrial permeability transition pore (mPTP) opening in human astrocytes. Importantly, IL-1 β - and abacavir-induced UPR and mPTP opening were inhibited by the intracellular calcium chelation, indicating the critical role of calcium signaling in HAND-relevant ER stress in astrocytes. In summary, our study highlights that

Apicoplast and Endoplasmic Reticulum Cooperate in Fatty Acid Biosynthesis in Apicomplexan Parasite Toxoplasma gondii*

PubMed Central

Ramakrishnan, Srinivasan; Docampo, Melissa D.; MacRae, James I.; Pujol, François M.; Brooks, Carrie F.; van Dooren, Giel G.; Hiltunen, J. Kalervo; Kastaniotis, Alexander J.; McConville, Malcolm J.; Striepen, Boris

2012-01-01

Apicomplexan parasites are responsible for high impact human diseases such as malaria, toxoplasmosis, and cryptosporidiosis. These obligate intracellular pathogens are dependent on both de novo lipid biosynthesis as well as the uptake of host lipids for biogenesis of parasite membranes. Genome annotations and biochemical studies indicate that apicomplexan parasites can synthesize fatty acids via a number of different biosynthetic pathways that are differentially compartmentalized. However, the relative contribution of each of these biosynthetic pathways to total fatty acid composition of intracellular parasite stages remains poorly defined. Here, we use a combination of genetic, biochemical, and metabolomic approaches to delineate the contribution of fatty acid biosynthetic pathways in Toxoplasma gondii. Metabolic labeling studies with [13C]glucose showed that intracellular tachyzoites synthesized a range of long and very long chain fatty acids (C14:0–26:1). Genetic disruption of the apicoplast-localized type II fatty-acid synthase resulted in greatly reduced synthesis of saturated fatty acids up to 18 carbons long. Ablation of type II fatty-acid synthase activity resulted in reduced intracellular growth that was partially restored by addition of long chain fatty acids. In contrast, synthesis of very long chain fatty acids was primarily dependent on a fatty acid elongation system comprising three elongases, two reductases, and a dehydratase that were localized to the endoplasmic reticulum. The function of these enzymes was confirmed by heterologous expression in yeast. This elongase pathway appears to have a unique role in generating very long unsaturated fatty acids (C26:1) that cannot be salvaged from the host. PMID:22179608

Endoplasmic reticulum stress in amelogenesis imperfecta and phenotypic rescue using 4-phenylbutyrate

PubMed Central

Brookes, Steven J.; Barron, Martin J.; Boot-Handford, Ray; Kirkham, Jennifer; Dixon, Michael J.

2014-01-01

Inherited diseases caused by genetic mutations can arise due to loss of protein function. Alternatively, mutated proteins may mis-fold, impairing endoplasmic reticulum (ER) trafficking, causing ER stress and triggering the unfolded protein response (UPR). The UPR attempts to restore proteostasis but if unsuccessful drives affected cells towards apoptosis. Previously, we reported that in mice, the p.Tyr64His mutation in the enamel extracellular matrix (EEM) protein amelogenin disrupts the secretory pathway in the enamel-forming ameloblasts, resulting in eruption of malformed tooth enamel that phenocopies human amelogenesis imperfecta (AI). Defective amelogenin post-secretory self-assembly and processing within the developing EEM has been suggested to underlie the pathogenesis of X chromosome-linked AI. Here, we challenge this concept by showing that AI pathogenesis associated with the p.Tyr64His amelogenin mutation involves ameloblast apoptosis induced by ER stress. Furthermore, we show that 4-phenylbutyrate can rescue the enamel phenotype in affected female mice by promoting cell survival over apoptosis such that they are able to complete enamel formation despite the presence of the mutation, offering a potential therapeutic option for patients with this form of AI and emphasizing the importance of ER stress in the pathogenesis of this inherited conformational disease. PMID:24362885

Endoplasmic reticulum stress in amelogenesis imperfecta and phenotypic rescue using 4-phenylbutyrate.

PubMed

Brookes, Steven J; Barron, Martin J; Boot-Handford, Ray; Kirkham, Jennifer; Dixon, Michael J

2014-05-01

Inherited diseases caused by genetic mutations can arise due to loss of protein function. Alternatively, mutated proteins may mis-fold, impairing endoplasmic reticulum (ER) trafficking, causing ER stress and triggering the unfolded protein response (UPR). The UPR attempts to restore proteostasis but if unsuccessful drives affected cells towards apoptosis. Previously, we reported that in mice, the p.Tyr64His mutation in the enamel extracellular matrix (EEM) protein amelogenin disrupts the secretory pathway in the enamel-forming ameloblasts, resulting in eruption of malformed tooth enamel that phenocopies human amelogenesis imperfecta (AI). Defective amelogenin post-secretory self-assembly and processing within the developing EEM has been suggested to underlie the pathogenesis of X chromosome-linked AI. Here, we challenge this concept by showing that AI pathogenesis associated with the p.Tyr64His amelogenin mutation involves ameloblast apoptosis induced by ER stress. Furthermore, we show that 4-phenylbutyrate can rescue the enamel phenotype in affected female mice by promoting cell survival over apoptosis such that they are able to complete enamel formation despite the presence of the mutation, offering a potential therapeutic option for patients with this form of AI and emphasizing the importance of ER stress in the pathogenesis of this inherited conformational disease.

Effect of the low- versus high-intensity exercise training on endoplasmic reticulum stress and GLP-1 in adolescents with type 2 diabetes mellitus.

PubMed

Lee, Sung Soo; Yoo, Jae Ho; So, Yong Seok

2015-10-01

[Purpose] The primary objective of this study was to investigate the effect of low-intensity exercise training compare with high-intensity exercise training on endoplasmic reticulum stress and glucagon-like peptide-1 in adolescents with type 2 diabetes mellitus. [Subjects and Methods] The low-intensity exercise training group performed aerobic exercise training at an intensity of ≤ 45% of the heart rate reserve. The high-intensity interval exercise training group performed interval exercise training at an intensity of ≥ 80% of the heart rate reserve. The exercise-related energy consumption was determined for both groups on a per-week basis (1,200 kcal/week). [Results] Both groups showed improvement in the glucose-regulated protein 78 and dipeptidyl peptidase-4, but the size of the between-group effect was not statistically significant. The high-intensity interval exercise training group showed a significant reduction in percentage body fat. The C-peptide level increased after the 12-weeks programs and was significantly different, between the groups. Fasting glucose, insulin resistance in the fasting state according to homeostasis model assessment, and leptin decreased after the 12-weeks exercise program and were significantly different between the groups, and glucagon-like peptide-1 increased after the 12-week exercise programs and was significantly different between the groups. [Conclusion] In conclusion high-intensity interval exercise training, as defined in this study, may lead to improvements in body composition, glycemic control, endoplasmic reticulum stress, and the glucagon-like peptide-1 in adolescents with type 2 diabetes mellitus.

Effect of the low- versus high-intensity exercise training on endoplasmic reticulum stress and GLP-1 in adolescents with type 2 diabetes mellitus

PubMed Central

Lee, Sung Soo; Yoo, Jae Ho; So, Yong Seok

2015-01-01

[Purpose] The primary objective of this study was to investigate the effect of low-intensity exercise training compare with high-intensity exercise training on endoplasmic reticulum stress and glucagon-like peptide-1 in adolescents with type 2 diabetes mellitus. [Subjects and Methods] The low-intensity exercise training group performed aerobic exercise training at an intensity of ≤ 45% of the heart rate reserve. The high-intensity interval exercise training group performed interval exercise training at an intensity of ≥ 80% of the heart rate reserve. The exercise-related energy consumption was determined for both groups on a per-week basis (1,200 kcal/week). [Results] Both groups showed improvement in the glucose-regulated protein 78 and dipeptidyl peptidase-4, but the size of the between-group effect was not statistically significant. The high-intensity interval exercise training group showed a significant reduction in percentage body fat. The C-peptide level increased after the 12-weeks programs and was significantly different, between the groups. Fasting glucose, insulin resistance in the fasting state according to homeostasis model assessment, and leptin decreased after the 12-weeks exercise program and were significantly different between the groups, and glucagon-like peptide-1 increased after the 12-week exercise programs and was significantly different between the groups. [Conclusion] In conclusion high-intensity interval exercise training, as defined in this study, may lead to improvements in body composition, glycemic control, endoplasmic reticulum stress, and the glucagon-like peptide-1 in adolescents with type 2 diabetes mellitus. PMID:26644644

Deficiency of Suppressor Enhancer Lin12 1 Like (SEL1L) in Mice Leads to Systemic Endoplasmic Reticulum Stress and Embryonic Lethality*

PubMed Central

Francisco, Adam B.; Singh, Rajni; Li, Shuai; Vani, Anish K.; Yang, Liu; Munroe, Robert J.; Diaferia, Giuseppe; Cardano, Marina; Biunno, Ida; Qi, Ling; Schimenti, John C.; Long, Qiaoming

2010-01-01

Stress in the endoplasmic reticulum (ER) plays an important causal role in the pathogenesis of several chronic diseases such as Alzheimer, Parkinson, and diabetes mellitus. Insight into the genetic determinants responsible for ER homeostasis will greatly facilitate the development of therapeutic strategies for the treatment of these debilitating diseases. Suppressor enhancer Lin12 1 like (SEL1L) is an ER membrane protein and was thought to be involved in the quality control of secreted proteins. Here we show that the mice homozygous mutant for SEL1L were embryonic lethal. Electron microscopy studies revealed a severely dilated ER in the fetal liver of mutant embryos, indicative of alteration in ER homeostasis. Consistent with this, several ER stress responsive genes were significantly up-regulated in the mutant embryos. Mouse embryonic fibroblast cells deficient in SEL1L exhibited activated unfolded protein response at the basal state, impaired ER-associated protein degradation, and reduced protein secretion. Furthermore, markedly increased apoptosis was observed in the forebrain and dorsal root ganglions of mutant embryos. Taken together, our results demonstrate an essential role for SEL1L in protein quality control during mouse embryonic development. PMID:20197277

Deficiency of suppressor enhancer Lin12 1 like (SEL1L) in mice leads to systemic endoplasmic reticulum stress and embryonic lethality.

PubMed

Francisco, Adam B; Singh, Rajni; Li, Shuai; Vani, Anish K; Yang, Liu; Munroe, Robert J; Diaferia, Giuseppe; Cardano, Marina; Biunno, Ida; Qi, Ling; Schimenti, John C; Long, Qiaoming

2010-04-30

Stress in the endoplasmic reticulum (ER) plays an important causal role in the pathogenesis of several chronic diseases such as Alzheimer, Parkinson, and diabetes mellitus. Insight into the genetic determinants responsible for ER homeostasis will greatly facilitate the development of therapeutic strategies for the treatment of these debilitating diseases. Suppressor enhancer Lin12 1 like (SEL1L) is an ER membrane protein and was thought to be involved in the quality control of secreted proteins. Here we show that the mice homozygous mutant for SEL1L were embryonic lethal. Electron microscopy studies revealed a severely dilated ER in the fetal liver of mutant embryos, indicative of alteration in ER homeostasis. Consistent with this, several ER stress responsive genes were significantly up-regulated in the mutant embryos. Mouse embryonic fibroblast cells deficient in SEL1L exhibited activated unfolded protein response at the basal state, impaired ER-associated protein degradation, and reduced protein secretion. Furthermore, markedly increased apoptosis was observed in the forebrain and dorsal root ganglions of mutant embryos. Taken together, our results demonstrate an essential role for SEL1L in protein quality control during mouse embryonic development.

Ursolic acid, a natural pentacyclic triterpenoid, inhibits intracellular trafficking of proteins and induces accumulation of intercellular adhesion molecule-1 linked to high-mannose-type glycans in the endoplasmic reticulum

PubMed Central

Mitsuda, Satoshi; Yokomichi, Tomonobu; Yokoigawa, Junpei; Kataoka, Takao

2014-01-01

Ursolic acid (3β-hydroxy-urs-12-en-28-oic acid) is a natural pentacyclic triterpenoid that is present in many plants, including medicinal herbs, and foods. Ursolic acid was initially identified as an inhibitor of the expression of intercellular adhesion molecule-1 (ICAM-1) in response to interleukin-1α (IL-1α). We report here a novel biological activity: ursolic acid inhibits intracellular trafficking of proteins. Ursolic acid markedly inhibited the IL-1α-induced cell-surface ICAM-1 expression in human cancer cell lines and human umbilical vein endothelial cells. By contrast, ursolic acid exerted weak inhibitory effects on the IL-1α-induced ICAM-1 expression at the protein level. Surprisingly, we found that ursolic acid decreased the apparent molecular weight of ICAM-1 and altered the structures of N-linked oligosaccharides bound to ICAM-1. Ursolic acid induced the accumulation of ICAM-1 in the endoplasmic reticulum, which was linked mainly to high-mannose-type glycans. Moreover, in ursolic-acid-treated cells, the Golgi apparatus was fragmented into pieces and distributed over the cells. Thus, our results reveal that ursolic acid inhibits intracellular trafficking of proteins and induces the accumulation of ICAM-1 linked to high-mannose-type glycans in the endoplasmic reticulum. PMID:24649404

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

PubMed

Márquez, Saioa; Fernández, José Javier; Terán-Cabanillas, Eli; Herrero, Carmen; Alonso, Sara; Azogil, Alicia; Montero, Olimpio; Iwawaki, Takao; Cubillos-Ruiz, Juan R; Fernández, Nieves; Crespo, Mariano Sánchez

2017-01-01

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.

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

PubMed Central

Márquez, Saioa; Fernández, José Javier; Terán-Cabanillas, Eli; Herrero, Carmen; Alonso, Sara; Azogil, Alicia; Montero, Olimpio; Iwawaki, Takao; Cubillos-Ruiz, Juan R.; Fernández, Nieves; Crespo, Mariano Sánchez

2017-01-01

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. PMID:28674530

Sphingosin-1-phosphate Receptor 1: a Potential Target to Inhibit Neuroinflammation and Restore the Sphingosin-1-phosphate Metabolism.

PubMed

Kolahdooz, Zeynab; Nasoohi, Sanaz; Asle-Rousta, Masoumeh; Ahmadiani, Abolhassan; Dargahi, Leila

2015-05-01

Recent evidence suggests that an extreme shift may occur in sphingosine metabolism in neuroinflammatory contexts. Sphingosine 1-phosphate (S1P)-metabolizing enzymes (SMEs) regulate the level of S1P. We recently found that FTY720, a S1P analogue, and SEW2871, a selective S1P receptor 1 (S1P1) agonist, provide protection against neural damage and memory deficit in amyloid beta (Aβ)-injected animals. This study aimed to evaluate the effects of these two analogues on the expression of SMEs as well as their anti-inflammatory roles. Rats were treated with intracerebral lipopolysaccharide (LPS) or Aβ. Memory impairment was assessed by Morris water maze and the effects of drugs on SMEs as well as inflammatory markers, TNF- α and COX-II, were determined by immunoblotting. Aβ and LPS differentially altered the expression profile of SMEs. In Aβ-injected animals, FTY720 and SEW2871 treatments exerted anti-inflammatory effects and restored the expression profile of SMEs, in parallel to our previous findings. In LPS animals however, in spite of anti-inflammatory effects of the two analogues, only FTY720 restored the levels of SMEs and prevented memory deficit. The observed ameliorating effects of FTY720 and SEW7821 can be partly attributed to the interruption of the vicious cycle of abnormal S1P metabolism and neuro-inflammation. The close imitation of the FTY720 effects by SW2871 in Aβ-induced neuro-inflammation may highlight the attractive role of S1P1 as a potential target to restore S1P metabolism and inhibit inflammatory processes.

An iterative algorithm for L1-TV constrained regularization in image restoration

NASA Astrophysics Data System (ADS)

Chen, K.; Loli Piccolomini, E.; Zama, F.

2015-11-01

We consider the problem of restoring blurred images affected by impulsive noise. The adopted method restores the images by solving a sequence of constrained minimization problems where the data fidelity function is the ℓ1 norm of the residual and the constraint, chosen as the image Total Variation, is automatically adapted to improve the quality of the restored images. Although this approach is general, we report here the case of vectorial images where the blurring model involves contributions from the different image channels (cross channel blur). A computationally convenient extension of the Total Variation function to vectorial images is used and the results reported show that this approach is efficient for recovering nearly optimal images.

The metabolomic signature of Leber's hereditary optic neuropathy reveals endoplasmic reticulum stress.

PubMed

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

the greater expression of C/EBP homologous protein and the increased XBP1 splicing, in fibroblasts from affected patients, all these changes being reversed by the endoplasmic reticulum stress inhibitor, TUDCA (tauroursodeoxycholic acid). Thus, our metabolomic analysis reveals a pharmacologically-reversible endoplasmic reticulum stress in complex I-related Leber's hereditary optic neuropathy fibroblasts, a finding that may open up new therapeutic perspectives for the treatment of Leber's hereditary optic neuropathy with endoplasmic reticulum-targeting drugs. © The Author (2016). Published by Oxford University Press on behalf of the Guarantors of Brain. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Agonist-activated Ca2+ influx occurs at stable plasma membrane and endoplasmic reticulum junctions

PubMed Central

Treves, Susan; Vukcevic, Mirko; Griesser, Johanna; Armstrong, Clara-Franzini; Zhu, Michael X.; Zorzato, Fancesco

2010-01-01

Junctate is a 33 kDa integral protein of sarco(endo)plasmic reticulum membranes that forms a macromolecular complex with inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] receptors and TRPC3 channels. TIRF microscopy shows that junctate enhances the number of fluorescent puncta on the plasma membrane. The size and distribution of these puncta are not affected by the addition of agonists that mobilize Ca2+ from Ins(1,4,5)P3-sensitive stores. Puncta are associated with a significantly larger number of peripheral junctions between endoplasmic reticulum and plasma membrane, which are further enhanced upon stable co-expression of junctate and TRPC3. The gap between the membranes of peripheral junctions is bridged by regularly spaced electron-dense structures of 10 nm. Ins(1,4,5)P3 inhibits the interaction of the cytoplasmic N-terminus of junctate with the ligand-binding domain of the Ins(1,4,5)P3 receptor. Furthermore, Ca2+ influx evoked by activation of Ins(1,4,5)P3 receptors is increased where puncta are located. We conclude that stable peripheral junctions between the plasma membrane and endoplasmic reticulum are the anatomical sites of agonist-activated Ca2+ entry. PMID:21062895

1. N elevation of Tender Frame Shop showing restored, standing ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

1. N elevation of Tender Frame Shop showing restored, standing seam metal roof. - Central of Georgia Railway, Savannah Repair Shops & Terminal Facilities, Tender Frame Shop, Bounded by West Broad, Jones, West Boundary & Hull Streets, Savannah, Chatham County, GA

Bi-polarized translation of ascidian maternal mRNA determinant pem-1 associated with regulators of the translation machinery on cortical Endoplasmic Reticulum (cER).

PubMed

Paix, Alexandre; Le Nguyen, Phuong Ngan; Sardet, Christian

2011-09-01

Polarized cortical mRNA determinants such as maternal macho-1 and pem-1 in ascidians, like budding yeast mating factor ASH1 reside on the cER-mRNA domain a subdomain of cortical Endoplasmic Reticulum(ER) and are translated in its vicinity. Using high resolution imaging and isolated cortical fragments prepared from eggs and embryos we now find that macho-1 and pem-1 RNAs co-localize with phospho-protein regulators of translation initiation (MnK/4EBP/S6K). Translation of cortical pem-1 RNA follows its bi-polarized relocalization. About 10 min after fertilization or artificial activation with a calcium ionophore, PEM1 protein is detected in the vegetal cortex in the vicinity of pem-1 RNA. About 40 min after fertilization-when pem-1 RNA and P-MnK move to the posterior pole-PEM1 protein remains in place forming a network of cortical patches anchored at the level of the zygote plasma membrane before disappearing. Cortical PEM1 protein is detected again at the 4 cell stage in the posterior centrosome attracting body (CAB) region where the cER-mRNA domain harboring pem-1/P-MnK/P-4EBP/P-S6K is concentrated. Bi-polarized PEM1 protein signals are not detected when pem-1 morpholinos are injected into eggs or zygotes or when MnK is inhibited. We propose that localized translation of the pem-1 RNA determinant is triggered by the fertilization/calcium wave and that the process is controlled by phospho-protein regulators of translation initiation co-localized with the RNA determinant on a sub-domain of the cortical Endoplasmic Reticulum. Copyright © 2011 Elsevier Inc. All rights reserved.

Requirement of Zinc Transporter SLC39A7/ZIP7 for Dermal Development to Fine-Tune Endoplasmic Reticulum Function by Regulating Protein Disulfide Isomerase.

PubMed

Bin, Bum-Ho; Bhin, Jinhyuk; Seo, Juyeon; Kim, Se-Young; Lee, Eunyoung; Park, Kyuhee; Choi, Dong-Hwa; Takagishi, Teruhisa; Hara, Takafumi; Hwang, Daehee; Koseki, Haruhiko; Asada, Yoshinobu; Shimoda, Shinji; Mishima, Kenji; Fukada, Toshiyuki

2017-08-01

Skin is the first area that manifests zinc deficiency. However, the molecular mechanisms by which zinc homeostasis affects skin development remain largely unknown. Here, we show that zinc-regulation transporter-/iron-regulation transporter-like protein 7 (ZIP7) localized to the endoplasmic reticulum plays critical roles in connective tissue development. Mice lacking the Slc39a7/Zip7 gene in collagen 1-expressing tissue exhibited dermal dysplasia. Ablation of ZIP7 in mesenchymal stem cells inhibited cell proliferation thereby preventing proper dermis formation, indicating that ZIP7 is required for dermal development. We also found that mesenchymal stem cells lacking ZIP7 accumulated zinc in the endoplasmic reticulum, which triggered zinc-dependent aggregation and inhibition of protein disulfide isomerase, leading to endoplasmic reticulum dysfunction. These results suggest that ZIP7 is necessary for endoplasmic reticulum function in mesenchymal stem cells and, as such, is essential for dermal development. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

The yeast p5 type ATPase, spf1, regulates manganese transport into the endoplasmic reticulum.

PubMed

Cohen, Yifat; Megyeri, Márton; Chen, Oscar C W; Condomitti, Giuseppe; Riezman, Isabelle; Loizides-Mangold, Ursula; Abdul-Sada, Alaa; Rimon, Nitzan; Riezman, Howard; Platt, Frances M; Futerman, Anthony H; Schuldiner, Maya

2013-01-01

The endoplasmic reticulum (ER) is a large, multifunctional and essential organelle. Despite intense research, the function of more than a third of ER proteins remains unknown even in the well-studied model organism Saccharomyces cerevisiae. One such protein is Spf1, which is a highly conserved, ER localized, putative P-type ATPase. Deletion of SPF1 causes a wide variety of phenotypes including severe ER stress suggesting that this protein is essential for the normal function of the ER. The closest homologue of Spf1 is the vacuolar P-type ATPase Ypk9 that influences Mn(2+) homeostasis. However in vitro reconstitution assays with Spf1 have not yielded insight into its transport specificity. Here we took an in vivo approach to detect the direct and indirect effects of deleting SPF1. We found a specific reduction in the luminal concentration of Mn(2+) in ∆spf1 cells and an increase following it's overexpression. In agreement with the observed loss of luminal Mn(2+) we could observe concurrent reduction in many Mn(2+)-related process in the ER lumen. Conversely, cytosolic Mn(2+)-dependent processes were increased. Together, these data support a role for Spf1p in Mn(2+) transport in the cell. We also demonstrate that the human sequence homologue, ATP13A1, is a functionally conserved orthologue. Since ATP13A1 is highly expressed in developing neuronal tissues and in the brain, this should help in the study of Mn(2+)-dependent neurological disorders.

Induction of endoplasmic reticulum calcium pump expression during early leukemic B cell differentiation.

PubMed

Aït Ghezali, Lamia; Arbabian, Atousa; Roudot, Hervé; Brouland, Jean-Philippe; Baran-Marszak, Fanny; Salvaris, Evelyn; Boyd, Andrew; Drexler, Hans G; Enyedi, Agnes; Letestu, Remi; Varin-Blank, Nadine; Papp, Bela

2017-06-26

Endoplasmic reticulum (ER) calcium storage and release play important roles in B lymphocyte maturation, survival, antigen-dependent cell activation and immunoglobulin synthesis. Calcium is accumulated in the endoplasmic reticulum (ER) by Sarco/Endoplasmic Reticulum Calcium ATPases (SERCA enzymes). Because lymphocyte function is critically dependent on SERCA activity, it is important to understand qualitative and quantitative changes of SERCA protein expression that occur during B lymphoid differentiation and leukemogenesis. In this work we investigated the modulation of SERCA expression during the pharmacologically induced differentiation of leukemic precursor B lymphoblast cell lines that carry the E2A-PBX1 fusion oncoprotein. Changes of SERCA levels during differentiation were determined and compared to those of established early B lymphoid differentiation markers. SERCA expression of the cells was compared to that of mature B cell lines as well, and the effect of the direct inhibition of SERCA-dependent calcium transport on the differentiation process was investigated. We show that E2A-PBX1 + leukemia cells simultaneously express SERCA2 and SERCA3-type calcium pumps; however, their SERCA3 expression is markedly inferior to that of mature B cells. Activation of protein kinase C enzymes by phorbol ester leads to phenotypic differentiation of the cells, and this is accompanied by the induction of SERCA3 expression. Direct pharmacological inhibition of SERCA-dependent calcium transport during phorbol ester treatment interferes with the differentiation process. These data show that the calcium pump composition of the ER is concurrent with increased SERCA3 expression during the differentiation of precursor B acute lymphoblastic leukemia cells, that a cross-talk exists between SERCA function and the control of differentiation, and that SERCA3 may constitute an interesting new marker for the study of early B cell phenotype.

Traffic of Human α-Mannosidase in Plant Cells Suggests the Presence of a New Endoplasmic Reticulum-to-Vacuole Pathway without Involving the Golgi Complex1[W

PubMed Central

De Marchis, Francesca; Bellucci, Michele; Pompa, Andrea

2013-01-01

The transport of secretory proteins from the endoplasmic reticulum to the vacuole requires sorting signals as well as specific transport mechanisms. This work is focused on the transport in transgenic tobacco (Nicotiana tabacum) plants of a human α-mannosidase, MAN2B1, which is a lysosomal enzyme involved in the turnover of N-linked glycoproteins and can be used in enzyme replacement therapy. Although ubiquitously expressed, α-mannosidases are targeted to lysosomes or vacuoles through different mechanisms according to the organisms in which these proteins are produced. In tobacco cells, MAN2B1 reaches the vacuole even in the absence of mannose-6-phosphate receptors, which are responsible for its transport in animal cells. We report that MAN2B1 is targeted to the vacuole without passing through the Golgi complex. In addition, a vacuolar targeting signal that is recognized in plant cells is located in the MAN2B1 amino-terminal region. Indeed, when this amino-terminal domain is removed, the protein is retained in the endoplasmic reticulum. Moreover, when this domain is added to a plant-secreted protein, the resulting fusion protein is partially redirected to the vacuole. These results strongly suggest the existence in plants of a new type of vacuolar traffic that can be used by leaf cells to transport vacuolar proteins. PMID:23449646

Translocation of the ABC transporter ABCD4 from the endoplasmic reticulum to lysosomes requires the escort protein LMBD1.

PubMed

Kawaguchi, Kosuke; Okamoto, Takumi; Morita, Masashi; Imanaka, Tsuneo

2016-07-26

We previously demonstrated that ABCD4 does not localize to peroxisomes but rather, the endoplasmic reticulum (ER), because it lacks the NH2-terminal hydrophilic region required for peroxisomal targeting. It was recently reported that mutations in ABCD4 result in a failure to release vitamin B12 from lysosomes. A similar phenotype is caused by mutations in LMBRD1, which encodes the lysosomal membrane protein LMBD1. These findings suggested to us that ABCD4 translocated from the ER to lysosomes in association with LMBD1. In this report, it is demonstrated that ABCD4 interacts with LMBD1 and then localizes to lysosomes, and this translocation depends on the lysosomal targeting ability of LMBD1. Furthermore, endogenous ABCD4 was localized to both lysosomes and the ER, and its lysosomal localization was disturbed by knockout of LMBRD1. To the best of our knowledge, this is the first report demonstrating that the subcellular localization of the ABC transporter is determined by its association with an adaptor protein.

Installation Restoration Program. Confirmation/Quantification Stage 1. Phase 2

DTIC Science & Technology

1985-03-07

INSTALLATION RESTORATION PROGRAM i0 PHASE II - CONFIRMATION/QUANTIFICATION 0STAGE 1 KIRTLAND AFB KIRTLAND AFB, NEW MEXICO 87117 IIl PREPARED BY SCIENCE...APPLICATIONS INTERNATIONAL CORPORATION 505 MARQUETTE NW, SUITE 1200 ALBUQUERQUE, NEW MEXICO 871021 5MARCH 1985 FINAL REPORT FROM FEB 1983 TO MAR 1985...QUANTIFICATION STAGE 1 i FINAL REPORT FOR IKIRTLAND AFB KIRTLAND AFB, NEW MEXICO 87117U HEADQUARTERS MILITARY AIRLIFT COMMAND COMMAND SURGEON’S OFFICE (HQ MAC

GOLGI TRANSPORT 1B Regulates Protein Export from the Endoplasmic Reticulum in Rice Endosperm Cells[OPEN

PubMed Central

Liu, Feng; Wang, Yunlong; Liu, Xi; Wang, Di; Zhu, Xiaopin; Jing, Ruonan; Wu, Mingming; Hao, Yuanyuan; Jiang, Ling; Wang, Chunming

2016-01-01

Coat protein complex II (COPII) mediates the first step of anterograde transport of newly synthesized proteins from the endoplasmic reticulum (ER) to other endomembrane compartments in eukaryotes. A group of evolutionarily conserved proteins (Sar1, Sec23, Sec24, Sec13, and Sec31) constitutes the basic COPII coat machinery; however, the details of how the COPII coat assembly is regulated remain unclear. Here, we report a protein transport mutant of rice (Oryza sativa), named glutelin precursor accumulation4 (gpa4), which accumulates 57-kD glutelin precursors and forms two types of ER-derived abnormal structures. GPA4 encodes the evolutionarily conserved membrane protein GOT1B (also known as GLUP2), homologous to the Saccharomyces cerevisiae GOT1p. The rice GOT1B protein colocalizes with Arabidopsis thaliana Sar1b at Golgi-associated ER exit sites (ERESs) when they are coexpressed in Nicotiana benthamiana. Moreover, GOT1B physically interacts with rice Sec23, and both proteins are present in the same complex(es) with rice Sar1b. The distribution of rice Sar1 in the endomembrane system, its association with rice Sec23c, and the ERES organization pattern are significantly altered in the gpa4 mutant. Taken together, our results suggest that GOT1B plays an important role in mediating COPII vesicle formation at ERESs, thus facilitating anterograde transport of secretory proteins in plant cells. PMID:27803308

GLP-1 promotes mitochondrial metabolism in vascular smooth muscle cells by enhancing endoplasmic reticulum-mitochondria coupling.

PubMed

Morales, Pablo E; Torres, Gloria; Sotomayor-Flores, Cristian; Peña-Oyarzún, Daniel; Rivera-Mejías, Pablo; Paredes, Felipe; Chiong, Mario

2014-03-28

Incretin GLP-1 has important metabolic effects on several tissues, mainly through the regulation of glucose uptake and usage. One mechanism for increasing cell metabolism is modulating endoplasmic reticulum (ER)-mitochondria communication, as it allows for a more efficient transfer of Ca(2+) into the mitochondria, thereby increasing activity. Control of glucose metabolism is essential for proper vascular smooth muscle cell (VSMC) function. GLP-1 has been shown to produce varied metabolic actions, but whether it regulates glucose metabolism in VSMC remains unknown. In this report, we show that GLP-1 increases mitochondrial activity in the aortic cell line A7r5 by increasing ER-mitochondria coupling. GLP-1 increases intracellular glucose and diminishes glucose uptake without altering glycogen content. ATP, mitochondrial potential and oxygen consumption increase at 3h of GLP-1 treatment, paralleled by increased Ca(2+) transfer from the ER to the mitochondria. Furthermore, GLP-1 increases levels of Mitofusin-2 (Mfn2), an ER-mitochondria tethering protein, via a PKA-dependent mechanism. Accordingly, PKA inhibition and Mfn2 down-regulation prevented mitochondrial Ca(2+) increases in GLP-1 treated cells. Inhibiting both Ca(2+) release from the ER and Ca(2+) entry into mitochondria as well as diminishing Mfn2 levels blunted the increase in mitochondrial activity in response to GLP-1. Altogether, these results strongly suggest that GLP-1 increases ER-mitochondria communication in VSMC, resulting in higher mitochondrial activity. Copyright © 2014 Elsevier Inc. All rights reserved.

Tributyltin induces apoptotic signaling in hepatocytes through pathways involving the endoplasmic reticulum and mitochondria

DOE Office of Scientific and Technical Information (OSTI.GOV)

Grondin, Melanie; Marion, Michel; Denizeau, Francine

2007-07-01

Tri-n-butyltin is a widespread environmental toxicant, which accumulates in the liver. This study investigates whether tri-n-butyltin induces pro-apoptotic signaling in rat liver hepatocytes through pathways involving the endoplasmic reticulum and mitochondria. Tri-n-butyltin activated the endoplasmic reticulum pathway of apoptosis, which was demonstrated by the activation of the protease calpain, its translocation to the plasma membrane, followed by cleavage of the calpain substrates, cytoskeletal protein vinculin, and caspase-12. Caspase-12 is localized to the cytoplasmic side of the endoplasmic reticulum and is involved in apoptosis mediated by the endoplasmic reticulum. Tri-n-butyltin also caused translocation of the pro-apoptotic proteins Bax and Bad frommore » the cytosol to mitochondria, as well as changes in mitochondrial membrane permeability, events which can activate the mitochondrial death pathway. Tri-n-butyltin induced downstream apoptotic events in rat hepatocytes at the nuclear level, detected by chromatin condensation and by confocal microscopy using acridine orange. We investigated whether the tri-n-butyltin-induced pro-apoptotic events in hepatocytes could be linked to perturbation of intracellular calcium homeostasis, using confocal microscopy. Tri-n-butyltin caused changes in intracellular calcium distribution, which were similar to those induced by thapsigargin. Calcium was released from a subcellular compartment, which is likely to be the endoplasmic reticulum, into the cytosol. Cytosolic acidification, which is known to trigger apoptosis, also occurred and involved the Cl{sup -}/HCO{sub 3} {sup -} exchanger. Pro-apoptotic events in hepatocytes were inhibited by the calcium chelator, Bapta-AM, and by a calpain inhibitor, which suggests that changes in intracellular calcium homeostasis are involved in tri-n-butyltin-induced apoptotic signaling in rat hepatocytes.« less

Signal sequence-independent targeting of MID2 mRNA to the endoplasmic reticulum by the yeast RNA-binding protein Khd1p.

PubMed

Syed, Muhammad Ibrahim; Moorthy, Balaji T; Jenner, Andreas; Fetka, Ingrid; Jansen, Ralf-Peter

2018-05-17

Localization of mRNAs depends on specific RNA-binding proteins (RBPs) and critically contributes not only to cell polarization but also to basal cell function. The yeast RBP Khd1p binds to several hundred mRNAs, the majority of which encodes secreted or membrane proteins. We demonstrate that a subfraction of Khd1p associates with artificial liposomes and endoplasmic reticulum (ER), and that Khd1p endomembrane association is partially dependent on its binding to RNA. ER targeting of at least two mRNAs, MID2 and SLG1/WSC1, requires KHD1 but is independent of their translation. Together, our results suggest interdependence of Khd1p and mRNA for their targeting to the ER and presents additional evidence for signal sequence-independent, RBP-mediated mRNA targeting. © 2018 Federation of European Biochemical Societies.

Central leptin action on euglycemia restoration in type 1 diabetes: Restraining responses normally induced by fasting?

PubMed

Xu, Yuanzhong; Tong, Qingchun

2017-07-01

Leptin monotherapy is sufficient to restore euglycemia in insulinopenic type 1 diabetes (T1D), yet the underlying mechanism remains poorly understood. Accumulating evidence demonstrates that the brain mediates the leptin action on euglycemia restoration. Here, we first review evidence supporting that symptoms in T1D resemble an uncontrolled response to fasting. Then, we discuss recent research progress on brain neurons and their neurotransmitters that potentially mediate the leptin action. Finally, peripheral effective pathways, which are normally involved in fasting responses and associated with leptin action on euglycemia restoration in T1D, will also be discussed. This summary complements several previous excellent reviews on this topic (Meek and Morton, 2016; Perry et al., 2016; Fujikawa and Coppari, 2015). A deep understanding of neurocircuitry and the peripheral effective pathways that mediate the leptin action on euglycemia restoration will likely lead to novel targets for an insulin-independent therapeutics against T1D. Copyright © 2016 Elsevier Ltd. All rights reserved.

A transgenic zebrafish model for monitoring xbp1 splicing and endoplasmic reticulum stress in vivo.

PubMed

Li, Junling; Chen, Zhiliang; Gao, Lian-Yong; Colorni, Angelo; Ucko, Michal; Fang, Shengyun; Du, Shao Jun

2015-08-01

Accumulation of misfolded or unfolded proteins in the endoplasmic reticulum (ER) triggers ER stress that initiates unfolded protein response (UPR). XBP1 is a transcription factor that mediates one of the key signaling pathways of UPR to cope with ER stress through regulating gene expression. Activation of XBP1 involves an unconventional mRNA splicing catalyzed by IRE1 endonuclease that removes an internal 26 nucleotides from xbp1 mRNA transcripts in the cytoplasm. Researchers have taken advantage of this unique activation mechanism to monitor XBP1 activation, thereby UPR, in cell culture and transgenic models. Here we report a Tg(ef1α:xbp1δ-gfp) transgenic zebrafish line to monitor XBP1 activation using GFP as a reporter especially in zebrafish oocytes and developing embryos. The Tg(ef1α:xbp1δ-gfp) transgene was constructed using part of the zebrafish xbp1 cDNA containing the splicing element. ER stress induced splicing results in the cDNA encoding a GFP-tagged partial XBP1 without the transactivation activation domain (XBP1Δ-GFP). The results showed that xbp1 transcripts mainly exist as the spliced active isoform in unfertilized oocytes and zebrafish embryos prior to zygotic gene activation at 3 hours post fertilization. A strong GFP expression was observed in unfertilized oocytes, eyes, brain and skeletal muscle in addition to a weak expression in the hatching gland. Incubation of transgenic zebrafish embryos with (dithiothreitol) DTT significantly induced XBP1Δ-GFP expression. Collectively, these studies unveil the presence of maternal xbp1 splicing in zebrafish oocytes, fertilized eggs and early stage embryos. The Tg(ef1α:xbp1δ-gfp) transgenic zebrafish provides a useful model for in vivo monitoring xbp1 splicing during development and under ER stress conditions. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

26 CFR 1.1341-1 - Restoration of amounts received or accrued under claim of right.

Code of Federal Regulations, 2010 CFR

2010-04-01

... TREASURY (CONTINUED) INCOME TAX (CONTINUED) INCOME TAXES Claim of Right § 1.1341-1 Restoration of amounts... income for a prior taxable year (or years) under a claim of right, the tax imposed by chapter 1 of the Internal Revenue Code of 1954 for the taxable year shall be the tax provided in paragraph (b) of this...

Alteration of the proteostasis network of plant cells promotes the post-endoplasmic reticulum trafficking of recombinant mutant (L444P) human β-glucocerebrosidase

PubMed Central

Babajani, Gholamreza; Kermode, Allison R

2014-01-01

Gaucher disease is a prevalent lysosomal storage disease characterized by a deficiency in the activity of lysosomal acid β-glucosidase (glucocerebrosidase, GCase, EC 3.2.1.45). One of the most prevalent disease-causing mutations in humans is a L444P missense mutation in the GCase protein, which results in its disrupted folding in the endoplasmic reticulum (ER) and impaired post-ER trafficking. To determine whether the post-ER trafficking of this severely malfolded protein can be restored, we expressed the mutant L444P GCase as a recombinant protein in transgenic tobacco (Nicotiana tabacum L. cv Bright Yellow 2 [BY2]) cells, in which the GCase variant was equipped with a plant signal peptide to allow for secretion upon rescued trafficking out of the ER. The recombinant L444P mutant GCase was retained in the plant endoplasmic reticulum (ER). Kifunensine and Eeyarestatin I, both inhibitors of ER-associated degradation (ERAD), and the proteostasis regulators, celastrol and MG-132, increased the steady-state levels of the mutant protein inside the plant cells and further promoted the post-ER trafficking of L444P GCase, as indicated by endoglycosidase-H sensitivity- and secretion- analyses. Transcript profiling of genes encoding ER-molecular chaperones, ER stress responsive proteins, and cytoplasmic heat shock response proteins, revealed insignificant or only very modest changes in response to the ERAD inhibitors and proteostasis regulators. An exception was the marked response to celastrol which reduced the steady-state levels of cytoplasmic HSP90 transcripts and protein. As HSP90 participates in the targeting of misfolded proteins to the proteasome pathway, its down-modulation in response to celastrol may partly account for the mechanism of improved homeostasis of L444P GCase mediated by this triterpene. PMID:24713615

Asna1/TRC40 Controls β-Cell Function and Endoplasmic Reticulum Homeostasis by Ensuring Retrograde Transport.

PubMed

Norlin, Stefan; Parekh, Vishal S; Naredi, Peter; Edlund, Helena

2016-01-01

Type 2 diabetes (T2D) is characterized by insulin resistance and β-cell failure. Insulin resistance per se, however, does not provoke overt diabetes as long as compensatory β-cell function is maintained. The increased demand for insulin stresses the β-cell endoplasmic reticulum (ER) and secretory pathway, and ER stress is associated with β-cell failure in T2D. The tail recognition complex (TRC) pathway, including Asna1/TRC40, is implicated in the maintenance of endomembrane trafficking and ER homeostasis. To gain insight into the role of Asna1/TRC40 in maintaining endomembrane homeostasis and β-cell function, we inactivated Asna1 in β-cells of mice. We show that Asna1(β-/-) mice develop hypoinsulinemia, impaired insulin secretion, and glucose intolerance that rapidly progresses to overt diabetes. Loss of Asna1 function leads to perturbed plasma membrane-to-trans Golgi network and Golgi-to-ER retrograde transport as well as to ER stress in β-cells. Of note, pharmacological inhibition of retrograde transport in isolated islets and insulinoma cells mimicked the phenotype of Asna1(β-/-) β-cells and resulted in reduced insulin content and ER stress. These data support a model where Asna1 ensures retrograde transport and, hence, ER and insulin homeostasis in β-cells. © 2016 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.

Disruption of Protein Processing in the Endoplasmic Reticulum of DYT1 Knock-in Mice Implicates Novel Pathways in Dystonia Pathogenesis.

PubMed

Beauvais, Genevieve; Bode, Nicole M; Watson, Jaime L; Wen, Hsiang; Glenn, Kevin A; Kawano, Hiroyuki; Harata, N Charles; Ehrlich, Michelle E; Gonzalez-Alegre, Pedro

2016-10-05

Dystonia type 1 (DYT1) is a dominantly inherited neurological disease caused by mutations in TOR1A, the gene encoding the endoplasmic reticulum (ER)-resident protein torsinA. Previous work mostly completed in cell-based systems suggests that mutant torsinA alters protein processing in the secretory pathway. We hypothesized that inducing ER stress in the mammalian brain in vivo would trigger or exacerbate mutant torsinA-induced dysfunction. To test this hypothesis, we crossed DYT1 knock-in with p58(IPK)-null mice. The ER co-chaperone p58(IPK) interacts with BiP and assists in protein maturation by helping to fold ER cargo. Its deletion increases the cellular sensitivity to ER stress. We found a lower generation of DYT1 knock-in/p58 knock-out mice than expected from this cross, suggesting a developmental interaction that influences viability. However, surviving animals did not exhibit abnormal motor function. Analysis of brain tissue uncovered dysregulation of eiF2α and Akt/mTOR translational control pathways in the DYT1 brain, a finding confirmed in a second rodent model and in human brain. Finally, an unbiased proteomic analysis identified relevant changes in the neuronal protein landscape suggesting abnormal ER protein metabolism and calcium dysregulation. Functional studies confirmed the interaction between the DYT1 genotype and neuronal calcium dynamics. Overall, these findings advance our knowledge on dystonia, linking translational control pathways and calcium physiology to dystonia pathogenesis and identifying potential new pharmacological targets. Dystonia type 1 (DYT1) is one of the different forms of inherited dystonia, a neurological disorder characterized by involuntary, disabling movements. DYT1 is caused by mutations in the gene that encodes the endoplasmic reticulum (ER)-resident protein torsinA. How mutant torsinA causes neuronal dysfunction remains unknown. Here, we show the behavioral and molecular consequences of stressing the ER in DYT1 mice by

Dystonin/Bpag1 is a necessary endoplasmic reticulum/nuclear envelope protein in sensory neurons

DOE Office of Scientific and Technical Information (OSTI.GOV)

Young, Kevin G.; University of Ottawa Center for Neuromuscular Disease, University of Ottawa, Ottawa, Ontario; Kothary, Rashmi

2008-09-10

Dystonin/Bpag1 proteins are cytoskeletal linkers whose loss of function in mice results in a hereditary sensory neuropathy with a progressive loss of limb coordination starting in the second week of life. These mice, named dystonia musculorum (dt), succumb to the disease and die of unknown causes prior to sexual maturity. Previous evidence indicated that cytoskeletal defects in the axon are a primary cause of dt neurodegeneration. However, more recent data suggests that other factors may be equally important contributors to the disease process. In the present study, we demonstrate perikaryal defects in dorsal root ganglion (DRG) neurons at stages precedingmore » the onset of loss of limb coordination in dt mice. Abnormalities include alterations in endoplasmic reticulum (ER) chaperone protein expression, indicative of an ER stress response. Dystonin in sensory neurons localized in association with the ER and nuclear envelope (NE). A fusion protein ofthe dystonin-a2 isoform, which harbors an N-terminal transmembrane domain, associated with and reorganized the ER in cell culture. This isoform also interacts with the NE protein nesprin-3{alpha}, but not nesprin-3{beta}. Defects in dt mice, as demonstrated here, may ultimately result in pathogenesis involving ER dysfunction and contribute significantly to the dt phenotype.« less

Specificity in endoplasmic reticulum-stress signaling in yeast entails a step-wise engagement of HAC1 mRNA to clusters of the stress sensor Ire1.

PubMed

van Anken, Eelco; Pincus, David; Coyle, Scott; Aragón, Tomás; Osman, Christof; Lari, Federica; Gómez Puerta, Silvia; Korennykh, Alexei V; Walter, Peter

2014-12-30

Insufficient protein-folding capacity in the endoplasmic reticulum (ER) induces the unfolded protein response (UPR). In the ER lumen, accumulation of unfolded proteins activates the transmembrane ER-stress sensor Ire1 and drives its oligomerization. In the cytosol, Ire1 recruits HAC1 mRNA, mediating its non-conventional splicing. The spliced mRNA is translated into Hac1, the key transcription activator of UPR target genes that mitigate ER-stress. In this study, we report that oligomeric assembly of the ER-lumenal domain is sufficient to drive Ire1 clustering. Clustering facilitates Ire1's cytosolic oligomeric assembly and HAC1 mRNA docking onto a positively charged motif in Ire1's cytosolic linker domain that tethers the kinase/RNase to the transmembrane domain. By the use of a synthetic bypass, we demonstrate that mRNA docking per se is a pre-requisite for initiating Ire1's RNase activity and, hence, splicing. We posit that such step-wise engagement between Ire1 and its mRNA substrate contributes to selectivity and efficiency in UPR signaling.

Lysosome and endoplasmic reticulum quality control pathways in Niemann-Pick type C disease

PubMed Central

Schultz, Mark L.; Krus, Kelsey L.; Lieberman, Andrew P.

2017-01-01

Lysosomal storage diseases result from inherited deficiencies of lysosomal hydrolytic activities or lipid transport. Collectively, these disorders are a common cause of morbidity in the pediatric population and are often associated with severe neurodegeneration. Among this group of diseases is Niemann-Pick type C, an autosomal recessive disorder of lipid trafficking that causes cognitive impairment, ataxia and death, most often in childhood. Here, we review the current knowledge of disease pathogenesis, with particular focus on insights gleaned from genetics and the study of model systems. Critical advances in understanding mechanisms that regulate intracellular cholesterol trafficking have emerged from this work and are highlighted. We review effects of disease-causing mutations on quality control pathways involving the lysosome and endoplasmic reticulum, and discuss how they function to clear the most common mutant protein found in Niemann-Pick type C patients, NPC1-I1061T. Finally, we summarize insights into the mechanisms that degrade misfolded transmembrane proteins in the endoplasmic reticulum and how manipulating these quality control pathways may lead to the identification of novel targets for disease-modifying therapies. PMID:27026653

26 CFR 1.412(c)(1)-3 - Applying the minimum funding requirements to restored plans.

Code of Federal Regulations, 2010 CFR

2010-04-01

... balance in the funding standard account is set equal to zero when the initial restoration amortization... that are sufficient to prevent the outstanding balance of the initial restoration amortization base..., the maximum permitted outstanding balance of the initial restoration amortization base at the end of...

Ghrelin Ameliorates Asthma by Inhibiting Endoplasmic Reticulum Stress.

PubMed

Fu, Tian; Wang, Lei; Zeng, Qingdi; Zhang, Yan; Sheng, Baowei; Han, Liping

2017-12-01

This study aimed to confirm the ameliorative effect of ghrelin on asthma and investigate its mechanism. The murine model of asthma was induced by ovalbumin (OVA) treatment and assessed by histological pathology and airway responsiveness to methacholine. The total and differential leukocytes were counted. Tumor necrosis factor α, interferon γ, interleukin-5 and interleukin-13 levels in bronchoalveolar lavage fluid were quantified by commercial kits. The protein levels in pulmonary tissues were measured by Western blot analysis. Ghrelin ameliorated the histological pathology and airway hyperresponsiveness in the OVA-induced asthmatic mouse model. Consistently, OVA-increased total and differential leukocytes and levels of tumor necrosis factor α, interferon γ, interleukin-5 and interleukin-13 in bronchoalveolar lavage fluid were significantly attenuated by ghrelin. Ghrelin prevented the increased protein levels of the endoplasmic reticulum stress markers glucose regulated protein 78 and CCAAT/enhancer binding protein homologous protein and reversed the reduced levels of p-Akt in asthmatic mice. Ghrelin might prevent endoplasmic reticulum stress activation by stimulating the Akt signaling pathway, which attenuated inflammation and ameliorated asthma in mice. Ghrelin might be a new target for asthma therapy. Copyright © 2017. Published by Elsevier Inc.

Lon in maintaining mitochondrial and endoplasmic reticulum homeostasis.

PubMed

Yang, Jieyeqi; Chen, Wenying; Zhang, Boyang; Tian, Fengli; Zhou, Zheng; Liao, Xin; Li, Chen; Zhang, Yi; Han, Yanyan; Wang, Yan; Li, Yuzhe; Wang, Guo-Qing; Shen, Xiao Li

2018-06-01

As a vital member of AAA+ (ATPase associated with diverse cellular activities) protein superfamily, Lon, a homo-hexameric ring-shaped protein complex with a serine-lysine catalytic dyad, is highly conserved throughout almost all prokaryotic and eukaryotic organisms. Lon protease (LONP) plays an important role in maintaining mitoproteostasis through selectively recognizing and degrading oxidatively modified mitoproteins within mitochondrial matrix, such as oxidized aconitase, phosphorylated mitochondrial transcription factor A, etc. Furthermore, the up-regulated LONP increased mitochondrial ROS generation to promote cell survival, cell proliferation, epithelial-mesenchymal transition, and cell migration, which was attributed to the up-regulation of NADH:ubiquinone oxidoreductase core subunit S8 via interaction with chaperone Lon under hypoxic or oxidative stress in tumorigenesis. In addition, Lon also participated in protein kinase RNA (PKR)-like endoplasmic reticulum kinase signaling pathway under endoplasmic reticulum (ER) stress. In short, Lon, as a pivotal stress-responsive protein that involved in the crosstalks among mitochondria, ER and nucleus, participated in multifarious important cellular processes crucial for cell survival, such as the mitochondrial protein quality control system, the mitochondrial unfolded protein response, the mtDNA maintenance, and the ER unfolded protein response.

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

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sun, Jing; Department of Dental Implantology, School of Stomatology, Tongji University, Shanghai 200072; Ogata, Shigenori

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 withmore » 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}.« less

Inhibition of Reticulon-1A-Mediated Endoplasmic Reticulum Stress in Early AKI Attenuates Renal Fibrosis Development.

PubMed

Fan, Ying; Xiao, Wenzhen; Lee, Kyung; Salem, Fadi; Wen, Jiejun; He, Li; Zhang, Jing; Fei, Yang; Cheng, Dongsheng; Bao, Hongda; Liu, Yumei; Lin, Fujun; Jiang, Gengru; Guo, Zhiyong; Wang, Niansong; He, John Cijiang

2017-07-01

Several animal studies have shown an important role for endoplasmic reticulum (ER) stress in AKI, whereas human studies are lacking. We recently reported that Reticulon-1A (RTN1A) is a key mediator of ER stress and kidney cell injury. Here, we investigated whether modulation of RTN1A expression during AKI contributes to the progression to CKD. In a retrospective study of 51 patients with AKI, increased expression of RTN1A and other ER stress markers were associated with the severity of kidney injury and with progression to CKD. In an inducible tubular cell-specific RTN1A-knockdown mouse model subjected to folic acid nephropathy (FAN) or aristolochic acid nephropathy, reduction of RTN1A expression during the initial stage of AKI attenuated ER stress and kidney cell injury in early stages and renal fibrosis development in later stages. Treatment of wild-type mice with tauroursodeoxycholic acid, an inhibitor of ER stress, after the induction of kidney injury with FA facilitated renoprotection similar to that observed in RTN1A-knockdown mice. Conversely, in transgenic mice with inducible tubular cell-specific overexpression of RTN1A subjected to FAN, induction of RTN1A overexpression aggravated ER stress and renal injury at the early stage and renal fibrosis at the late stage of FAN. Together, our human and mouse data suggest that the RTN1A-mediated ER stress response may be an important determinant in the severity of AKI and maladaptive repair that may promote progression to CKD. Copyright © 2017 by the American Society of Nephrology.

Endoplasmic Reticulum Stress and Unfolded Protein Response in Cartilage Pathophysiology; Contributing Factors to Apoptosis and Osteoarthritis.

PubMed

Hughes, Alexandria; Oxford, Alexandra E; Tawara, Ken; Jorcyk, Cheryl L; Oxford, Julia Thom

2017-03-20

Chondrocytes of the growth plate undergo apoptosis during the process of endochondral ossification, as well as during the progression of osteoarthritis. Although the regulation of this process is not completely understood, alterations in the precisely orchestrated programmed cell death during development can have catastrophic results, as exemplified by several chondrodystrophies which are frequently accompanied by early onset osteoarthritis. Understanding the mechanisms that underlie chondrocyte apoptosis during endochondral ossification in the growth plate has the potential to impact the development of therapeutic applications for chondrodystrophies and associated early onset osteoarthritis. In recent years, several chondrodysplasias and collagenopathies have been recognized as protein-folding diseases that lead to endoplasmic reticulum stress, endoplasmic reticulum associated degradation, and the unfolded protein response. Under conditions of prolonged endoplasmic reticulum stress in which the protein folding load outweighs the folding capacity of the endoplasmic reticulum, cellular dysfunction and death often occur. However, unfolded protein response (UPR) signaling is also required for the normal maturation of chondrocytes and osteoblasts. Understanding how UPR signaling may contribute to cartilage pathophysiology is an essential step toward therapeutic modulation of skeletal disorders that lead to osteoarthritis.

Fracture Resistance of Teeth Restored with Direct and Indirect Composite Restorations

PubMed Central

Torabzadeh, Hassan; Ghasemi, Amir; Dabestani, Atoosa; Razmavar, Sara

2013-01-01

Objective: Tooth fracture is a common dental problem. By extension of cavity dimensions, the remaining tooth structure weakens and occlusal forces may cause tooth fracture. The aim of this study was to evaluate and compare the fracture resistance of teeth restored with direct and indirect composite restorations. Materials and Methods: Sixty-five sound maxillary premolar teeth were chosen and randomly divided into five groups each comprising thirteen. Fifty-two teeth received mesio-occluso-distal (MOD) cavities with 4.5mm bucco-lingual width, 4mm pulpal depth and 3mm gingival depth and were divided into the following four groups. G-1: restored with direct composite (Z-250, 3M/ESPE) with cusp coverage, G-2: restored with direct composite (Z-250) without cusp coverage, G-3: restored with direct composite (Gradia, GC-international) with cusp coverage, G-4: restored with indirect composite (Gradia, GC-International) with cusp coverage. Intact teeth were used in G-5 as control. The teeth were subjected to a compressive axial loading using a 4 mm diameter rod in a universal testing machine with 1 mm/min speed. Data were analyzed using one-way ANOVA and Tukey tests. Results: The mean fracture strength recorded was: G-1: 1148.46N±262, G-2: 791.54N±235, G-3: 880.00N±123, G-4: 800.00N±187, G-5: 1051.54N±345. ANOVA revealed significant differences between groups (p<0.05). Tukey test showed significant difference between group 1 and the other groups. There was no significant difference among other groups. Conclusion: Direct composite (Z-250) with cusp coverage is a desirable treatment for weakened teeth. Treatment with Z-250 without cusp coverage, direct and indirect Gradia with cusp coverage restored the strength of the teeth to the level of intact teeth. PMID:24910649

Regulation of calcium release from the endoplasmic reticulum by the serine hydrolase ABHD2.

PubMed

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.

Chemical Chaperones Reduce ER Stress and Restore Glucose Homeostasis in a Mouse Model of Type 2 Diabetes

NASA Astrophysics Data System (ADS)

Özcan, Umut; Yilmaz, Erkan; Özcan, Lale; Furuhashi, Masato; Vaillancourt, Eric; Smith, Ross O.; Görgün, Cem Z.; Hotamisligil, Gökhan S.

2006-08-01

Endoplasmic reticulum (ER) stress is a key link between obesity, insulin resistance, and type 2 diabetes. Here, we provide evidence that this mechanistic link can be exploited for therapeutic purposes with orally active chemical chaperones. 4-Phenyl butyric acid and taurine-conjugated ursodeoxycholic acid alleviated ER stress in cells and whole animals. Treatment of obese and diabetic mice with these compounds resulted in normalization of hyperglycemia, restoration of systemic insulin sensitivity, resolution of fatty liver disease, and enhancement of insulin action in liver, muscle, and adipose tissues. Our results demonstrate that chemical chaperones enhance the adaptive capacity of the ER and act as potent antidiabetic modalities with potential application in the treatment of type 2 diabetes.

Silver nanoparticles induce SH-SY5Y cell apoptosis via endoplasmic reticulum- and mitochondrial pathways that lengthen endoplasmic reticulum-mitochondria contact sites and alter inositol-3-phosphate receptor function.

PubMed

Li, Lin; Cui, Jiahui; Liu, Zi; Zhou, Xuejiao; Li, Zengqiang; Yu, Yang; Jia, Yuanyuan; Zuo, Daiying; Wu, Yingliang

2018-03-15

Silver nanoparticles (AgNPs) have many medical and commercial applications, but their effects on human health are poorly understood. The aim of this study was to assess the effect of AgNPs on the human neuroblastoma cell line SH-SY5Y and to explore their potential mechanisms of action. We found that AgNPs decreased SH-SY5Y cell viability in a dose- and time-dependent manner. Exposure to AgNPs activated endoplasmic reticulum (ER) stress, as reflected by upregulated expression of glucose-regulated protein 78 (GRP78), phosphorylated PKR-like endoplasmic reticulum kinase (p-PERK), phosphorylated eukaryotic translation initiation factor 2α (p-eIF2α), C/EBP homology protein (CHOP), spliced X-box binding protein-1 (XBP1), and phosphorylated inositol-requiring enzyme (p-IRE), all of which are involved in the cellular unfolded protein response. Prolonged exposure of cells to AgNPs damaged calcium (Ca 2+ ) homeostasis, increased the length of contact sites between the ER and mitochondria, altered IP 3 R function by the increased levels of phosphatase and tensin homolog deleted on chromosome ten (PTEN) in the ER and enhanced mitochondrial Ca 2+ uptake. Finally, Ca 2+ overload and disrupted homeostasis in the mitochondria triggered apoptotic cell death. Our results suggest that caution should be exercised in the use of AgNPs in humans. Copyright © 2018 Elsevier B.V. All rights reserved.

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

PubMed

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.

Biosynthesis and processing of ribophorins in the endoplasmic reticulum.

PubMed

Rosenfeld, M G; Marcantonio, E E; Hakimi, J; Ort, V M; Atkinson, P H; Sabatini, D; Kreibich, G

1984-09-01

Ribophorins are two transmembrane glycoproteins characteristic of the rough endoplasmic reticulum, which are thought to be involved in the binding of ribosomes. Their biosynthesis was studied in vivo using lines of cultured rat hepatocytes (clone 9) and pituitary cells (GH 3.1) and in cell-free synthesis experiments. In vitro translation of mRNA extracted from free and bound polysomes of clone 9 cells demonstrated that ribophorins are made exclusively on bound polysomes. The primary translation products of ribophorin messengers obtained from cultured hepatocytes or from regenerating livers co-migrated with the respective mature proteins, but had slightly higher apparent molecular weights (2,000) than the unglycosylated forms immunoprecipitated from cells treated with tunicamycin. This indicates that ribophorins, in contrast to all other endoplasmic reticulum membrane proteins previously studied, contain transient amino-terminal insertion signals which are removed co-translationally. Kinetic and pulse-chase experiments with [35S]methionine and [3H]mannose demonstrated that ribophorins are not subjected to electrophoretically detectable posttranslational modifications, such as proteolytic cleavage or trimming and terminal glycosylation of oligosaccharide side chain(s). Direct analysis of the oligosaccharides of ribophorin l showed that they do not contain the terminal sugars characteristic of complex oligosaccharides and that they range in composition from Man8GlcNAc to Man5GlcNAc. These findings, as well as the observation that the mature proteins are sensitive to endoglycosidase H and insensitive to endoglycosidase D, are consistent with the notion that the biosynthetic pathway of the ribophorins does not require a stage of passage through the Golgi apparatus.

Chemical chaperon 4-phenylbutyrate protects against the endoplasmic reticulum stress-mediated renal fibrosis in vivo and in vitro.

PubMed

Liu, Shing-Hwa; Yang, Ching-Chin; Chan, Ding-Cheng; Wu, Cheng-Tien; Chen, Li-Ping; Huang, Jenq-Wen; Hung, Kuan-Yu; Chiang, Chih-Kang

2016-04-19

Renal tubulointerstitial fibrosis is the common and final pathologic change of kidney in end-stage renal disease. Interesting, endoplasmic reticulum (ER) stress is known to contribute to the pathophysiological mechanisms during the development of renal fibrosis. Here, we investigated the effects of chemical chaperon sodium 4-phenylbutyrate (4-PBA) on renal fibrosis in vivo and in vitro. In a rat unilateral ureteral obstruction (UUO) model, 4-PBA mimicked endogenous ER chaperon in the kidneys and significantly reduced glucose regulated protein 78 (GRP78), CCAAT/enhancer binding protein (C/EBP) homologous protein (CHOP), activating transcription factor 4 (ATF4), and phosphorylated JNK protein expressions as well as restored spliced X-box-binding protein 1 (XBP1) expressions in the kidneys of UUO rats. 4-PBA also attenuated the increases of α-smooth muscle actin (α-SMA), connective tissue growth factor (CTGF) protein expressions, tubulointerstitial fibrosis, and apoptosis in the kidneys of UUO rats. Moreover, transforming growth factor (TGF)-β markedly increased ER stress-associated molecules, profibrotic factors, and apoptotic markers in the renal tubular cells (NRK-52E), all of which could be significantly counteracted by 4-PBA treatment. 4-PBA also diminished TGF-β-increased CTGF promoter activity and CTGF mRNA expression in NRK-52E cells. Taken together, our results indicated that 4-PBA acts as an ER chaperone to ameliorate ER stress-induced renal tubular cell apoptosis and renal fibrosis.

[Comparison of color reappearance between metal-ceram restoration and foundry-ceram restoration using crystaleye spectrophotometer].

PubMed

Shi, Tao; Zhang, Ning; Kong, Fan-wen; Zhan, De-song

2010-10-01

To study the color reappearance effect of metal-ceram restoration and foundry-ceram restoration using Crystaleye spectrophotometer. 58 metal-ceram restorations and 58 foundry-ceram restorations according to the result of the Crystaleye spectrophotometer were made respectively. The deltaE between restorations and natural teeth as referenced were analyzed. And satisfaction of dentists and patients were evaluated. The deltaE between metal-ceram restorations and natural teeth was 7.13 +/- 0.74. The deltaE between foundry-ceram restorations and teeth was 1.47 +/- 0.84. There were statistical differences between the deltaE (P < 0.01). The satisnatural faction of dentists and patients to the foundry-ceram restorations were higher than to the metal-ceram restorations (P < 0.05). Crystaleye spectrophotometer can provide accurate reference for foundry-ceram restoration, but for metal-ceram restoration it is not accurate.

Chlorhexidine-induced apoptosis or necrosis in L929 fibroblasts: A role for endoplasmic reticulum stress

DOE Office of Scientific and Technical Information (OSTI.GOV)

Faria, Gisele; Cardoso, Cristina R.B.; Department of Biological Sciences, Federal University of Triangulo Mineiro, Uberaba, Minas Gerais

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 ofmore » 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.« less

Early Restoration | NOAA Gulf Spill Restoration

Science.gov Websites

Early Restoration Plan. On April 20, 2011 we reached an agreement with BP to start restoration planning draft plan for the third phase of early restoration in December 2013. We are considering your comments : All Phase III information and documents Phase II Useful Links: Phase II Early Restoration Plan &

Restoring the impaired cardiac calcium homeostasis and cardiac function in iron overload rats by the combined deferiprone and N-acetyl cysteine

PubMed Central

Wongjaikam, Suwakon; Kumfu, Sirinart; Khamseekaew, Juthamas; Chattipakorn, Siriporn C.; Chattipakorn, Nipon

2017-01-01

Intracellular calcium [Ca2+]i dysregulation plays an important role in the pathophysiology of iron overload cardiomyopathy. Although either iron chelators or antioxidants provide cardioprotection, a comparison of the efficacy of deferoxamine (DFO), deferiprone (DFP), deferasirox (DFX), N-acetyl cysteine (NAC) or a combination of DFP plus NAC on cardiac [Ca2+]i homeostasis in chronic iron overload has never been investigated. Male Wistar rats were fed with either a normal diet or a high iron (HFe) diet for 4 months. At 2 months, HFe rats were divided into 6 groups and treated with either a vehicle, DFO (25 mg/kg/day), DFP (75 mg/kg/day), DFX (20 mg/kg/day), NAC (100 mg/kg/day), or combined DFP plus NAC. At 4 months, the number of cardiac T-type calcium channels was increased, whereas cardiac sarcoplasmic-endoplasmic reticulum Ca2+ ATPase (SERCA) was decreased, leading to cardiac iron overload and impaired cardiac [Ca2+]i homeostasis. All pharmacological interventions restored SERCA levels. Although DFO, DFP, DFX or NAC alone shared similar efficacy in improving cardiac [Ca2+]i homeostasis, only DFP + NAC restored cardiac [Ca2+]i homeostasis, leading to restoring left ventricular function in the HFe-fed rats. Thus, the combined DFP + NAC was more effective than any monotherapy in restoring cardiac [Ca2+]i homeostasis, leading to restored myocardial contractility in iron-overloaded rats. PMID:28287621

Metformin Restores Parkin-Mediated Mitophagy, Suppressed by Cytosolic p53

PubMed Central

Song, Young Mi; Lee, Woo Kyung; Lee, Yong-ho; Kang, Eun Seok; Cha, Bong-Soo; Lee, Byung-Wan

2016-01-01

Metformin is known to alleviate hepatosteatosis by inducing 5’ adenosine monophosphate (AMP)-kinase-independent, sirtuin 1 (SIRT1)-mediated autophagy. Dysfunctional mitophagy in response to glucolipotoxicities might play an important role in hepatosteatosis. Here, we investigated the mechanism by which metformin induces mitophagy through restoration of the suppressed Parkin-mediated mitophagy. To this end, our ob/ob mice were divided into three groups: (1) ad libitum feeding of a standard chow diet; (2) intraperitoneal injections of metformin 300 mg/kg; and (3) 3 g/day caloric restriction (CR). HepG2 cells were treated with palmitate (PA) plus high glucose in the absence or presence of metformin. We detected enhanced mitophagy in ob/ob mice treated with metformin or CR, whereas mitochondrial spheroids were observed in mice fed ad libitum. Metabolically stressed ob/ob mice and PA-treated HepG2 cells showed an increase in expression of endoplasmic reticulum (ER) stress markers and cytosolic p53. Cytosolic p53 inhibited mitophagy by disturbing the mitochondrial translocation of Parkin, as demonstrated by immunoprecipitation. However, metformin decreased ER stress and p53 expression, resulting in induction of Parkin-mediated mitophagy. Furthermore, pifithrin-α, a specific inhibitor of p53, increased mitochondrial incorporation into autophagosomes. Taken together, these results indicate that metformin treatment facilitates Parkin-mediated mitophagy rather than mitochondrial spheroid formation by decreasing the inhibitory interaction with cytosolic p53 and increasing degradation of mitofusins. PMID:26784190

32 CFR 644.453 - Major restoration cases-determining extent of restoration required.

Code of Federal Regulations, 2010 CFR

2010-07-01

... Improvements § 644.453 Major restoration cases—determining extent of restoration required. (a) Engineer... the item numbers on the Recapitulation sheet, part I of the form: (1) “1” to “6” Self-explanatory. (2) “7. Original Cost (Actual or Estimated) of Government-owned improvements, fixtures and alterations...

32 CFR 644.453 - Major restoration cases-determining extent of restoration required.

Code of Federal Regulations, 2014 CFR

2014-07-01

... Improvements § 644.453 Major restoration cases—determining extent of restoration required. (a) Engineer... the item numbers on the Recapitulation sheet, part I of the form: (1) “1” to “6” Self-explanatory. (2) “7. Original Cost (Actual or Estimated) of Government-owned improvements, fixtures and alterations...

32 CFR 644.453 - Major restoration cases-determining extent of restoration required.

Code of Federal Regulations, 2012 CFR

2012-07-01

... Improvements § 644.453 Major restoration cases—determining extent of restoration required. (a) Engineer... the item numbers on the Recapitulation sheet, part I of the form: (1) “1” to “6” Self-explanatory. (2) “7. Original Cost (Actual or Estimated) of Government-owned improvements, fixtures and alterations...

32 CFR 644.453 - Major restoration cases-determining extent of restoration required.

Code of Federal Regulations, 2013 CFR

2013-07-01

... Improvements § 644.453 Major restoration cases—determining extent of restoration required. (a) Engineer... the item numbers on the Recapitulation sheet, part I of the form: (1) “1” to “6” Self-explanatory. (2) “7. Original Cost (Actual or Estimated) of Government-owned improvements, fixtures and alterations...

Exploring the genetics of fertility restoration controlled by Rf1 in common wheat (Triticum aestivum L.) using high-density linkage maps.

PubMed

Geyer, Manuel; Albrecht, Theresa; Hartl, Lorenz; Mohler, Volker

2018-04-01

Hybrid wheat breeding has the potential to significantly increase wheat productivity compared to line breeding. The induction of male sterility by the cytoplasm of Triticum timopheevii Zhuk. is a widely discussed approach to ensure cross-pollination between parental inbred lines in hybrid wheat seed production. As fertility restoration in hybrids with this cytoplasm is often incomplete, understanding the underlying genetics is a prerequisite to apply this technology. A promising component for fertility restoration is the restorer locus Rf1, which was first detected on chromosome 1A of the restorer accession R3. In the present study, we performed quantitative trait locus (QTL) analyses to locate Rf1 and estimate its effect in populations involving the restorer lines R3, R113 and L19. Molecular markers linked to Rf1 in these populations were used to analyse the genomic target region in T. timopheevii accessions and common wheat breeding lines. The QTL analyses revealed that Rf1 interacted with a modifier locus on chromosome 1BS and the restorer locus Rf4 on chromosome 6B. The modifier locus significantly influenced both the penetrance and expressivity of Rf1. Whereas Rf1 exhibited expressivity higher than that of Rf4, the effects of these loci were not additive. Evaluating the marker haplotype for the Rf1 region, we propose that the restoring Rf1 allele may be derived exclusively from T. timopheevii. The present study demonstrates that interactions between restorer and modifier loci play a critical role in fertility restoration of common wheat with the cytoplasm of T. timopheevii.

MoYvh1 subverts rice defense through functions of ribosomal protein MoMrt4 in Magnaporthe oryzae.

PubMed

Liu, Xinyu; Yang, Jie; Qian, Bin; Cai, Yongchao; Zou, Xi; Zhang, Haifeng; Zheng, Xiaobo; Wang, Ping; Zhang, Zhengguang

2018-04-01

The accumulation of the reactive oxygen species (ROS) in rice is important in its interaction with the rice blast fungus Magnaporthe oryzae during which the pathogen scavenges ROS through the production of extracellular enzymes that promote blast. We previously characterized the MoYvh1 protein phosphatase from M. oryzae that plays a role in scavenging of ROS. To understand the underlying mechanism, we found that MoYvh1 is translocated into the nucleus following oxidative stress and that this translocation is dependent on MoSsb1 and MoSsz1 that are homologous to heat-shock protein 70 (Hsp70) proteins. In addition, we established a link between MoYvh1 and MoMrt4, a ribosome maturation factor homolog whose function also involves shuttling between the cytoplasm and the nucleus. Moreover, we found that MoYvh1 regulates the production of extracellular proteins that modulate rice-immunity. Taking together, our evidence suggests that functions of MoYvh1 in regulating ROS scavenging require its nucleocytoplasmic shuttling and the partner proteins MoSsb1 and MoSsz1, as well as MoMrt4. Our findings provide novel insights into the mechanism by which M. oryzae responds to and subverts host immunity through the regulation of ribosome biogenesis and protein biosynthesis.

TANGO1 and Mia2/cTAGE5 (TALI) cooperate to export bulky pre-chylomicrons/VLDLs from the endoplasmic reticulum.

PubMed

Santos, António J M; Nogueira, Cristina; Ortega-Bellido, Maria; Malhotra, Vivek

2016-05-09

Procollagens, pre-chylomicrons, and pre-very low-density lipoproteins (pre-VLDLs) are too big to fit into conventional COPII-coated vesicles, so how are these bulky cargoes exported from the endoplasmic reticulum (ER)? We have shown that TANGO1 located at the ER exit site is necessary for procollagen export. We report a role for TANGO1 and TANGO1-like (TALI), a chimeric protein resulting from fusion of MIA2 and cTAGE5 gene products, in the export of pre-chylomicrons and pre-VLDLs from the ER. TANGO1 binds TALI, and both interact with apolipoprotein B (ApoB) and are necessary for the recruitment of ApoB-containing lipid particles to ER exit sites for their subsequent export. Although export of ApoB requires the function of both TANGO1 and TALI, the export of procollagen XII by the same cells requires only TANGO1. These findings reveal a general role for TANGO1 in the export of bulky cargoes from the ER and identify a specific requirement for TALI in assisting TANGO1 to export bulky lipid particles. © 2016 Santos et al.

Detergent-Resistant Microdomains Determine the Localization of σ-1 Receptors to the Endoplasmic Reticulum-Mitochondria JunctionS⃞

PubMed Central

Fujimoto, Michiko

2010-01-01

σ-1 receptors (Sig-1Rs) that bind diverse synthetic and endogenous compounds have been implicated in the pathophysiology of several human diseases such as drug addiction, depression, neurodegenerative disorders, pain-related disorders, and cancer. Sig-1Rs were identified recently as novel ligand-operated molecular chaperones. Although Sig-1Rs are predominantly expressed at endoplasmic reticulum (ER) subdomains apposing mitochondria [i.e., the mitochondria-associated ER membrane (MAM)], they dynamically change the cellular distribution, thus regulating both MAM-specific and plasma membrane proteins. However, what determines the location of Sig-1R at the MAM and how the receptor translocation is initiated is unknown. Here we report that the detergent-resistant membranes (DRMs) play an important role in anchoring Sig-1Rs to the MAM. The MAM, which is highly capable of accumulating ceramides, is enriched with both cholesterol and simple sphingolipids, thus forming Triton X-114-resistant DRMs. Sig-1Rs associate with MAM-derived DRMs but not with those from microsomes. A lipid overlay assay found that solubilized Sig-1Rs preferentially associate with simple sphingolipids such as ceramides. Disrupting DRMs by lowering cholesterol or inhibiting de novo synthesis of ceramides at the ER largely decreases Sig-1R at DRMs and causes translocation of Sig-1R from the MAM to ER cisternae. These findings suggest that the MAM, bearing cholesterol and ceramide-enriched microdomains at the ER, may use the microdomains to anchor Sig-1Rs to the location; thus, it serves to stage Sig-1R at ER-mitochondria junctions. PMID:20053954

Identification of discrete sites in Yip1A necessary for regulation of endoplasmic reticulum structure.

PubMed

Dykstra, Kaitlyn M; Ulengin, Idil; Delrose, Nicholas; Lee, Tina H

2013-01-01

The endoplasmic reticulum (ER) of specialized cells can undergo dramatic changes in structural organization, including formation of concentric whorls. We previously reported that depletion of Yip1A, an integral membrane protein conserved between yeast and mammals, caused ER whorl formation reminiscent of that seen in specialized cells. Yip1A and its yeast homologue Yip1p cycle between the ER and early Golgi, have been implicated in a number of distinct trafficking steps, and interact with a conserved set of binding partners including Yif1p/Yif1A and the Ypt1/Ypt31 Rab GTPases. Here, we carried out a mutational analysis of Yip1A to obtain insight into how it regulates ER whorl formation. Most of the Yip1A cytoplasmic domain was dispensable, whereas the transmembrane (TM) domain, especially residues within predicted TM helices 3 and 4, were sensitive to mutagenesis. Comprehensive analysis revealed two discrete functionally required determinants. One was E95 and flanking residues L92 and L96 within the cytoplasmic domain; the other was K146 and nearby residue V152 within the TM domain. Notably, the identified determinants correspond closely to two sites previously found to be essential for yeast viability (E76 and K130 in Yip1p corresponding to E95 and K146 in Yip1A, respectively). In contrast, a third site (E89) also essential for yeast viability (E70 in Yip1p) was dispensable for regulation of whorl formation. Earlier work showed that E76 (E95) was dispensable for binding Yif1p or Ypt1p/Ypt31p, whereas E70 (E89) was required. Collectively, these findings suggest that the ability of Yip1A to bind its established binding partners may be uncoupled from its ability to control ER whorl formation. In support, Yif1A knockdown did not cause ER whorl formation. Thus Yip1A may use the sites identified herein to interact with a novel binding partner to regulate ER membrane organization.

Hierarchically Self-Assembled Star-Shaped ZnO Microparticles for Electrochemical Sensing of Amines.

PubMed

Du, Jianping; Huang, Xiaoxi; Zhao, Ruihua; Li, Jinping; Asefa, Tewodros

2016-06-06

Novel, hierarchically nanostructured, star-shaped ZnO (SSZ) microparticles are synthesized by a hydrothermal synthetic route. The SSZ microparticles serve as effective platforms for electrochemical detection of amines in solution. The morphology and structure of the materials are characterized by X-ray diffraction, scanning electron microscopy, transmission electron microscopy, Raman spectroscopy, and UV/Vis spectroscopy. The as-synthesized SSZ microparticles comprise self-assembled hexagonal prisms that possess nanometer and micrometer pores in their structure and on their surfaces-structural features that are conducive to sensing applications. An electrode fabricated by using the hierarchically nanostructured SSZ materials serve as a sensitive electrochemical sensor for detection of low concentrations of ethylenediamine, with a sensitivity of 2.98×10(-2)  mA cm(-2)  mm(-1) , a detection limit of 2.36×10(-2)  mm, and a short response time of 8 s. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Calneuron 1 Increased Ca2+ in the Endoplasmic Reticulum and Aldosterone Production in Aldosterone-Producing Adenoma.

PubMed

Kobuke, Kazuhiro; Oki, Kenji; Gomez-Sanchez, Celso E; Gomez-Sanchez, Elise P; Ohno, Haruya; Itcho, Kiyotaka; Yoshii, Yoko; Yoneda, Masayasu; Hattori, Noboru

2018-01-01

Aldosterone production is initiated by angiotensin II stimulation and activation of intracellular Ca 2+ signaling. In aldosterone-producing adenoma (APA) cells, the activation of intracellular Ca 2+ signaling is independent of the renin-angiotensin-aldosterone systems. The purpose of our study was to clarify molecular mechanisms of aldosterone production related to Ca 2+ signaling. Transcriptome analysis revealed that the CALN1 gene encoding calneuron 1 had the strongest correlation with CYP11B2 (aldosterone synthase) among genes encoding Ca 2+ -binding proteins in APA. CALN1 modulation and synthetic or fluorescent compounds were used for functional studies in human adrenocortical carcinoma (HAC15) cells. CALN1 expression was 4.4-fold higher in APAs than nonfunctioning adrenocortical adenomas. CALN1 expression colocalized with CYP11B2 expression as investigated using immunohistochemistry in APA and zona glomerulosa of male rats fed by a low-salt diet. CALN1 expression was detected in the endoplasmic reticulum (ER) by using GFP-fused CALN1, CellLight ER-RFP, and the corresponding antibodies. CALN1 -overexpressing HAC15 cells showed increased Ca 2+ in the ER and cytosol fluorescence-based studies. Aldosterone production was potentiated in HAC15 cells by CALN1 expression, and dose-responsive inhibition with TMB-8 showed that CALN1-mediated Ca 2+ storage in ER involved sarcoendoplasmic reticulum calcium transport ATPase. The silencing of CALN1 decreased Ca 2+ in ER, and abrogated angiotensin II- or KCNJ5 T158A-mediated aldosterone production in HAC15 cells. Increased CALN1 expression in APA was associated with elevated Ca 2+ storage in ER and aldosterone overproduction. Suppression of CALN1 expression prevented angiotensin II- or KCNJ5 T158A-mediated aldosterone production in HAC15 cells, suggesting that CALN1 is a potential therapeutic target for excess aldosterone production. © 2017 American Heart Association, Inc.

Gene regulatory network of unfolded protein response genes in endoplasmic reticulum stress.

PubMed

Takayanagi, Sayuri; Fukuda, Riga; Takeuchi, Yuuki; Tsukada, Sakiko; Yoshida, Kenichi

2013-01-01

In the endoplasmic reticulum (ER), secretory and membrane proteins are properly folded and modified, and the failure of these processes leads to ER stress. At the same time, unfolded protein response (UPR) genes are activated to maintain homeostasis. Despite the thorough characterization of the individual gene regulation of UPR genes to date, further investigation of the mutual regulation among UPR genes is required to understand the complex mechanism underlying the ER stress response. In this study, we aimed to reveal a gene regulatory network formed by UPR genes, including immunoglobulin heavy chain-binding protein (BiP), X-box binding protein 1 (XBP1), C/EBP [CCAAT/enhancer-binding protein]-homologous protein (CHOP), PKR-like endoplasmic reticulum kinase (PERK), inositol-requiring 1 (IRE1), activating transcription factor 6 (ATF6), and ATF4. For this purpose, we focused on promoter-luciferase reporters for BiP, XBP1, and CHOP genes, which bear an ER stress response element (ERSE), and p5 × ATF6-GL3, which bears an unfolded protein response element (UPRE). We demonstrated that the luciferase activities of the BiP and CHOP promoters were upregulated by all the UPR genes, whereas those of the XBP1 promoter and p5 × ATF6-GL3 were upregulated by all the UPR genes except for BiP, CHOP, and ATF4 in HeLa cells. Therefore, an ERSE- and UPRE-centered gene regulatory network of UPR genes could be responsible for the robustness of the ER stress response. Finally, we revealed that BiP protein was degraded when cells were treated with DNA-damaging reagents, such as etoposide and doxorubicin; this finding suggests that the expression level of BiP is tightly regulated at the post-translational level, rather than at the transcriptional level, in the presence of DNA damage.

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

PubMed

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.

Selective Modulation of Endoplasmic Reticulum Stress Markers in Prostate Cancer Cells by a Standardized Mangosteen Fruit Extract

PubMed Central

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. PMID:24367485

Ermelin, an endoplasmic reticulum transmembrane protein, contains the novel HELP domain conserved in eukaryotes.

PubMed

Suzuki, Akiko; Endo, Takeshi

2002-02-06

We have cloned a cDNA encoding a novel protein referred to as ermelin from mouse C2 skeletal muscle cells. This protein contained six hydrophobic amino acid stretches corresponding to transmembrane domains, two histidine-rich sequences, and a sequence homologous to the fusion peptides of certain fusion proteins. Ermelin also contained a novel modular sequence, designated as HELP domain, which was highly conserved among eukaryotes, from yeast to higher plants and animals. All these HELP domain-containing proteins, including mouse KE4, Drosophila Catsup, and Arabidopsis IAR1, possessed multipass transmembrane domains and histidine-rich sequences. Ermelin was predominantly expressed in brain and testis, and induced during neuronal differentiation of N1E-115 neuroblastoma cells but downregulated during myogenic differentiation of C2 cells. The mRNA was accumulated in hippocampus and cerebellum of brain and central areas of seminiferous tubules in testis. Epitope-tagging experiments located ermelin and KE4 to a network structure throughout the cytoplasm. Staining with the fluorescent dye DiOC(6)(3) identified this structure as the endoplasmic reticulum. These results suggest that at least some, if not all, of the HELP domain-containing proteins are multipass endoplasmic reticulum membrane proteins with functions conserved among eukaryotes.

Endoplasmic reticulum-Golgi intermediate compartment protein 3 knockdown suppresses lung cancer through endoplasmic reticulum stress-induced autophagy.

PubMed

Hong, Seong-Ho; Chang, Seung-Hee; Cho, Kyung-Cho; Kim, Sanghwa; Park, Sungjin; Lee, Ah Young; Jiang, Hu-Lin; Kim, Hyeon-Jeong; Lee, Somin; Yu, Kyeong-Nam; Seo, Hwi Won; Chae, Chanhee; Kim, Kwang Pyo; Park, Jongsun; Cho, Myung-Haing

2016-10-04

Trafficking from the endoplasmic reticulum (ER) to the Golgi apparatus is elevated in cancer cells. Therefore, proteins of the ER-Golgi intermediate compartment (ERGIC) attract significant attention as targets for cancer treatment. Enhanced cancer cell growth and epithelial-mesenchymal transition by ERGICs correlates with poor-prognosis of lung cancer. This prompted us to assess whether knockdown of ERGIC3 may decrease lung cancer growth. To test the hypothesis, the effects of ERGIC3 short hairpin RNA (shERGIC3) on ER stress-induced cell death and lung tumorigenesis were investigated both in vitro and in vivo. Knockdown of ERGIC3 led to ER stress-induced autophagic cell death and suppression of proliferation in the A549 human lung cancer cell-line. Moreover, non-invasive aerosol-delivery of shERGIC3 using the biocompatible carrier glycerol propoxylate triacrylate and spermine (GPT-SPE) inhibited lung tumorigenesis in the K-rasLA1 murine model of lung cancer. Our data suggest that suppression of ERGIC3 could provide a framework for the development of effective lung cancer therapies.

Molecular Characterization of the Elaeis guineensis Medium-Chain Fatty Acid Diacylglycerol Acyltransferase DGAT1-1 by Heterologous Expression in Yarrowia lipolytica.

PubMed

Aymé, Laure; Jolivet, Pascale; Nicaud, Jean-Marc; Chardot, Thierry

2015-01-01

Diacylglycerol acyltransferases (DGAT) are involved in the acylation of sn-1,2-diacylglycerol. Palm kernel oil, extracted from Elaeis guineensis (oil palm) seeds, has a high content of medium-chain fatty acids mainly lauric acid (C12:0). A putative E. guineensis diacylglycerol acyltransferase gene (EgDGAT1-1) is expressed at the onset of lauric acid accumulation in the seed endosperm suggesting that it is a determinant of medium-chain triacylglycerol storage. To test this hypothesis, we thoroughly characterized EgDGAT1-1 activity through functional complementation of a Yarrowia lipolytica mutant strain devoid of neutral lipids. EgDGAT1-1 expression is sufficient to restore triacylglycerol accumulation in neosynthesized lipid droplets. A comparative functional study with Arabidopsis thaliana DGAT1 highlighted contrasting substrate specificities when the recombinant yeast was cultured in lauric acid supplemented medium. The EgDGAT1-1 expressing strain preferentially accumulated medium-chain triacylglycerols whereas AtDGAT1 expression induced long-chain triacylglycerol storage in Y. lipolytica. EgDGAT1-1 localized to the endoplasmic reticulum where TAG biosynthesis takes place. Reestablishing neutral lipid accumulation in the Y. lipolytica mutant strain did not induce major reorganization of the yeast microsomal proteome. Overall, our findings demonstrate that EgDGAT1-1 is an endoplasmic reticulum DGAT with preference for medium-chain fatty acid substrates, in line with its physiological role in palm kernel. The characterized EgDGAT1-1 could be used to promote medium-chain triacylglycerol accumulation in microbial-produced oil for industrial chemicals and cosmetics.

Molecular Characterization of the Elaeis guineensis Medium-Chain Fatty Acid Diacylglycerol Acyltransferase DGAT1-1 by Heterologous Expression in Yarrowia lipolytica

PubMed Central

Aymé, Laure; Jolivet, Pascale; Nicaud, Jean-Marc; Chardot, Thierry

2015-01-01

Diacylglycerol acyltransferases (DGAT) are involved in the acylation of sn-1,2-diacylglycerol. Palm kernel oil, extracted from Elaeis guineensis (oil palm) seeds, has a high content of medium-chain fatty acids mainly lauric acid (C12:0). A putative E. guineensis diacylglycerol acyltransferase gene (EgDGAT1-1) is expressed at the onset of lauric acid accumulation in the seed endosperm suggesting that it is a determinant of medium-chain triacylglycerol storage. To test this hypothesis, we thoroughly characterized EgDGAT1-1 activity through functional complementation of a Yarrowia lipolytica mutant strain devoid of neutral lipids. EgDGAT1-1 expression is sufficient to restore triacylglycerol accumulation in neosynthesized lipid droplets. A comparative functional study with Arabidopsis thaliana DGAT1 highlighted contrasting substrate specificities when the recombinant yeast was cultured in lauric acid supplemented medium. The EgDGAT1-1 expressing strain preferentially accumulated medium-chain triacylglycerols whereas AtDGAT1 expression induced long-chain triacylglycerol storage in Y. lipolytica. EgDGAT1-1 localized to the endoplasmic reticulum where TAG biosynthesis takes place. Reestablishing neutral lipid accumulation in the Y. lipolytica mutant strain did not induce major reorganization of the yeast microsomal proteome. Overall, our findings demonstrate that EgDGAT1-1 is an endoplasmic reticulum DGAT with preference for medium-chain fatty acid substrates, in line with its physiological role in palm kernel. The characterized EgDGAT1-1 could be used to promote medium-chain triacylglycerol accumulation in microbial-produced oil for industrial chemicals and cosmetics. PMID:26581109

Novel Roles of the Non-catalytic Elements of Yeast Protein-disulfide Isomerase in Its Interplay with Endoplasmic Reticulum Oxidoreductin 1*

PubMed Central

Niu, Yingbo; Zhang, Lihui; Yu, Jiaojiao; Wang, Chih-chen; Wang, Lei

2016-01-01

The formation of disulfide bonds in the endoplasmic reticulum (ER) of eukaryotic cells is catalyzed by the sulfhydryl oxidase, ER oxidoreductin 1 (Ero1), and protein-disulfide isomerase (PDI). PDI is oxidized by Ero1 to continuously introduce disulfides into substrates, and feedback regulates Ero1 activity by manipulating the regulatory disulfides of Ero1. In this study we find that yeast Ero1p is enzymatically active even with its regulatory disulfides intact, and further activation of Ero1p by reduction of the regulatory disulfides requires the reduction of non-catalytic Cys90-Cys97 disulfide in Pdi1p. The principal client-binding site in the Pdi1p b′ domain is necessary not only for the functional Ero1p-Pdi1p disulfide relay but also for the activation of Ero1p. We also demonstrate by complementary activation assays that the regulatory disulfides in Ero1p are much more stable than those in human Ero1α. These new findings on yeast Ero1p-Pdi1p interplay reveal significant differences from our previously identified mode of human Ero1α-PDI interplay and provide insights into the evolution of the eukaryotic oxidative protein folding pathway. PMID:26846856

The Cytoplasmic Region of α-1,6-Mannosyltransferase Mnn9p Is Crucial for Retrograde Transport from the Golgi Apparatus to the Endoplasmic Reticulum in Saccharomyces cerevisiae▿ †

PubMed Central

Okamoto, Michiyo; Yoko-o, Takehiko; Miyakawa, Tokichi; Jigami, Yoshifumi

2008-01-01

In Saccharomyces cerevisiae, Och1p and Mnn9p mannosyltransferases are localized in the cis-Golgi. Attempts to live image Och1p and Mnn9p tagged with green fluorescent protein or red fluorescent protein, respectively, using a high-performance confocal laser scanning microscope system resulted in simultaneous visualization of the native proteins in a living cell. Our observations revealed that Och1p and Mnn9p are not always colocalized to the same cisternae. The difference in the dynamics of these mannosyltransferases may reflect differences in the mechanisms for their retention in the cis-Golgi, since it has been reported that Mnn9p cycles between the endoplasmic reticulum and the cis-Golgi whereas Och1p does not (Z. Todorow, A. Spang, E. Carmack, J. Yates, and R. Schekman, Proc. Natl. Acad. Sci. USA 97:13643-13648, 2000). We investigated the localization of chimeric proteins of Mnn9p and Och1p in sec12 and erd1 mutant cells. A chimeric protein, M16/O16, which consists of the N-terminal cytoplasmic region of Mnn9p and the transmembrane and luminal region of Och1p, behaved like Mnn9p, suggesting that the N-terminal cytoplasmic region is important for the intracellular dynamics of Mnn9p. This observation is supported by results from subcellular-fractionation experiments. Mutational analysis revealed that two arginine residues in the N-terminal region of Mnn9p are important for the chimeric protein to cycle between the endoplasmic reticulum and the Golgi apparatus. PMID:18083825

Acetic acid induces Sch9p-dependent translocation of Isc1p from the endoplasmic reticulum into mitochondria.

PubMed

Rego, António; Cooper, Katrina F; Snider, Justin; Hannun, Yusuf A; Costa, Vítor; Côrte-Real, Manuela; Chaves, Susana R

2018-06-01

Changes in sphingolipid metabolism have been linked to modulation of cell fate in both yeast and mammalian cells. We previously assessed the role of sphingolipids in cell death regulation using a well characterized yeast model of acetic acid-induced regulated cell death, finding that Isc1p, inositol phosphosphingolipid phospholipase C, plays a pro-death role in this process. Indeed, isc1∆ mutants exhibited a higher resistance to acetic acid associated with reduced mitochondrial alterations. Here, we show that Isc1p is regulated by Sch9p under acetic acid stress, since both single and double mutants lacking Isc1p or/and Sch9p have the same resistant phenotype, and SCH9 deletion leads to a higher retention of Isc1p in the endoplasmic reticulum upon acetic acid exposure. We also found that the higher resistance of all mutants correlates with higher levels of endogenous mitochondrial phosphorylated long chain bases (LCBPs), suggesting that changing the sphingolipid balance in favour of LCBPs in mitochondria results in increased survival to acetic acid. In conclusion, our results suggest that Sch9p pathways modulate acetic acid-induced cell death, through the regulation of Isc1p cellular distribution, thus affecting the sphingolipid balance that regulates cell fate. Copyright © 2018 Elsevier B.V. All rights reserved.

Cytoplasmic Nucleation and Atypical Branching Nucleation Generate Endoplasmic Microtubules in Physcomitrella patens[OPEN

PubMed Central

Nakaoka, Yuki; Kimura, Akatsuki; Tani, Tomomi; Goshima, Gohta

2015-01-01

The mechanism underlying microtubule (MT) generation in plants has been primarily studied using the cortical MT array, in which fixed-angled branching nucleation and katanin-dependent MT severing predominate. However, little is known about MT generation in the endoplasm. Here, we explored the mechanism of endoplasmic MT generation in protonemal cells of Physcomitrella patens. We developed an assay that utilizes flow cell and oblique illumination fluorescence microscopy, which allowed visualization and quantification of individual MT dynamics. MT severing was infrequently observed, and disruption of katanin did not severely affect MT generation. Branching nucleation was observed, but it showed markedly variable branch angles and was occasionally accompanied by the transport of nucleated MTs. Cytoplasmic nucleation at seemingly random locations was most frequently observed and predominated when depolymerized MTs were regrown. The MT nucleator γ-tubulin was detected at the majority of the nucleation sites, at which a single MT was generated in random directions. When γ-tubulin was knocked down, MT generation was significantly delayed in the regrowth assay. However, nucleation occurred at a normal frequency in steady state, suggesting the presence of a γ-tubulin-independent backup mechanism. Thus, endoplasmic MTs in this cell type are generated in a less ordered manner, showing a broader spectrum of nucleation mechanisms in plants. PMID:25616870

The endoplasmic reticulum is a hub to sort proteins toward unconventional traffic pathways and endosymbiotic organelles.

PubMed

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.

A plasma membrane-type Ca[sup 2+]-ATPase of 120 kilodaltons on the endoplasmic reticulum from carrot (Daucus carota) cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chen, F.H.; Ratterman, D.M.; Sze, H.

1993-06-01

Cytosolic Ca[sup 2+] levels are regulated in part by Ca[sup 2+]-pumping ATPases that export Ca[sup 2+] from the cytoplasm; The types and properties of Ca[sup 2+] pumps in plants are not well understood. The kinetic properties of a 120-kD phosphoenzyme (PE) intermediate formed during the reaction cycle of a Ca[sup 2+]-ATPase from suspension-cultured carrot (Daucus carota) cells are characterized. Only one Ca[sup 2+]-dependent phosphoprotein was formed when carrot membrane vesicles were incubated with [[gamma]-[sup 32]P]ATP. Formation of this 120-kD phosphoprotein was inhibited by vanadate, enhanced by La[sup 3+], and decreased by hydroxylamine, confirming its identification as an intermediate of amore » phosphorylated-type Ca[sup 2+]-translocating ATPase. The 120-kD Ca[sup 2+]-ATPase was most abundant in endoplasmic reticulum-enriched fractions, in which the Ca[sup 2+]-ATPase was estimated to be 0.1% of membrane protein. Direct quantitation of Ca[sup 2+]-dependent phosphoprotein was used to examine the kinetics of PE formation. PE formation exhibited a K[sub m] for Ca[sup 2+] of 1 to 2 [mu]m and a K[sub m] for ATP of 67 nm. Relative affinities of substrates, determined by competition experiments, were 0.075 [mu]m for ATP, 1 [mu]m for ADP, 100 [mu]m for ITP, and 250 [mu]m for GTP. Thapsigargin and cyclopiazonic acid, specific inhibitors of animal sarcoplasmic/endoplasmic reticulum Ca[sup 2+]-ATPase, had no effect on PE formation; erythrosin B inhibited with 50% inhibition at <0.1 [mu]m. Calmodulin (1 [mu]m) stimulated PE formation by 25%. The results indicate that the carrot 120-kD Ca[sup 2+]-ATPase is similar but not identical to animal plasma membrane-type Ca[sup 2+]-ATPase and yet is located on endomembranes, such as the endoplasmic reticulum. This type of Ca[sup 2+] pump may reside on the cortical endoplasmic reticulum, thought to play a major role in anchoring the cytoskeleton and in facilitating secretion. 34 refs., 9 figs., 3 tabs.« less

Survival of atraumatic restorative treatment (ART) sealants and restorations: a meta-analysis.

PubMed

de Amorim, Rodrigo G; Leal, Soraya C; Frencken, Jo E

2012-04-01

The purpose of this study is to perform a systematic investigation plus meta-analysis into survival of atraumatic restorative treatment (ART) sealants and restorations using high-viscosity glass ionomers and to compare the results with those from the 2005 ART meta-analysis. Until February 2010, four databases were searched. Two hundred four publications were found, and 66 reported on ART restorations or sealant survival. Based on five exclusion criteria, two independent reviewers selected the 29 publications that accounted for the meta-analysis. Confidence intervals (CI) and or standard errors were calculated and the heterogeneity variance of the survival rates was estimated. Location (school/clinic) was an independent variable. The survival rates of single-surface and multiple-surface ART restorations in primary teeth over the first 2 years were 93% (CI, 91-94%) and 62% (CI, 51-73%), respectively; for single-surface ART restorations in permanent teeth over the first 3 and 5 years it was 85% (CI, 77-91%) and 80% (CI, 76-83%), respectively and for multiple-surface ART restorations in permanent teeth over 1 year it was 86% (CI, 59-98%). The mean annual dentine lesion incidence rate, in pits and fissures previously sealed using ART, over the first 3 years was 1%. No location effect and no differences between the 2005 and 2010 survival rates of ART restorations and sealants were observed. The short-term survival rates of single-surface ART restorations in primary and permanent teeth, and the caries-preventive effect of ART sealants were high. ART can safely be used in single-surface cavities in both primary and permanent teeth. ART sealants have a high caries preventive effect.

Retention of glucose units added by the UDP-GLC:glycoprotein glucosyltransferase delays exit of glycoproteins from the endoplasmic reticulum

PubMed Central

1995-01-01

It has been proposed that the UDP-Glc:glycoprotein glucosyltransferase, an endoplasmic reticulum enzyme that only glucosylates improperly folded glycoproteins forming protein-linked Glc1Man7-9-GlcNAc2 from the corresponding unglucosylated species, participates together with lectin- like chaperones that recognize monoglucosylated oligosaccharides in the control mechanism by which cells only allow passage of properly folded glycoproteins to the Golgi apparatus. Trypanosoma cruzi cells were used to test this model as in trypanosomatids addition of glucosidase inhibitors leads to the accumulation of only monoglucosylated oligosaccharides, their formation being catalyzed by the UDP- Glc:glycoprotein glucosyltransferase. In all other eukaryotic cells the inhibitors produce underglycosylation of proteins and/or accumulation of oliogosaccharides containing two or three glucose units. Cruzipain, a lysosomal proteinase having three potential N-glycosylation sites, two at the catalytic domain and one at the COOH-terminal domain, was isolated in a glucosylated form from cells grown in the presence of the glucosidase II inhibitor 1-deoxynojirimycin. The oligosaccharides present at the single glycosylation site of the COOH-terminal domain were glucosylated in some cruzipain molecules but not in others, this result being consistent with an asynchronous folding of glycoproteins in the endoplasmic reticulum. In spite of not affecting cell growth rate or the cellular general metabolism in short and long term incubations, 1-deoxynojirimycin caused a marked delay in the arrival of cruzipain to lysosomes. These results are compatible with the model proposed by which monoglucosylated glycoproteins may be transiently retained in the endoplasmic reticulum by lectin-like anchors recognizing monoglucosylated oligosaccharides. PMID:7642696

Restoring proximal caries lesions conservatively with tunnel restorations

PubMed Central

Chu, Chun-Hung; Mei, May L; Cheung, Chloe; Nalliah, Romesh P

2013-01-01

The tunnel restoration has been suggested as a conservative alternative to the conventional box preparation for treating proximal caries. The main advantage of tunnel restoration over the conventional box or slot preparation includes being more conservative and increasing tooth integrity and strength by preserving the marginal ridge. However, tunnel restoration is technique-sensitive and can be particularly challenging for inexperienced restorative dentists. Recent advances in technology, such as the contemporary design of dental handpieces with advanced light-emitting diode (LED) and handheld comfort, offer operative dentists better vision, illumination, and maneuverability. The use of magnifying loupes also enhances the visibility of the preparation. The advent of digital radiographic imaging has improved dental imaging and reduced radiation. The new generation of restorative materials has improved mechanical properties. Tunnel restoration can be an option to restore proximal caries if the dentist performs proper case selection and pays attention to the details of the restorative procedures. This paper describes the clinical technique of tunnel restoration and reviews the studies of tunnel restorations. PMID:24019754

Restoring proximal caries lesions conservatively with tunnel restorations.

PubMed

Chu, Chun-Hung; Mei, May L; Cheung, Chloe; Nalliah, Romesh P

2013-07-30

The tunnel restoration has been suggested as a conservative alternative to the conventional box preparation for treating proximal caries. The main advantage of tunnel restoration over the conventional box or slot preparation includes being more conservative and increasing tooth integrity and strength by preserving the marginal ridge. However, tunnel restoration is technique-sensitive and can be particularly challenging for inexperienced restorative dentists. Recent advances in technology, such as the contemporary design of dental handpieces with advanced light-emitting diode (LED) and handheld comfort, offer operative dentists better vision, illumination, and maneuverability. The use of magnifying loupes also enhances the visibility of the preparation. The advent of digital radiographic imaging has improved dental imaging and reduced radiation. The new generation of restorative materials has improved mechanical properties. Tunnel restoration can be an option to restore proximal caries if the dentist performs proper case selection and pays attention to the details of the restorative procedures. This paper describes the clinical technique of tunnel restoration and reviews the studies of tunnel restorations.

XBP-1 Regulates a Subset of Endoplasmic Reticulum Resident Chaperone Genes in the Unfolded Protein Response

PubMed Central

Lee, Ann-Hwee; Iwakoshi, Neal N.; Glimcher, Laurie H.

2003-01-01

The mammalian unfolded protein response (UPR) protects the cell against the stress of misfolded proteins in the endoplasmic reticulum (ER). We have investigated here the contribution of the UPR transcription factors XBP-1, ATF6α, and ATF6β to UPR target gene expression. Gene profiling of cell lines lacking these factors yielded several XBP-1-dependent UPR target genes, all of which appear to act in the ER. These included the DnaJ/Hsp40-like genes, p58IPK, ERdj4, and HEDJ, as well as EDEM, protein disulfide isomerase-P5, and ribosome-associated membrane protein 4 (RAMP4), whereas expression of BiP was only modestly dependent on XBP-1. Surprisingly, given previous reports that enforced expression of ATF6α induced a subset of UPR target genes, cells deficient in ATF6α, ATF6β, or both had minimal defects in upregulating UPR target genes by gene profiling analysis, suggesting the presence of compensatory mechanism(s) for ATF6 in the UPR. Since cells lacking both XBP-1 and ATF6α had significantly impaired induction of select UPR target genes and ERSE reporter activation, XBP-1 and ATF6α may serve partially redundant functions. No UPR target genes that required ATF6β were identified, nor, in contrast to XBP-1 and ATF6α, did the activity of the UPRE or ERSE promoters require ATF6β, suggesting a minor role for it during the UPR. Collectively, these results suggest that the IRE1/XBP-1 pathway is required for efficient protein folding, maturation, and degradation in the ER and imply the existence of subsets of UPR target genes as defined by their dependence on XBP-1. Further, our observations suggest the existence of additional, as-yet-unknown, key regulators of the UPR. PMID:14559994

Toll-like receptor 4-induced endoplasmic reticulum stress contributes to endothelial dysfunction

USDA-ARS?s Scientific Manuscript database

Impairment of vasodilator action of insulin is associated with endothelial dysfunction and insulin resistance. Endoplasmic reticulum (ER) stress is implicated as one of the mechanisms for pathophysiology of various cardiometabolic syndromes, including insulin resistance and endothelial dysfunction. ...

Gene therapy restores vision in rd1 mice after removal of a confounding mutation in Gpr179.

PubMed

Nishiguchi, Koji M; Carvalho, Livia S; Rizzi, Matteo; Powell, Kate; Holthaus, Sophia-Martha kleine; Azam, Selina A; Duran, Yanai; Ribeiro, Joana; Luhmann, Ulrich F O; Bainbridge, James W B; Smith, Alexander J; Ali, Robin R

2015-01-23

The rd1 mouse with a mutation in the Pde6b gene was the first strain of mice identified with a retinal degeneration. However, AAV-mediated gene supplementation of rd1 mice only results in structural preservation of photoreceptors, and restoration of the photoreceptor-mediated a-wave, but not in restoration of the bipolar cell-mediated b-wave. Here we show that a mutation in Gpr179 prevents the full restoration of vision in rd1 mice. Backcrossing rd1 with C57BL6 mice reveals the complete lack of b-wave in a subset of mice, consistent with an autosomal recessive Mendelian inheritance pattern. We identify a mutation in the Gpr179 gene, which encodes for a G-protein coupled receptor localized to the dendrites of ON-bipolar cells. Gene replacement in rd1 mice that are devoid of the mutation in Gpr179 successfully restores the function of both photoreceptors and bipolar cells, which is maintained for up to 13 months. Our discovery may explain the failure of previous gene therapy attempts in rd1 mice, and we propose that Grp179 mutation status should be taken into account in future studies involving rd1 mice.

Wildlife Habitat Restoration: Chapter 12

USGS Publications Warehouse

Conway, Courtney J.; Borgmann, Kathi L.; Morrison, Michael L.; Mathewson, Heather A.

2015-01-01

As the preceding chapters point out, many wildlife species and the habitat they depend on are in peril. However, opportunities exist to restore habitat for many imperiled wildlife species. But what is wildlife habitat restoration? We begin this chapter by defining habitat restoration and then provide recommendations on how to maximize success of future habitat restoration efforts for wildlife. Finally, we evaluate whether we have been successful in restoring wildlife habitat and supply recommendations to advance habitat restoration. Successful restoration requires clear and explicit goals that are based on our best understanding of what the habitat was like prior to the disturbing event. Ideally, a restoration project would include: (1) a summary of prerestoration conditions that define the existing status of wildlife populations and their habitat; (2) a description of habitat features required by the focal or indicator species for persistence; (3) an a priori description of measurable, quantitative metrics that define restoration goals and measures of success; (4) a monitoring plan; (5) postrestoration comparisons of habitat features and wildlife populations with adjacent unmodified areas that are similar to the restoration site; and (6) expert review of the entire restoration plan (i.e., the five aforementioned components).

Hydrogen alleviates hyperoxic acute lung injury related endoplasmic reticulum stress in rats through upregulation of SIRT1.

PubMed

Sun, Qiang; Han, Wenjie; Hu, Huijun; Fan, Danfeng; Li, Yanbo; Zhang, Yu; Lv, Yan; Li, Mingxin; Pan, Shuyi

2017-06-01

Hyperoxic acute lung injury (HALI) is a major clinical problem for patients undergoing supplemental oxygen therapy. Currently in clinical settings there exist no effective means of prevention or treatment methods. Our previous study found that: hydrogen could reduce HALI, as well as oxidative stress. This research will further explore the mechanism underlying the protective effect of hydrogen on oxygen toxicity. Rats were randomly assigned into three experimental groups and were exposed in a oxygen chamber for 60 continuous hours: 100% balanced air (control); 100% oxygen (HALI); 100% oxygen with hydrogen treatment (HALI + HRS). We examined lung function by wet to dry ratio of lung, lung pleural effusion and cell apoptosis. We also detected endoplasmic reticulum stress (ERS) by examining the expression of CHOP, GRP78 and XBP1. We further investigated the role of Sirtuin 1 (SIRT1) in HALI, which contributes to cellular regulation including ERS, by examining its expression after hydrogen treatment with SIRT1 inhibitor. Hydrogen could significantly reduce HALI by reducing lung edema and apoptosis, inhibiting the elevating of ERS and increased SIRT1 expression. By inhibition of SIRT1 expression, the effect of hydrogen on prevention of HALI is significantly weakened, the inhibition of the ERS was also reversed. Our findings indicate that hydrogen could reduce HALI related ERS and the mechanism of hydrogen may be associated with upregulation of SIRT1, this study reveals the molecular mechanisms underlying the protective effect of hydrogen, which provides a new theoretical basis for clinical application of hydrogen.

Partial-coverage posterior ceramic restorations. Part 1: a return to diligence.

PubMed

Liebenberg, W H

2001-01-01

The application of multisurfaced tooth-colored restorations in the posterior dentition is an exercise in risk tolerance when dentin occupies the bulk of the tooth substrate. Not only is interfacial integrity capricious, but also a recent in vivo study has confirmed that dentin bond strengths deteriorate with time. Although the literature is replete with esthetic guidelines for posterior restitution, most practicing clinicians appreciate the prime tenet that clinical success involves more than esthetic realism in the posterior dentition. Success with indirect ceramic restorations is dependent on interfacial integrity, which, although multitudinous, is contingently related to operative competence. Innovative clinical techniques are described in this two-part article, along with a discussion of the probationary status of current adhesive options and the need for excellence in all phases of this demanding restorative sequence. Restorative success in the posterior dentition is profoundly influenced by the variability of operative competence and diligence. This article discusses the precincts of posterior indirect ceramic restorations and submits a number of innovative solutions to the clinical challenge.

Calcium transport in tonoplast and endoplasmic reticulum vesicles isolated from cultured carrot cells. [Daucus carota Danvers

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bush, D.R.; Sze, H.

1986-02-01

Two active calcium (Ca/sup 2 +/) transport systems have been identified and partially characterized in membrane vesicles isolated from cultured carrot cells (Daucus carota Danvers). Both transport systems required MgATP for activity and were enhanced by 10 millimolar oxalate. Ca/sup 2 +/ transport in membrane vesicles derived from isolated vacuoles equilibrated at 1.10 grams per cubic centimeter and comigrated with Cl/sup -/-stimulated, NO/sub 3//sup -/-inhibited ATPase activity on sucrose density gradients. Ca/sup 2 +/ transport in this system was insensitive to vanadate, but was inhibited by nitrate, carbonyl cyanide-m-chlorophenylhydrazone (CCCP), N,N'-dicyclohexylcarbodiimide (DCCD), and 4,4-diisothiocyano-2,2'-stilbene disulfonic acid (DIDS). The K/sub m/more » for MgATP and Ca/sup 2 +/ were 0.1 mM and 21 micromolar, respectively. The predominant Ca/sup 2 +/ transport system detectable in microsomal membrane preparations equilibrated at a density of 1.13 grams per cubic centimeter and comigrated with the endoplasmic reticulum (ER) marker, antimycin A-insensitive NADH-dependent cytochrome c reductase. Ca/sup 2 +/ transport activity and the ER marker also shifted in parallel in ER shifting experiments. This transport system was inhibited by vanadate (I/sub 50/ = 12 micromolar) and was insensitive to nitrate, CCCP, DCCD, and DIDS. Transport exhibited cooperative MgATP dependent kinetics. Ca/sup 2 +/ dependent kinetics were complex with an apparent K/sub m/ ranging from 0.7 to 2 micromolar. We conclude that the vacuolar-derived system is a Ca/sup 2 +//H/sup +/ antiport located on the tonoplast and that the microsomal transport system is a Ca,Mg-ATPase enriched on the ER. These two Ca/sup 2 +/ transport systems are proposed to restore and maintain cytoplasmic Ca/sup 2 +/ homeostasis under changing cellular and environmental conditions.« less

Caspase-12 is involved in stretch-induced apoptosis mediated endoplasmic reticulum stress.

PubMed

Zhang, Qiang; Liu, Jianing; Chen, Shulan; Liu, Jing; Liu, Lijuan; Liu, Guirong; Wang, Fang; Jiang, Wenxin; Zhang, Caixia; Wang, Shuangyu; Yuan, Xiao

2016-04-01

It is well recognized that mandibular growth, which is caused by a variety of functional appliances, is considered to be the result of both neuromuscular and skeletal adaptations. Accumulating evidence has demonstrated that apoptosis plays an important role in the adaptation of skeletal muscle function. However, the underlying mechanism of apoptosis that is induced by stretch continues to be incompletely understood. Endoplasmic reticulum stress (ERS), a newly defined signaling pathway, initiates apoptosis. This study seeks to determine if caspase-12 is involved in stretch-induced apoptosis mediated endoplasmic reticulum stress in myoblast and its underlying mechanism. Apoptosis was assessed by Hochest staining, DAPI staining and annexin V binding and PI staining. ER chaperones, such as GRP78, CHOP and caspase-12, were determined by reverse transcription polymerase chain reaction (RT-PCR) and Western blot. Furthermore, caspase-12 inhibitor was used to value the mechanism of the caspase-12 pathway. Apoptosis of myoblast, which is subjected to cyclic stretch, was observed in a time-dependent manner. We found that GRP78 mRNA and protein were significantly increased and CHOP and caspase-12 were activated in myoblast that was exposed to cyclic stretch. Caspase-12 inhibition reduced stretch-induced apoptosis, and caspase-12 activated caspase-3 to induce apoptosis. We concluded that caspase-12 played an important role in stretch-induced apoptosis that is associated by endoplasmic reticulum stress by activating caspase-3.

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

PubMed

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.

Installation Restoration Program. Phase 1 - Records Search, Elmendorf AFB, Alaska

DTIC Science & Technology

1983-09-01

Installation Restoration Hazardous Waste Management Past Solid Waste Disposal Sites Ground Water Contamination 26. ABSTRACT (CO- ffew. - reev. ilde It necessar...Activity Review 4-1 Industrial Operations (Shops) 4-2 Fire Training 4-13 Fuels Management 4-15 Description of Past On-sane Disposal Methods 4-23 Waste...characteristics, potential for contaminant migration and waste management practices. The details of the rating procedure are presented in Appendix H and the

The Endoplasmic Reticulum: A Central Player in Cell Signalling and Protein Synthesis

NASA Astrophysics Data System (ADS)

Llewelyn Roderick, H.; Berridge, Michael J.; Bootman, Martin D.

In addition to being the principle intracellular Ca 2+ store, the endoplasmic reticulum (ER) is the initial site of synthesis and folding of membrane and secretory proteins. These two roles of the ER are intimately linked. First, the function of many proteins involved in Ca 2+ handling are modulated by Ca 2+ and second, ER lumenal Ca 2+ modulates protein synthesis and folding. Within the ER, Ca 2+ is stored by low affinity high capacity Ca 2+ binding proteins and is maintained at a free concentration between 0.1 and 1 μM relative to 0.1 μM cytosolic Ca 2+. This concentration gradient is maintained by the action of the Sarco-Endoplasmic Reticulum ATPases (SERCa) which hydrolyse ATP to pump Ca 2+ into the ER. Following stimulation Ca 2+ is released from the ER through several classes of ligand gated channels. The most well characterized of these being the inositol 1,4,5-trisphosphate receptor ( IP 3 R) and the Ryanodine receptor (RyR) families of proteins. This release of Ca 2+ results in a drop of ER free Ca 2+ to levels as low as 10 μM. This decrease in lumenal Ca 2+ inhibits further release through the channels and increases the rate of re-sequestration of Ca 2+ into the ER by the SERCa pumps. Under these conditions, in addition to effects on Ca 2+ handling proteins, protein synthesis is inhibited, chaperones dissociate from their substrates, secondary modifications of proteins are inhibited and the retention of many proteins within the ER is lost. Furthermore, a signalling cascade resulting in the up-regulation of many proteins involved in protein folding and Ca 2+ homeostasis is initiated. This review will focus on the proteins involved in the regulation ER lumenal Ca 2+ and the role of ER lumenal Ca 2+ in cell signalling and protein synthesis.

Role of dorsal hippocampal orexin-1 receptors in memory restoration induced by morphine sensitization phenomenon.

PubMed

Alijanpour, S; Tirgar, F; Zarrindast, M-R

2016-01-15

The present study was examined the blockade of CA1 orexin-1 receptors (OX1Rs) of the dorsal hippocampus in the induction or expression phase on morphine sensitization-induced memory restoration using the Morris water maze (MWM) apparatus. Results showed that pre-training administration of morphine (5mg/kg, s.c.) increases escape latency and traveled distance, while does not alter swimming speed. This supports the impairing effect of morphine on the spatial memory acquisition in male adult rats. Also, in the retrieval session (probe trial) this treatment decreased the time spent in the target quadrant. Moreover, morphine-induced sensitization (15 or 20mg/kg, s.c.; once daily for 3days and followed by 5days no drug treatment) restored the memory acquisition/retrieval deficit which had been induced by pre-training administration of morphine (5mg/kg, s.c.). Intra-CA1 microinjection of subthreshold doses of SB-334867 (OX1Rs antagonist; 10, 20 and 40nmol/rat), 5min before morphine (20mg/kg/day×3days, s.c.; induction phase for morphine sensitization) did not alter restoration of memory acquisition/retrieval produced by the morphine sensitization phenomenon. In contrast, microinjection of subthreshold doses of SB-334867 (10, 20 and 40nmol/rat) into the CA1 region in the training session, 5min prior to morphine (5mg/kg, s.c.; expression phase for morphine sensitization) blocked the spatial memory acquisition/retrieval in morphine-sensitized rats. In conclusion, these findings show that morphine sensitization reverses morphine-induced amnesia. Furthermore, the blockade of CA1 OX1Rs in the expression phase, but not in the induction phase, disrupts memory restoration induced by morphine sensitization. Copyright © 2015 IBRO. Published by Elsevier Ltd. All rights reserved.

Identification and Functional Characterization of Hypoxia-Induced Endoplasmic Reticulum Stress Regulating lncRNA (HypERlnc) in Pericytes.

PubMed

Bischoff, Florian C; Werner, Astrid; John, David; Boeckel, Jes-Niels; Melissari, Maria-Theodora; Grote, Phillip; Glaser, Simone F; Demolli, Shemsi; Uchida, Shizuka; Michalik, Katharina M; Meder, Benjamin; Katus, Hugo A; Haas, Jan; Chen, Wei; Pullamsetti, Soni S; Seeger, Werner; Zeiher, Andreas M; Dimmeler, Stefanie; Zehendner, Christoph M

2017-08-04

Pericytes are essential for vessel maturation and endothelial barrier function. Long noncoding RNAs regulate many cellular functions, but their role in pericyte biology remains unexplored. Here, we investigate the effect of hypoxia-induced endoplasmic reticulum stress regulating long noncoding RNAs (HypERlnc, also known as ENSG00000262454) on pericyte function in vitro and its regulation in human heart failure and idiopathic pulmonary arterial hypertension. RNA sequencing in human primary pericytes identified hypoxia-regulated long noncoding RNAs, including HypERlnc. Silencing of HypERlnc decreased cell viability and proliferation and resulted in pericyte dedifferentiation, which went along with increased endothelial permeability in cocultures consisting of human primary pericyte and human coronary microvascular endothelial cells. Consistently, Cas9-based transcriptional activation of HypERlnc was associated with increased expression of pericyte marker genes. Moreover, HypERlnc knockdown reduced endothelial-pericyte recruitment in Matrigel assays ( P <0.05). Mechanistically, transcription factor reporter arrays demonstrated that endoplasmic reticulum stress-related transcription factors were prominently activated by HypERlnc knockdown, which was confirmed via immunoblotting for the endoplasmic reticulum stress markers IRE1α ( P <0.001), ATF6 ( P <0.01), and soluble BiP ( P <0.001). Kyoto encyclopedia of genes and gene ontology pathway analyses of RNA sequencing experiments after HypERlnc knockdown indicate a role in cardiovascular disease states. Indeed, HypERlnc expression was significantly reduced in human cardiac tissue from patients with heart failure ( P <0.05; n=19) compared with controls. In addition, HypERlnc expression significantly correlated with pericyte markers in human lungs derived from patients diagnosed with idiopathic pulmonary arterial hypertension and from donor lungs (n=14). Here, we show that HypERlnc regulates human pericyte function and the

Critical Role of Interdomain Interactions in the Conformational Change and Catalytic Mechanism of Endoplasmic Reticulum Aminopeptidase 1.

PubMed

Stamogiannos, Athanasios; Maben, Zachary; Papakyriakou, Athanasios; Mpakali, Anastasia; Kokkala, Paraskevi; Georgiadis, Dimitris; Stern, Lawrence J; Stratikos, Efstratios

2017-03-14

Endoplasmic reticulum aminopeptidase 1 (ERAP1) is an intracellular enzyme that is important for the generation of antigenic epitopes and major histocompatibility class I-restricted adaptive immune responses. ERAP1 processes a vast variety of different peptides but still shows length and sequence selectivity, although the mechanism behind these properties is poorly understood. X-ray crystallographic analysis has revealed that ERAP1 can assume at least two distinct conformations in which C-terminal domain IV is either proximal or distal to active site domain II. To improve our understanding of the role of this conformational change in the catalytic mechanism of ERAP1, we used site-directed mutagenesis to perturb key salt bridges between domains II and IV. Enzymatic analysis revealed that these mutations, although located away from the catalytic site, greatly reduce the catalytic efficiency and change the allosteric kinetic behavior. The variants were more efficiently activated by small peptides and bound a competitive inhibitor with weaker affinity and faster dissociation kinetics. Molecular dynamics analysis suggested that the mutations affect the conformational distribution of ERAP1, reducing the population of closed states. Small-angle X-ray scattering indicated that both the wild type and the ERAP1 variants are predominantly in an open conformational state in solution. Overall, our findings suggest that electrostatic interactions between domains II and IV in ERAP1 are crucial for driving a conformational change that regulates the structural integrity of the catalytic site. The extent of domain opening in ERAP1 probably underlies its specialization for antigenic peptide precursors and should be taken into account in inhibitor development efforts.

Sequestration of Mutated α1-Antitrypsin into Inclusion Bodies Is a Cell-protective Mechanism to Maintain Endoplasmic Reticulum Function

PubMed Central

Granell, Susana; Baldini, Giovanna; Mohammad, Sameer; Nicolin, Vanessa; Narducci, Paola; Storrie, Brian

2008-01-01

A variant α1-antitrypsin with E342K mutation has a high tendency to form intracellular polymers, and it is associated with liver disease. In the hepatocytes of individuals carrying the mutation, α1-antitrypsin localizes both to the endoplasmic reticulum (ER) and to membrane-surrounded inclusion bodies (IBs). It is unclear whether the IBs contribute to cell toxicity or whether they are protective to the cell. We found that in hepatoma cells, mutated α1-antitrypsin exited the ER and accumulated in IBs that were negative for autophagosomal and lysosomal markers, and contained several ER components, but not calnexin. Mutated α1-antitrypsin induced IBs also in neuroendocrine cells, showing that formation of these organelles is not cell type specific. In the presence of IBs, ER function was largely maintained. Increased levels of calnexin, but not of protein disulfide isomerase, inhibited formation of IBs and lead to retention of mutated α1-antitrypsin in the ER. In hepatoma cells, shift of mutated α1-antitrypsin localization to the ER by calnexin overexpression lead to cell shrinkage, ER stress, and impairment of the secretory pathway at the ER level. We conclude that segregation of mutated α1-antitrypsin from the ER to the IBs is a protective cell response to maintain a functional secretory pathway. PMID:18045994

Endoplasmic reticulum stress regulates tumor growth and anti-tumor immunity: a promising opportunity for cancer immunotherapy.

PubMed

Mohamed, Eslam; Cao, Yu; Rodriguez, Paulo C

2017-08-01

The endoplasmic reticulum (ER) stress is a cellular process that occurs as a consequence of several stress circumstances, such as the accumulation of unfolded proteins in the lumen of the ER or distinct insults that disturb the ER normal function. Different conditions in the tumor microenvironment (TME), including hypoxia, nutrient deprivation, and the elevated production of reactive oxygen and nitrogen species destabilize the loading and dispatching of the newly synthesized proteins, triggering ER stress in cancer cells and tumor-infiltrating leukocytes. In order to cope with TME-induced ER stress, tumor and stromal cells initiate an adaptive response process that aims to resolve ER stress and to restore cellular homeostasis, which is referred as the unfolded protein responses (UPR). Paradoxically, the UPR can also induce cell death under severe and/or permanent ER stress. The UPR is started through three mediators, the activation of the inositol-requiring enzyme-1α, the pancreatic ER kinase-like ER kinase, and the activating transcription factor 6. In this minireview, we will discuss the pro- and anti-tumorigenic role of the UPR in cancer cells. In addition, we will describe the effects of the TME-induced ER stress in the immunosuppressive activity of tumor-infiltrating myeloid cells. Also, we will review the results of emerging therapeutic interventions that target ER stress and the UPR mediators in cancer. We postulate that the inhibition of ER stress or the UPR-related elements could represent a significant approach to increase the efficacy of various forms of cancer immunotherapy.

Endoplasmic reticulum stress-induced apoptosis accompanies enhanced expression of multiple inositol polyphosphate phosphatase 1 (Minpp1): a possible role for Minpp1 in cellular stress response.

PubMed

Kilaparty, Surya P; Agarwal, Rakhee; Singh, Pooja; Kannan, Krishnaswamy; Ali, Nawab

2016-07-01

Inositol polyphosphates represent a group of differentially phosphorylated inositol metabolites, many of which are implicated to regulate diverse cellular processes such as calcium mobilization, vesicular trafficking, differentiation, apoptosis, etc. The metabolic network of these compounds is complex and tightly regulated by various kinases and phosphatases present predominantly in the cytosol. Multiple inositol polyphosphate phosphatase 1 (Minpp1) is the only known endoplasmic reticulum (ER) luminal enzyme that hydrolyzes various inositol polyphosphates in vitro as well as in vivo conditions. However, access of the Minpp1 to cytosolic substrates has not yet been demonstrated clearly and hence its physiological function. In this study, we examined a potential role for Minpp1 in ER stress-induced apoptosis. We generated a custom antibody and characterized its specificity to study the expression of Minpp1 protein in multiple mammalian cells under experimentally induced cellular stress conditions. Our results demonstrate a significant increase in the expression of Minpp1 in response to a variety of cellular stress conditions. The protein expression was corroborated with the expression of its mRNA and enzymatic activity. Further, in an attempt to link the role of Minpp1 to apoptotic stress, we studied the effect of Minpp1 expression on apoptosis following silencing of the Minpp1 gene by its specific siRNA. Our results suggest an attenuation of apoptotic parameters following knockdown of Minpp1. Thus, in addition to its known role in inositol polyphosphate metabolism, we have identified a novel role for Minpp1 as a stress-responsive protein. In summary, our results provide, for the first time, a probable link between ER stress-induced apoptosis and Minpp1 expression.

Redox cycling induced Ni exsolution in Gd0.1Ce0.8Ni0.1O2 - (Sr0.9La0.1)0.9Ti0.9Ni0.1O3 composite solid oxide fuel cell anodes

NASA Astrophysics Data System (ADS)

Shen, X.; Chen, T.; Bishop, S. R.; Perry, N. H.; Tuller, H. L.; Sasaki, K.

2017-12-01

Oxide anodes composed of 60 wt% Gd0.1Ce0.8Ni0.1O2 (GDCN)- 40 wt% (Sr0.9La0.1)0.9Ti0.9Ni0.1O3 (SLTN) composites were prepared and tested on (ZrO2)0.89(Sc2O3)0.1(CeO2)0.01 (SSZ) electrolyte-supported SOFC cells utilizing a (La0.75Sr0.25)0.98MnO3 (LSM)-SSZ cathode, in 3%-humidified hydrogen fuel at 800 °C. Improved electrochemical performance was found compared to the cell using Ni-free 60 wt% Gd0.1Ce0.9O2 (GDC) - 40 wt % Sr0.9La0.1TiO3 (SLT) that was attributed to the exsolution of nano-sized Ni particles from the Ni-doped system. This exsolution process represents a simpler, more attractive method to improve performance than the more conventional but more complicated infiltration method for introducing catalytic nanoparticles. Redox cycling testing was performed to investigate the performance and structural stability of the Ni-doped GDC-SLT anode. The results indicated that the Ni exsolution and aggregation occurred while redox cycling proceeded, resulting in a gradually reduced anodic overvoltage. Symmetric cells with dense thin film Gd0.1Ce0.9-xNixO2 (x = 0, 0.05, 0.1, 0.15) electrodes were also tested, demonstrating lower area-specific resistances with increasing Ni content on the surface under reducing conditions. The steady improvement during redox cycling, despite Ni agglomeration, is related to the continuous increase in the overall Ni content on the anode surface, which may be enabled by kinetic limitations to Ni re-dissolving under oxidizing transients.

Expression of a truncated form of the endoplasmic reticulum chaperone protein, σ1 receptor, promotes mitochondrial energy depletion and apoptosis.

PubMed

Shioda, Norifumi; Ishikawa, Kiyoshi; Tagashira, Hideaki; Ishizuka, Toru; Yawo, Hiromu; Fukunaga, Kohji

2012-07-06

The σ1 receptor (σ(1)R) regulates endoplasmic reticulum (ER)/mitochondrial interorganellar Ca(2+) mobilization through the inositol 1,4,5-trisphosphate receptor (IP(3)R). Here, we observed that expression of a novel splice variant of σ(1)R, termed short form σ(1)R (σ(1)SR), has a detrimental effect on mitochondrial energy production and cell survival. σ(1)SR mRNA lacks 47 ribonucleotides encoding exon 2, resulting in a frameshift and formation of a truncated receptor. σ(1)SR localizes primarily in the ER at perinuclear regions and forms a complex with σ(1)R but not with IP(3)R in the mitochondrion-associated ER membrane. Overexpression of both σ(1)R and the truncated isoform promotes mitochondrial elongation with increased ER mitochondrial contact surface. σ(1)R overexpression increases the efficiency of mitochondrial Ca(2+) uptake in response to IP(3)R-driven stimuli, whereas σ(1)SR overexpression reduces it. Most importantly, σ(1)R promotes ATP production via increased mitochondrial Ca(2+) uptake, promoting cell survival in the presence of ER stress. By contrast, σ(1)SR suppresses ATP production following ER stress, enhancing cell death. Taken together, the newly identified σ(1)SR isoform interferes with σ(1)R function relevant to mitochondrial energy production under ER stress conditions, promoting cellular apoptosis.

Ricin A chain reaches the endoplasmic reticulum after endocytosis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu Qiong; Department of Biochemistry and Molecular Biology, Ningbo University Medical School, Ningbo 315211; Zhan Jinbiao

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 purifiedmore » 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.« less

The Endoplasmic Reticulum Coat Protein II Transport Machinery Coordinates Cellular Lipid Secretion and Cholesterol Biosynthesis*

PubMed Central

Fryer, Lee G. D.; Jones, Bethan; Duncan, Emma J.; Hutchison, Claire E.; Ozkan, Tozen; Williams, Paul A.; Alder, Olivia; Nieuwdorp, Max; Townley, Anna K.; Mensenkamp, Arjen R.; Stephens, David J.; Dallinga-Thie, Geesje M.; Shoulders, Carol C.

2014-01-01

Triglycerides and cholesterol are essential for life in most organisms. Triglycerides serve as the principal energy storage depot and, where vascular systems exist, as a means of energy transport. Cholesterol is essential for the functional integrity of all cellular membrane systems. The endoplasmic reticulum is the site of secretory lipoprotein production and de novo cholesterol synthesis, yet little is known about how these activities are coordinated with each other or with the activity of the COPII machinery, which transports endoplasmic reticulum cargo to the Golgi. The Sar1B component of this machinery is mutated in chylomicron retention disorder, indicating that this Sar1 isoform secures delivery of dietary lipids into the circulation. However, it is not known why some patients with chylomicron retention disorder develop hepatic steatosis, despite impaired intestinal fat malabsorption, and why very severe hypocholesterolemia develops in this condition. Here, we show that Sar1B also promotes hepatic apolipoprotein (apo) B lipoprotein secretion and that this promoting activity is coordinated with the processes regulating apoB expression and the transfer of triglycerides/cholesterol moieties onto this large lipid transport protein. We also show that although Sar1A antagonizes the lipoprotein secretion-promoting activity of Sar1B, both isoforms modulate the expression of genes encoding cholesterol biosynthetic enzymes and the synthesis of cholesterol de novo. These results not only establish that Sar1B promotes the secretion of hepatic lipids but also adds regulation of cholesterol synthesis to Sar1B's repertoire of transport functions. PMID:24338480

Cardiac Ablation of Rheb1 Induces Impaired Heart Growth, Endoplasmic Reticulum-Associated Apoptosis and Heart Failure in Infant Mice

PubMed Central

Cao, Yunshan; Tao, Lichan; Shen, Shutong; Xiao, Junjie; Wu, Hang; Li, Beibei; Wu, Xiangqi; Luo, Wen; Xiao, Qi; Hu, Xiaoshan; Liu, Hailang; Nie, Junwei; Lu, Shuangshuang; Yuan, Baiyin; Han, Zhonglin; Xiao, Bo; Yang, Zhongzhou; Li, Xinli

2013-01-01

Ras homologue enriched in brain 1 (Rheb1) plays an important role in a variety of cellular processes. In this study, we investigate the role of Rheb1 in the post-natal heart. We found that deletion of the gene responsible for production of Rheb1 from cardiomyocytes of post-natal mice resulted in malignant arrhythmias, heart failure, and premature death of these mice. In addition, heart growth impairment, aberrant metabolism relative gene expression, and increased cardiomyocyte apoptosis were observed in Rheb1-knockout mice prior to the development of heart failure and arrhythmias. Also, protein kinase B (PKB/Akt) signaling was enhanced in Rheb1-knockout mice, and removal of phosphatase and tensin homolog (Pten) significantly prolonged the survival of Rheb1-knockouts. Furthermore, signaling via the mammalian target of rapamycin complex 1 (mTORC1) was abolished and C/EBP homologous protein (CHOP) and phosphorylation levels of c-Jun N-terminal kinase (JNK) were increased in Rheb1 mutant mice. In conclusion, this study demonstrates that Rheb1 is important for maintaining cardiac function in post-natal mice via regulation of mTORC1 activity and stress on the endoplasmic reticulum. Moreover, activation of Akt signaling helps to improve the survival of mice with advanced heart failure. Thus, this study provides direct evidence that Rheb1 performs multiple important functions in the heart of the post-natal mouse. Enhancing Akt activity improves the survival of infant mice with advanced heart failure. PMID:24351823

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

USGS Publications Warehouse

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.

Marker-assisted identification of restorer gene(s) in iso-cytoplasmic restorer lines of WA cytoplasm in rice and assessment of their fertility restoration potential across environments.

PubMed

Kumar, Amit; Bhowmick, Prolay Kumar; Singh, Vikram Jeet; Malik, Manoj; Gupta, Ashish Kumar; Seth, R; Nagarajan, M; Krishnan, S Gopala; Singh, Ashok Kumar

2017-10-01

Iso-cytoplasmic restorers possess the same male sterile cytoplasm as the cytoplasmic male sterile (CMS) lines, thereby minimizing the potential cyto-nuclear conflict in the hybrids. Restoration of fertility of the wild abortive CMS is governed by two major genes namely, Rf3 and Rf4 . Therefore, assessing the allelic status of these restorer genes in the iso-cytoplasmic restorers using molecular markers will not only help in estimating the efficiency of these genes either alone or in combination, in fertility restoration in the hybrids in different environments, but will also be useful in determining the efficacy of these markers. In the present study, the efficiency of molecular markers in identifying genotypes carrying restorer allele of the gene(s) Rf3 and Rf4, restoring male fertility of WA cytoplasm in rice was assessed in a set of 100 iso-cytoplasmic rice restorers using gene linked as well as candidate gene based markers. In order to validate the efficacy of markers in identifying the restorers, a sub-set of selected 25 iso-cytoplasmic rice restorers were crossed with four different cytoplasmic male sterile lines namely, IR 79156A, IR 58025A, Pusa 6A and RTN 12A, and the pollen and spikelet fertility of the F 1 s were evaluated at three different locations. Marker analysis showed that Rf4 was the predominant fertility restorer gene in the iso-cytoplasmic restorers and Rf3 had a synergistic effect on fertility restoration. The efficiency of gene based markers, DRCG-RF4-14 and DRRM-RF3-10 for Rf4 (87%) and Rf3 (84%) genes was higher than respective gene-linked SSR markers RM6100 (80%) and RM3873 (82%). It is concluded that the gene based markers can be effectively used in identifying fertility restorer lines obviating the need for making crosses and evaluating the F 1 s. Though gene based markers are more efficient, there is a need to identify functional polymorphisms which can provide 100% efficiency. Three iso-cytoplasmic restorers namely, PRR 300, PRR 363

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

PubMed

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.

In vitro studies on the translocation of acid phosphatase into the endoplasmic reticulum of the yeast Saccharomyces cerevisiae.

PubMed

Krebs, H O; Hoffschulte, H K; Müller, M

1989-05-01

We demonstrate here the in vitro translocation of yeast acid phosphatase into rough endoplasmic reticulum. The precursor of the repressible acid phosphatase from Saccharomyces cerevisiae encoded by the PHO5 gene, was synthesized in a yeast lysate programmed with in vitro transcribed PHO5 mRNA. In the presence of yeast rough microsomes up to 16% of the acid phosphatase synthesized was found to be translocated into the microsomes, as judged by proteinase resistance, and fully core-glycosylated. The translocation efficiency however, decreased to 3% if yeast rough microsomes were added after synthesis of acid phosphatase had been terminated. When a wheat-germ extract was used for in vitro synthesis, the precursor of acid phosphatase was translocated into canine pancreatic rough microsomes and thereby core-glycosylated in a signal-recognition-particle-dependent manner. Replacing canine with yeast rough microsomes in the wheat-germ translation system, however, resulted in a significant decrease in the ability to translocate and glycosylate the precursor. Translocation and glycosylation were partially restored by a high-salt extract prepared from yeast ribosomes. The results presented here suggest that yeast-specific factors are needed to translocate and glycosylate acid phosphatase efficiently in vitro.

Hydrogen Gas Attenuates Myocardial Ischemia Reperfusion Injury Independent of Postconditioning in Rats by Attenuating Endoplasmic Reticulum Stress-Induced Autophagy.

PubMed

Gao, Yunan; Yang, Hongxiao; Chi, Jing; Xu, Qiannan; Zhao, Luqi; Yang, Weijia; Liu, Weifan; Yang, Wei

2017-01-01

To study the effect of inhaling hydrogen gas on myocardial ischemic/reperfusion(I/R) injury in rats. Seventy male Wistar albino rats were divided into five groups at random as the sham group (Sham). The I/R group (I/R), The ischemic postconditioning group (IPo), The I/R plus hydrogen group (IH2) and the ischemic postconditioning plus hydrogen group (IPoH2). The Sham group was without coronary occlusion. In I/R group, Ischemic/reperfusion injury was induced by coronary occlusion for 1 hour. Followed by 2 hours of reperfusion. In the IPo and IPoH2 group, four cycles of 1 min reperfusion/1 min ischemia was given at the end of 1 hour coronary occlusion. While 2% hydrogen was administered by inhalation 5 min before reperfusion till 2 hours after reperfusion in both the IPoH2 and IH2 group. The heart and blood samples were harvested at the end of the surgical protocol. Then the myocardium cell endoplasmic reticulum(ER) stress and autophagy was observed by electron microscope. In addition, the cardiac ER stress and autophagy related proteins expression were detected by Western blotting analysis. Both inhaling 2% hydrogen and ischemic postconditioning treatment reduced the ischemic size and serum troponin I level in rats with I/R injury, and inhaling hydrogen showed a more curative effect compared with ischemic postconditioning treatment. Meanwhile inhaling hydrogen showed a better protective effect in attenuating tissue reactive oxygen species. Malondialdehyde levels and immunoreactivities against 8-hydroxy-2'-deoxyguanosine and inhibiting cardiac endoplasmic reticulum stress and down-regulating autophagy as compared with ischemic postconditioning treatment. These results revealed a better protective effect of hydrogen on myocardial ischemic/reperfusion injury in rats by attenuating endoplasmic reticulum stress and down-regulating autophagy compared with ischemic postconditioning treatment. © 2017 The Author(s). Published by S. Karger AG, Basel.

Xc- inhibitor sulfasalazine sensitizes colorectal cancer to cisplatin by a GSH-dependent mechanism.

PubMed

Ma, Ming-zhe; Chen, Gang; Wang, Peng; Lu, Wen-hua; Zhu, Chao-feng; Song, Ming; Yang, Jing; Wen, Shijun; Xu, Rui-hua; Hu, Yumin; Huang, Peng

2015-11-01

Sulfasalazine (SSZ) is an anti-inflammatory drug that has been demonstrated to induce apoptosis and tumor regression through inhibition of plasma membrane cystine transporter xc(-). Cysteine is a rate-limiting precursor for intracellular glutathione (GSH) synthesis, which is vital for compound detoxification and maintaining redox balance. Platinum-based chemotherapy is an important regimen used in clinics for various cancers including colorectal cancer (CRC). We hypothesized that targeting xc(-) transporter by SSZ may annihilate cellular detoxification through interruption of GSH synthesis and may enhance the anti-cancer activity of cisplatin (CDDP) by increasing drug transport. In the present study, we revealed that xCT, the active subunit of xc(-), is highly expressed in CRC cell lines and human colorectal carcinoma tissues compared with their normal counterparts. SSZ effectively depleted cellular GSH, leading to significant accumulation of reactive oxygen species and growth inhibition in CRC cells. In contrast, the normal epithelial cells of colon origin were less sensitive to SSZ, showing a moderate ROS elevation. Importantly, SSZ effectively enhanced the intracellular platinum level and cytotoxicity of CDDP in CRC cells. The synergistic effect of SSZ and CDDP was reversed by antioxidant N-acetyl-L-cysteine (NAC). Together, these results suggest that SSZ, a relatively non-toxic drug that targets cystine transporter, may, in combination with CDDP, have effective therapy for colorectal cancer. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Restoration | NOAA Gulf Spill Restoration

Science.gov Websites

comprehensive look at the types of restoration that may be required to offset impacts from the spill during this long-term injury assessment continues. Types of possible early restoration projects include: Creating

σ-1 Receptor at the Mitochondrial-Associated Endoplasmic Reticulum Membrane Is Responsible for Mitochondrial Metabolic Regulation

PubMed Central

Marriott, Karla-Sue C.; Prasad, Manoj; Thapliyal, Veena

2012-01-01

The mitochondria-associated endoplasmic reticulum (ER) membrane (MAM) is a small section of the outer mitochondrial membrane tethered to the ER by lipid and protein filaments. One such MAM protein is the σ-1 receptor, which contributes to multiple signaling pathways. We found that short interfering RNA-mediated knockdown of σ-1 reduced pregnenolone synthesis by 95% without affecting expression of the inner mitochondrial membrane resident enzyme, 3-β-hydroxysteroid dehydrogenase 2. To explore the underlying mechanism of this effect, we generated a series of σ-receptor ligands: 5,6-dimethoxy-3-methyl-N-phenyl-N-(3-(piperidin-1-yl)propyl)benzofuran-2-carboxamide (KSCM-1), 3-methyl-N-phenyl-N-(3-(piperidin-1-yl)propyl)benzofuran-2-carboxamide (KSCM-5), and 6-methoxy-3-methyl-N-phenyl-N-(3-(piperidin-1-yl) propyl)benzofuran-2-carboxamide (KSCM-11) specifically bound to σ-1 in the nanomolar range, whereas KSCM-5 and KSCM-11 also bound to σ-2. Treatment of cells with the KSCM ligands led to decreased cell viability, with KSCM-5 having the most potent effect followed by KSCM-11. KSCM-1 increased σ-1 expression by 4-fold and progesterone synthesis, whereas the other compounds decreased progesterone synthesis. These differences probably are caused by ligand molecular structure. For example, KSCM-1 has two methoxy substituents at C-5 and C-6 of the benzofuran ring, whereas KSCM-11 has one at C-6. KSCM ligands or σ-1 knockdown did not alter the expression of ER resident enzymes that synthesize steroids. However, coimmunoprecipitation of the σ-1 receptor pulled down voltage-dependent anion channel 2 (VDAC2), whose expression was enhanced by KSCM-1. VDAC2 plays a key role in cholesterol transport into the mitochondria, suggesting that the σ-1 receptor at the MAM coordinates with steroidogenic acute regulatory protein for cholesterol trafficking into the mitochondria for metabolic regulation. PMID:22923735

Sequestosome 1 (SQSTM1/p62) maintains protein folding capacity under endoplasmic reticulum stress in mouse hypothalamic organotypic culture.

PubMed

Tominaga, Takashi; Goto, Motomitsu; Onoue, Takeshi; Mizoguchi, Akira; Sugiyama, Mariko; Tsunekawa, Taku; Hagiwara, Daisuke; Morishita, Yoshiaki; Ito, Yoshihiro; Iwama, Shintaro; Suga, Hidetaka; Banno, Ryoichi; Arima, Hiroshi

2017-08-24

Sequestosome 1 (SQSTM1) also known as ubiquitin-binding protein p62 (p62) is a cargo protein involved in the degradation of misfolded proteins via selective autophagy. Disruption of autophagy and resulting accumulation of misfolded proteins in the endoplasmic reticulum (ER) leads to ER stress. ER stress is implicated in several neurodegenerative diseases and obesity. As knockout of p62 (p62KO) reportedly induces obesity in mice, we examined how p62 contributes to ER stress and the ensuing unfolded protein response (UPR) in hypothalamus using mouse organotypic cultures in the present study. Cultures from p62KO mice showed significantly reduced formation of LC3-GFP puncta, an index of autophagosome formation, in response to the chemical ER stressor thapsigargin compared to wild-type (WT) cultures. Hypothalamic cultures from p62KO mice exhibited higher basal expression of the UPR/ER stress markers CHOP mRNA and ATF4 mRNA than WT cultures. Thapsigargin enhanced CHOP, ATF4, and BiP mRNA as well as p-eIF2α protein expression in both WT and p62KO cultures, but all peak values were greater in p62KO cultures. A proteasome inhibitor increased p62 expression in WT cultures and upregulated the UPR/ER stress markers CHOP mRNA and ATF4 mRNA in both genotypes, but to a greater extent in p62KO cultures. Therefore, p62 deficiency disturbed autophagosome formation and enhanced both basal and chemically induced ER stress, suggesting that p62 serves to prevent ER stress in mouse hypothalamus by maintaining protein folding capacity. Copyright © 2017 Elsevier B.V. All rights reserved.

Guiding Restoration Principles

DTIC Science & Technology

2004-12-01

novaeangliae). Rapid, unexplained declines in the populations of the southern pod of resident Puget Sound orcas (Orcinus orca ), groundfi sh such as...Guiding Restoration Principles 1 Technical Report 2004-03 Guiding Restoration Principles Prepared in support of the Puget Sound ...Kurt Fresh, Tom Mumford and Miles Logsdon Puget Sound Nearshore Partnership WDFW - P.O. Box 43145, Olympia Washington 98504-3145 U.S. Army Corps of

Evaluation of Four Different Restorative Materials for Restoration of the Periodontal Condition of Wedge-Shaped Defect: A Comparative Study.

PubMed

Ruan, Jian-Yong; Gong, Zheng-Lin; Zhang, Rui-Zhi; Zhang, Zhe; Xu, Ran; Li, Da-Xu; Ren, Le; Tao, Hong

2017-09-16

BACKGROUND This study aimed to conduct a clinical evaluation of four restorative materials for restoration of dental wedge-shaped defect (WSD) and their impacts on periodontal tissues. MATERIAL AND METHODS A total of 280 maxillary premolars with dental WSD were selected from 106 patients; the patient cases were divided into eight groups according to different combinations of restorative materials (flowable resin composites, Dyract compomers, glass ionomer cement (GIC), light-curing composite resin), and WSD positions (approaching gingival and subgingival positions). Gingival crevicular fluid (GCF) volume, levels of aspartate aminotransferase (AST), alkaline phosphatase (ALP), and interleukin-1β (IL-1β) in GCF were analyzed, while probing depth (PD), plaque index (PLI), and sulcus bleeding index (SBI) were also measured. The periodontal conditions of all patients were followed prior to restoration, as well as six months and 12 months after restoration. RESULTS After six months of restoration, the overall clinical success rates of flowable resin composites, Dyract compomers, and light-curing composite resin were greater than those of GIC. GCF volume, GCF-AST, IL-1β levels, PD, PLI, and SBI of cases restored by GIC were higher than those restored by the other three materials. After 12 months of restoration, the overall clinical success rates of flowable resin composites and Dyract compomers were greater than those of light-curing composite resin and GIC. GCF volume, GCF-AST, GCF-ALP, IL-1β levels, PD, PLI, and SBI of cases restored by GIC were higher than those restored by the other three materials. CONCLUSIONS Our study provided evidence that the clinical efficacy of flowable resin composites, Dyract compomers, and light-curing composite resin was greater than that of GIC for restoration of dental WSD.

Evaluation of Four Different Restorative Materials for Restoration of the Periodontal Condition of Wedge-Shaped Defect: A Comparative Study

PubMed Central

Ruan, Jian-Yong; Gong, Zheng-Lin; Zhang, Rui-Zhi; Zhang, Zhe; Xu, Ran; Li, Da-Xu; Ren, Le; Tao, Hong

2017-01-01

Background This study aimed to conduct a clinical evaluation of four restorative materials for restoration of dental wedge-shaped defect (WSD) and their impacts on periodontal tissues. Material/Methods A total of 280 maxillary premolars with dental WSD were selected from 106 patients; the patient cases were divided into eight groups according to different combinations of restorative materials (flowable resin composites, Dyract compomers, glass ionomer cement (GIC), light-curing composite resin), and WSD positions (approaching gingival and subgingival positions). Gingival crevicular fluid (GCF) volume, levels of aspartate aminotransferase (AST), alkaline phosphatase (ALP), and interleukin-1β (IL-1β) in GCF were analyzed, while probing depth (PD), plaque index (PLI), and sulcus bleeding index (SBI) were also measured. The periodontal conditions of all patients were followed prior to restoration, as well as six months and 12 months after restoration. Results After six months of restoration, the overall clinical success rates of flowable resin composites, Dyract compomers, and light-curing composite resin were greater than those of GIC. GCF volume, GCF-AST, IL-1β levels, PD, PLI, and SBI of cases restored by GIC were higher than those restored by the other three materials. After 12 months of restoration, the overall clinical success rates of flowable resin composites and Dyract compomers were greater than those of light-curing composite resin and GIC. GCF volume, GCF-AST, GCF-ALP, IL-1β levels, PD, PLI, and SBI of cases restored by GIC were higher than those restored by the other three materials. Conclusions Our study provided evidence that the clinical efficacy of flowable resin composites, Dyract compomers, and light-curing composite resin was greater than that of GIC for restoration of dental WSD. PMID:28917087

Endoplasmic reticulum stress in the pathogenesis of hypertension.

PubMed

Young, Colin N

2017-08-01

What is the topic of this review? This review highlights the emerging role of disruptions in endoplasmic reticulum (ER) function, namely ER stress, as a contributor to hypertension. What advances does it highlight? This review presents an integrative view of ER stress in cardiovascular control systems, including systems within the brain, kidney and peripheral vasculature, as related to development of hypertension. The endoplasmic reticulum (ER) is a cellular organelle specialized in the synthesis, folding, assembly and modification of proteins. In situations of increased protein demand, complex signalling pathways, termed the unfolded protein response, influence a series of cellular feedback loops to control ER function strictly. Although this is initially a compensatory attempt to maintain cellular homeostasis, chronic activation of the unfolded protein response, known as ER stress, leads to sustained changes in cellular function. A growing body of literature points to ER stress in diverse cardioregulatory systems, including the brain, kidney and vasculature, as central to the development of hypertension. Here, these recent findings from essential and obesity-related forms of hypertension are highlighted in an integrative manner, with discussion of the potential upstream causes and downstream consequences of ER stress. Given that hypertension is a leading medical and socio-economic global challenge, emerging findings suggest that targeting ER stress might represent a viable strategy for the treatment of hypertensive disease. © 2017 The Authors. Experimental Physiology © 2017 The Physiological Society.

Allelism analysis of BrRfp locus in different restorer lines and map-based cloning of a fertility restorer gene, BrRfp1, for pol CMS in Chinese cabbage (Brassica rapa L.).

PubMed

Zhang, Huamin; Wu, Junqing; Dai, Zihui; Qin, Meiling; Hao, Lingyu; Ren, Yanjing; Li, Qingfei; Zhang, Lugang

2017-03-01

In Chinese cabbage, there are two Rf loci for pol CMS and one of them was mapped to a 12.6-kb region containing a potential candidate gene encoding PPR protein. In Chinese cabbage (Brassica rapa), polima cytoplasmic male sterility (pol CMS) is an important CMS type and is widely used for hybrid breeding. By extensive test crossing in Chinese cabbage, four restorer lines (92s105, 01s325, 00s109, and 88s148) for pol CMS were screened. By analyzing the allelism of the four restorer lines, it was found that 92s105, 01s325, and 00s109 had the same "restorers of fertility" (Rf) locus (designated as BrRfp1), but 88s148 had a different Rf locus (designated as BrRfp2). For fine mapping the BrRfp1 locus of 92s105, a BC 1 F 1 population with 487 individuals and a BC 1 F 2 population with 2485 individuals were successively constructed. Using simple sequence repeat (SSR) markers developed from Brassica rapa reference genome and InDel markers derived from whole-genome resequencing data of 94c9 and 92s105, BrRfp1 was mapped to a 12.6-kb region containing a potential candidate gene encoding pentatricopeptide repeat-containing protein. Based on the nucleotide polymorphisms of the candidate gene sequence between the restoring and nonrestoring alleles, a co-segregating marker SC718 was developed, which would be helpful for hybrid breeding by marker-assisted screening and for detecting new restorer lines.

Endoplasmic-Reticulum Calcium Depletion and Disease

PubMed Central

Mekahli, Djalila; Bultynck, Geert; Parys, Jan B.; De Smedt, Humbert; Missiaen, Ludwig

2011-01-01

The endoplasmic reticulum (ER) as an intracellular Ca2+ store not only sets up cytosolic Ca2+ signals, but, among other functions, also assembles and folds newly synthesized proteins. Alterations in ER homeostasis, including severe Ca2+ depletion, are an upstream event in the pathophysiology of many diseases. On the one hand, insufficient release of activator Ca2+ may no longer sustain essential cell functions. On the other hand, loss of luminal Ca2+ causes ER stress and activates an unfolded protein response, which, depending on the duration and severity of the stress, can reestablish normal ER function or lead to cell death. We will review these various diseases by mainly focusing on the mechanisms that cause ER Ca2+ depletion. PMID:21441595

Image restoration, uncertainty, and information.

PubMed

Yu, F T

1969-01-01

Some of the physical interpretations about image restoration are discussed. From the theory of information the unrealizability of an inverse filter can be explained by degradation of information, which is due to distortion on the recorded image. The image restoration is a time and space problem, which can be recognized from the theory of relativity (the problem of image restoration is related to Heisenberg's uncertainty principle in quantum mechanics). A detailed discussion of the relationship between information and energy is given. Two general results may be stated: (1) the restoration of the image from the distorted signal is possible only if it satisfies the detectability condition. However, the restored image, at the best, can only approach to the maximum allowable time criterion. (2) The restoration of an image by superimposing the distorted signal (due to smearing) is a physically unrealizable method. However, this restoration procedure may be achieved by the expenditure of an infinite amount of energy.

Chemical chaperones reduce endoplasmic reticulum stress and prevent mutant HFE aggregate formation.

PubMed

de Almeida, Sérgio F; Picarote, Gonçalo; Fleming, John V; Carmo-Fonseca, Maria; Azevedo, Jorge E; de Sousa, Maria

2007-09-21

HFE C282Y, the mutant protein associated with hereditary hemochromatosis (HH), fails to acquire the correct conformation in the endoplasmic reticulum (ER) and is targeted for degradation. We have recently shown that an active unfolded protein response (UPR) is present in the cells of patients with HH. Now, by using HEK 293T cells, we demonstrate that the stability of HFE C282Y is influenced by the UPR signaling pathway that promotes its degradation. Treatment of HFE C282Y-expressing cells with tauroursodeoxycholic acid (TUDCA), a bile acid derivative with chaperone properties, or with the chemical chaperone sodium 4-phenylbutyrate (4PBA) impeded the UPR activation. However, although TUDCA led to an increased stability of the mutant protein, 4PBA contributed to a more efficient disposal of HFE C282Y to the degradation route. Fluorescence microscopy and biochemical analysis of the subcellular localization of HFE revealed that a major portion of the C282Y mutant protein forms intracellular aggregates. Although neither TUDCA nor 4PBA restored the correct folding and intracellular trafficking of HFE C282Y, 4PBA prevented its aggregation. These data suggest that TUDCA hampers the UPR activation by acting directly on its signal transduction pathway, whereas 4PBA suppresses ER stress by chemically enhancing the ER capacity to cope with the expression of misfolded HFE, facilitating its degradation. Together, these data shed light on the molecular mechanisms involved in HFE C282Y-related HH and open new perspectives on the use of orally active chemical chaperones as a therapeutic approach for HH.

Deficiency in the Lipid Exporter ABCA1 Impairs Retrograde Sterol Movement and Disrupts Sterol Sensing at the Endoplasmic Reticulum*♦

PubMed Central

Yamauchi, Yoshio; Iwamoto, Noriyuki; Rogers, Maximillian A.; Abe-Dohmae, Sumiko; Fujimoto, Toyoshi; Chang, Catherine C. Y.; Ishigami, Masato; Kishimoto, Takuma; Kobayashi, Toshihide; Ueda, Kazumitsu; Furukawa, Koichi; Chang, Ta-Yuan; Yokoyama, Shinji

2015-01-01

Cellular cholesterol homeostasis involves sterol sensing at the endoplasmic reticulum (ER) and sterol export from the plasma membrane (PM). Sterol sensing at the ER requires efficient sterol delivery from the PM; however, the macromolecules that facilitate retrograde sterol transport at the PM have not been identified. ATP-binding cassette transporter A1 (ABCA1) mediates cholesterol and phospholipid export to apolipoprotein A-I for the assembly of high density lipoprotein (HDL). Mutations in ABCA1 cause Tangier disease, a familial HDL deficiency. Several lines of clinical and experimental evidence suggest a second function of ABCA1 in cellular cholesterol homeostasis in addition to mediating cholesterol efflux. Here, we report the unexpected finding that ABCA1 also plays a key role in facilitating retrograde sterol transport from the PM to the ER for sterol sensing. Deficiency in ABCA1 delays sterol esterification at the ER and activates the SREBP-2 cleavage pathway. The intrinsic ATPase activity in ABCA1 is required to facilitate retrograde sterol transport. ABCA1 deficiency causes alternation of PM composition and hampers a clathrin-independent endocytic activity that is required for ER sterol sensing. Our finding identifies ABCA1 as a key macromolecule facilitating bidirectional sterol movement at the PM and shows that ABCA1 controls retrograde sterol transport by modulating a certain clathrin-independent endocytic process. PMID:26198636

Brucella Rough Mutant Induce Macrophage Death via Activating IRE1α Pathway of Endoplasmic Reticulum Stress by Enhanced T4SS Secretion

PubMed Central

Li, Peng; Tian, Mingxing; Bao, Yanqing; Hu, Hai; Liu, Jiameng; Yin, Yi; Ding, Chan; Wang, Shaohui; Yu, Shengqing

2017-01-01

Brucella is a Gram-negative facultative intracellular pathogen that causes the worldwide zoonosis, known as brucellosis. Brucella virulence relies mostly on its ability to invade and replicate within phagocytic cells. The type IV secretion system (T4SS) and lipopolysaccharide are two major Brucella virulence factors. Brucella rough mutants reportedly induce the death of infected macrophages, which is T4SS dependent. However, the underlying molecular mechanism remains unclear. In this study, the T4SS secretion capacities of Brucella rough mutant and its smooth wild-type strain were comparatively investigated, by constructing the firefly luciferase fused T4SS effector, BPE123 and VceC. In addition, quantitative real-time PCR and western blotting were used to analyze the T4SS expression. The results showed that T4SS expression and secretion were enhanced significantly in the Brucella rough mutant. We also found that the activity of the T4SS virB operon promoter was notably increased in the Brucella rough mutant, which depends on quorum sensing-related regulators of VjbR upregulation. Cell infection and cell death assays revealed that deletion of vjbR in the Brucella rough mutant absolutely abolished cytotoxicity within macrophages by downregulating T4SS expression. This suggests that up-regulation of T4SS promoted by VjbR in rough mutant ΔrfbE contribute to macrophage death. In addition, we found that the Brucella rough mutant induce macrophage death via activating IRE1α pathway of endoplasmic reticulum stress. Taken together, our study provide evidence that in comparison to the Brucella smooth wild-type strain, VjbR upregulation in the Brucella rough mutant increases transcription of the virB operon, resulting in overexpression of the T4SS gene, accompanied by the over-secretion of effecter proteins, thereby causing the death of infected macrophages via activating IRE1α pathway of endoplasmic reticulum stress, suggesting novel insights into the molecular

Brucella Rough Mutant Induce Macrophage Death via Activating IRE1α Pathway of Endoplasmic Reticulum Stress by Enhanced T4SS Secretion.

PubMed

Li, Peng; Tian, Mingxing; Bao, Yanqing; Hu, Hai; Liu, Jiameng; Yin, Yi; Ding, Chan; Wang, Shaohui; Yu, Shengqing

2017-01-01

Brucella is a Gram-negative facultative intracellular pathogen that causes the worldwide zoonosis, known as brucellosis. Brucella virulence relies mostly on its ability to invade and replicate within phagocytic cells. The type IV secretion system (T4SS) and lipopolysaccharide are two major Brucella virulence factors. Brucella rough mutants reportedly induce the death of infected macrophages, which is T4SS dependent. However, the underlying molecular mechanism remains unclear. In this study, the T4SS secretion capacities of Brucella rough mutant and its smooth wild-type strain were comparatively investigated, by constructing the firefly luciferase fused T4SS effector, BPE123 and VceC. In addition, quantitative real-time PCR and western blotting were used to analyze the T4SS expression. The results showed that T4SS expression and secretion were enhanced significantly in the Brucella rough mutant. We also found that the activity of the T4SS virB operon promoter was notably increased in the Brucella rough mutant, which depends on quorum sensing-related regulators of VjbR upregulation. Cell infection and cell death assays revealed that deletion of vjbR in the Brucella rough mutant absolutely abolished cytotoxicity within macrophages by downregulating T4SS expression. This suggests that up-regulation of T4SS promoted by VjbR in rough mutant Δ rfbE contribute to macrophage death. In addition, we found that the Brucella rough mutant induce macrophage death via activating IRE1α pathway of endoplasmic reticulum stress. Taken together, our study provide evidence that in comparison to the Brucella smooth wild-type strain, VjbR upregulation in the Brucella rough mutant increases transcription of the virB operon, resulting in overexpression of the T4SS gene, accompanied by the over-secretion of effecter proteins, thereby causing the death of infected macrophages via activating IRE1α pathway of endoplasmic reticulum stress, suggesting novel insights into the molecular

Bearing restoration by grinding

NASA Technical Reports Server (NTRS)

Hanau, H.; Parker, R. J.; Zaretsky, E. V.; Chen, S. M.; Bull, H. L.

1976-01-01

A joint program was undertaken by the NASA Lewis Research Center and the Army Aviation Systems Command to restore by grinding those rolling-element bearings which are currently being discarded at aircraft engine and transmission overhaul. Three bearing types were selected from the UH-1 helicopter engine (T-53) and transmission for the pilot program. No bearing failures occurred related to the restoration by grinding process. The risk and cost of a bearing restoration by grinding programs was analyzed. A microeconomic impact analysis was performed.

Endoplasmic reticulum and lysosomal Ca²⁺ stores are remodelled in GBA1-linked Parkinson disease patient fibroblasts.

PubMed

Kilpatrick, Bethan S; Magalhaes, Joana; Beavan, Michelle S; McNeill, Alisdair; Gegg, Matthew E; Cleeter, Michael W J; Bloor-Young, Duncan; Churchill, Grant C; Duchen, Michael R; Schapira, Anthony H; Patel, Sandip

2016-01-01

Mutations in β-glucocerebrosidase (encoded by GBA1) cause Gaucher disease (GD), a lysosomal storage disorder, and increase the risk of developing Parkinson disease (PD). The pathogenetic relationship between the two disorders is unclear. Here, we characterised Ca(2+) release in fibroblasts from type I GD and PD patients together with age-matched, asymptomatic carriers, all with the common N370S mutation in β-glucocerebrosidase. We show that endoplasmic reticulum (ER) Ca(2+) release was potentiated in GD and PD patient fibroblasts but not in cells from asymptomatic carriers. ER Ca(2+) signalling was also potentiated in fibroblasts from aged healthy subjects relative to younger individuals but not further increased in aged PD patient cells. Chemical or molecular inhibition of β-glucocerebrosidase in fibroblasts and a neuronal cell line did not affect ER Ca(2+) signalling suggesting defects are independent of enzymatic activity loss. Conversely, lysosomal Ca(2+) store content was reduced in PD fibroblasts and associated with age-dependent alterations in lysosomal morphology. Accelerated remodelling of Ca(2+) stores by pathogenic GBA1 mutations may therefore feature in PD. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

Provision of Atraumatic Restorative Treatment (ART) restorations to Chinese pre-school children--a 30-month evaluation.

PubMed

Lo, E C; Holmgren, C J

2001-01-01

The objectives of this study were: to provide restorations using the ART approach to pre-school children in Southern China in a kindergarten environment, using a high-strength glass-ionomer restorative material; to assess the acceptability of this approach and to evaluate on a longitudinal basis the restorations placed. A total of 170 ART restorations were placed in 95 children, aged 5.1 +/- 0.7 years, by seven final-year dental students using standard ART procedures and hand instruments. The restorations were evaluated every six months thereafter by two calibrated independent examiners using explorers and mouth-mirrors. 93% of the children reported that they did not feel pain during treatment and 86% were willing to receive ART restorations again. The cumulative 12- and 30-month survival rates of Class I restorations were 91% and 79%, respectively. The corresponding figures for Class V restorations were 79% and 70%, while those for Class II restorations were 75% and 51%. The failure rates of Class III and IV restorations were high with more than half of them scored as missing within the first year. The ART approach was shown to be acceptable to Chinese pre-school children for providing restorative dental care outside the traditional clinical setting. The success rates were high for Class I and V restorations in primary teeth, modest for Class II, and low for Class III and IV restorations.

5 CFR 353.301 - Restoration rights.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 5 Administrative Personnel 1 2010-01-01 2010-01-01 false Restoration rights. 353.301 Section 353... DUTY FROM UNIFORMED SERVICE OR COMPENSABLE INJURY Compensable Injury § 353.301 Restoration rights. (a.... Although these restoration rights are agencywide, the employee's basic entitlement is to the former...

Hydromorphological restoration stimulates river ecosystem metabolism

NASA Astrophysics Data System (ADS)

Kupilas, Benjamin; Hering, Daniel; Lorenz, Armin W.; Knuth, Christoph; Gücker, Björn

2017-04-01

Both ecosystem structure and functioning determine ecosystem status and are important for the provision of goods and services to society. However, there is a paucity of research that couples functional measures with assessments of ecosystem structure. In mid-sized and large rivers, effects of restoration on key ecosystem processes, such as ecosystem metabolism, have rarely been addressed and remain poorly understood. We compared three reaches of the third-order, gravel-bed river Ruhr in Germany: two reaches restored with moderate (R1) and substantial effort (R2) and one upstream degraded reach (D). Hydromorphology, habitat composition, and hydrodynamics were assessed. We estimated gross primary production (GPP) and ecosystem respiration (ER) using the one-station open-channel diel dissolved oxygen change method over a 50-day period at the end of each reach. Moreover, we estimated metabolic rates of the combined restored reaches (R1 + R2) using the two-station open-channel method. Values for hydromorphological variables increased with restoration intensity (D < R1 < R2). Restored reaches had lower current velocity, higher longitudinal dispersion and larger transient storage zones. However, fractions of median travel time due to transient storage were highest in R1 and lowest in R2, with intermediate values in D. The share of macrophyte cover of total wetted area was highest in R2 and lowest in R1, with intermediate values in D. Station R2 had higher average GPP and ER than R1 and D. The combined restored reaches R1 + R2 also exhibited higher GPP and ER than the degraded upstream river (station D). Restoration increased river autotrophy, as indicated by elevated GPP : ER, and net ecosystem production (NEP) of restored reaches. Temporal patterns of ER closely mirrored those of GPP, pointing to the importance of autochthonous production for ecosystem functioning. In conclusion, high reach-scale restoration effort had considerable effects on river hydrodynamics and

Lentiviral Expression of Retinal Guanylate Cyclase-1 (RetGC1) Restores Vision in an Avian Model of Childhood Blindness

PubMed Central

Haire, Shannon E; Aleman, Tomas S; Cideciyan, Artur V; Sokal, Izabel; Palczewski, Krzysztof; Jacobson, Samuel G; Semple-Rowland, Susan L

2006-01-01

Background Leber congenital amaurosis (LCA) is a genetically heterogeneous group of retinal diseases that cause congenital blindness in infants and children. Mutations in the GUCY2D gene that encodes retinal guanylate cyclase–1 (retGC1) were the first to be linked to this disease group (LCA type 1 [LCA1]) and account for 10%–20% of LCA cases. These mutations disrupt synthesis of cGMP in photoreceptor cells, a key second messenger required for function of these cells. The GUCY1*B chicken, which carries a null mutation in the retGC1 gene, is blind at hatching and serves as an animal model for the study of LCA1 pathology and potential treatments in humans. Methods and Findings A lentivirus-based gene transfer vector carrying the GUCY2D gene was developed and injected into early-stage GUCY1*B embryos to determine if photoreceptor function and sight could be restored to these animals. Like human LCA1, the avian disease shows early-onset blindness, but there is a window of opportunity for intervention. In both diseases there is a period of photoreceptor cell dysfunction that precedes retinal degeneration. Of seven treated animals, six exhibited sight as evidenced by robust optokinetic and volitional visual behaviors. Electroretinographic responses, absent in untreated animals, were partially restored in treated animals. Morphological analyses indicated there was slowing of the retinal degeneration. Conclusions Blindness associated with loss of function of retGC1 in the GUCY1*B avian model of LCA1 can be reversed using viral vector-mediated gene transfer. Furthermore, this reversal can be achieved by restoring function to a relatively low percentage of retinal photoreceptors. These results represent a first step toward development of gene therapies for one of the more common forms of childhood blindness. PMID:16700630

How is success or failure in river restoration projects evaluated? Feedback from French restoration projects.

PubMed

Morandi, Bertrand; Piégay, Hervé; Lamouroux, Nicolas; Vaudor, Lise

2014-05-01

Since the 1990s, French operational managers and scientists have been involved in the environmental restoration of rivers. The European Water Framework Directive (2000) highlights the need for feedback from restoration projects and for evidence-based evaluation of success. Based on 44 French pilot projects that included such an evaluation, the present study includes: 1) an introduction to restoration projects based on their general characteristics 2) a description of evaluation strategies and authorities in charge of their implementation, and 3) a focus on the evaluation of results and the links between these results and evaluation strategies. The results show that: 1) the quality of an evaluation strategy often remains too poor to understand well the link between a restoration project and ecological changes; 2) in many cases, the conclusions drawn are contradictory, making it difficult to determine the success or failure of a restoration project; and 3) the projects with the poorest evaluation strategies generally have the most positive conclusions about the effects of restoration. Recommendations are that evaluation strategies should be designed early in the project planning process and be based on clearly-defined objectives. Copyright © 2014 Elsevier Ltd. All rights reserved.

Endoplasmic reticulum stress is induced in the human placenta during labour

PubMed Central

Veerbeek, J.H.W.; Tissot Van Patot, M.C.; Burton, G.J.; Yung, H.W.

2015-01-01

Placental endoplasmic reticulum (ER) stress has been postulated in the pathophysiology of pre-eclampsia (PE) and intrauterine growth restriction (IUGR), but its activation remains elusive. Oxidative stress induced by ischaemia/hypoxia-reoxygenation activates ER stress in vitro. Here, we explored whether exposure to labour represents an in vivo model for the study of acute placental ER stress. ER stress markers, GRP78, P-eIF2α and XBP-1, were significantly higher in laboured placentas than in Caesarean-delivered controls localised mainly in the syncytiotrophoblast. The similarities to changes observed in PE/IUGR placentas suggest exposure to labour can be used to investigate induction of ER stress in pathological placentas. PMID:25434970

Ceramics in Restorative and Prosthetic DENTISTRY1

NASA Astrophysics Data System (ADS)

Kelly, J. Robert

1997-08-01

This review is intended to provide the ceramic engineer with information about the history and current use of ceramics in dentistry, contemporary research topics, and potential research agenda. Background material includes intra-oral design considerations, descriptions of ceramic dental components, and the origin, composition, and microstructure of current dental ceramics. Attention is paid to efforts involving net-shape processing, machining as a forming method, and the analysis of clinical failure. A rationale is presented for the further development of all-ceramic restorative systems. Current research topics receiving attention include microstructure/processing/property relationships, clinical failure mechanisms and in vitro testing, wear damage and wear testing, surface treatments, and microstructural modifications. The status of the field is critically reviewed with an eye toward future work. Significant improvements seem possible in the clinical use of ceramics based on engineering solutions derived from the study of clinically failed restorations, on the incorporation of higher levels of "biomimicry" in new systems, and on the synergistic developments in dental cements and adhesive dentin bonding.

15 CFR 990.55 - Restoration selection-developing restoration plans.

Code of Federal Regulations, 2011 CFR

2011-01-01

... 15 Commerce and Foreign Trade 3 2011-01-01 2011-01-01 false Restoration selection-developing... POLLUTION ACT REGULATIONS NATURAL RESOURCE DAMAGE ASSESSMENTS Restoration Planning Phase § 990.55 Restoration selection—developing restoration plans. (a) General. OPA requires that damages be based upon a...

15 CFR 990.55 - Restoration selection-developing restoration plans.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 15 Commerce and Foreign Trade 3 2010-01-01 2010-01-01 false Restoration selection-developing... POLLUTION ACT REGULATIONS NATURAL RESOURCE DAMAGE ASSESSMENTS Restoration Planning Phase § 990.55 Restoration selection—developing restoration plans. (a) General. OPA requires that damages be based upon a...

Forest restoration paradigms

Treesearch

John Stanturf; Brian J. Palik; Mary I. Williams; R. Kasten Dumroese

2014-01-01

An estimated 2 billion ha of forests are degraded globally and global change suggests even greater need for forest restoration. Four forest restoration paradigms are identified and discussed: revegetation, ecological restoration, functional restoration, and forest landscape restoration. Restoration is examined in terms of a degraded starting point and an ending point...

Unraveling the function of Arabidopsis thaliana OS9 in the endoplasmic reticulum-associated degradation of glycoproteins.

PubMed

Hüttner, Silvia; Veit, Christiane; Schoberer, Jennifer; Grass, Josephine; Strasser, Richard

2012-05-01

In the endoplasmic reticulum, immature polypeptides coincide with terminally misfolded proteins. Consequently, cells need a well-balanced quality control system, which decides about the fate of individual proteins and maintains protein homeostasis. Misfolded and unassembled proteins are sent for destruction via the endoplasmic reticulum-associated degradation (ERAD) machinery to prevent the accumulation of potentially toxic protein aggregates. Here, we report the identification of Arabidopsis thaliana OS9 as a component of the plant ERAD pathway. OS9 is an ER-resident glycoprotein containing a mannose-6-phosphate receptor homology domain, which is also found in yeast and mammalian lectins involved in ERAD. OS9 fused to the C-terminal domain of YOS9 can complement the ERAD defect of the corresponding yeast Δyos9 mutant. An A. thaliana OS9 loss-of-function line suppresses the severe growth phenotype of the bri1-5 and bri1-9 mutant plants, which harbour mutated forms of the brassinosteroid receptor BRI1. Co-immunoprecipitation studies demonstrated that OS9 associates with Arabidopsis SEL1L/HRD3, which is part of the plant ERAD complex and with the ERAD substrates BRI1-5 and BRI1-9, but only the binding to BRI1-5 occurs in a glycan-dependent way. OS9-deficiency results in activation of the unfolded protein response and reduces salt tolerance, highlighting the role of OS9 during ER stress. We propose that OS9 is a component of the plant ERAD machinery and may act specifically in the glycoprotein degradation pathway.

Alpha-lipoic acid attenuates endoplasmic reticulum stress-induced insulin resistance by improving mitochondrial function in HepG2 cells.

PubMed

Lei, Lin; Zhu, Yiwei; Gao, Wenwen; Du, Xiliang; Zhang, Min; Peng, Zhicheng; Fu, Shoupeng; Li, Xiaobing; Zhe, Wang; Li, Xinwei; Liu, Guowen

2016-10-01

Alpha-lipoic acid (ALA) has been reported to have beneficial effects for improving insulin sensitivity. However, the underlying molecular mechanism of the beneficial effects remains poorly understood. Endoplasmic reticulum (ER) stress and mitochondrial dysfunction are considered causal factors that induce insulin resistance. In this study, we investigated the effect of ALA on the modulation of insulin resistance in ER-stressed HepG2 cells, and we explored the potential mechanism of this effect. HepG2 cells were incubated with tunicamycin (Tun) for 6h to establish an ER stress cell model. Tun treatment induced ER stress, mitochondrial dysfunction and insulin resistance. Interestingly, ALA had no significant effect on ER stress signals. Pretreatment of the ER stress cell model with ALA for 24h improved insulin sensitivity, restored the expression levels of mitochondrial oxidative phosphorylation (OXPHOS) complexes and increased intracellular ATP production. Moreover, ALA augmented the β-oxidation capacity of the mitochondria. Importantly, ALA treatment could decrease oligomycin-induced mitochondrial dysfunction and then improved insulin resistance. Taken together, our data suggest that ALA prevents ER stress-induced insulin resistance by enhancing mitochondrial function. Copyright © 2016 Elsevier Inc. All rights reserved.

Endoplasmic reticulum polymers impair luminal protein mobility and sensitise to cellular stress in α1-antitrypsin deficiency

PubMed Central

Ordóñez, Adriana; Snapp, Erik L; Tan, Lu; Miranda, Elena; Marciniak, Stefan J; Lomas, David A

2013-01-01

Point mutants of α1-antitrypsin form ordered polymers that are retained as inclusions within the endoplasmic reticulum (ER) of hepatocytes in association with neonatal hepatitis, cirrhosis and hepatocellular carcinoma. These inclusions cause cell damage and predispose to ER stress in the absence of the classical unfolded protein response (UPR). The pathophysiology underlying this ER stress was explored by generating cell models that conditionally express wildtype α1-antitrypsin, two mutants that cause polymer-mediated inclusions and liver disease (E342K [the Z allele] and H334D) and a truncated mutant (Null Hong Kong, NHK) that induces classical ER stress and is removed by ER associated degradation. Expression of the polymeric mutants resulted in gross changes in the ER luminal environment that recapitulated the changes seen in liver sections from individuals with PI*ZZ α1-antitrypsin deficiency. In contrast expression of NHK α1-antitrypsin caused electron lucent dilatation and expansion of the ER throughout the cell. Photobleaching microscopy in live cells demonstrated a decrease in the mobility of soluble luminal proteins in cells that express E342K and H334D α1-antitrypsin when compared to those that express wildtype and NHK α1-antitrypsin (0.34±0.05, 0.22±0.03, 2.83±0.30 and 2.84±0.55 μm2/s respectively). There was no effect on protein mobility within ER membranes indicating that cisternal connectivity was not disrupted. Polymer expression alone was insufficient to induce the UPR but the resulting protein overload rendered cells hypersensitive to ER stress induced by either tunicamycin or glucose depletion. Conclusion Changes in protein diffusion provide an explanation for the cellular consequences of ER protein overload in mutants that cause inclusion body formation and α1-antitrypsin deficiency. PMID:23197448

Hydrogen Sulfide Inhibits Formaldehyde-Induced Endoplasmic Reticulum Stress in PC12 Cells by Upregulation of SIRT-1

PubMed Central

Zhang, Ping; Chen, Li-Xun; Wang, Li; Xie, Ming; Wang, Chun-Yan; Tang, Xiao-Qing

2014-01-01

Background Formaldehyde (FA), a well-known environmental pollutant, has been classified as a neurotoxic molecule. Our recent data demonstrate that hydrogen sulfide (H2S), the third gaseous transmitter, has a protective effect on the neurotoxicity of FA. However, the exact mechanisms underlying this protection remain largely unknown. Endoplasmic reticulum (ER) stress has been implicated in the neurotoxicity of FA. Silent mating type information regulator 2 homolog 1 (SIRT-1), a histone deacetylases, has various biological activities, including the extension of lifespan, the modulation of ER stress, and the neuroprotective action. Objective We hypothesize that the protection of H2S against FA-induced neurotoxicity involves in inhibiting ER stress by upregulation of SIRT-1. The present study attempted to investigate the protective effect of H2S on FA-induced ER stress in PC12 cells and the contribution of SIRT-1 to the protection of H2S against FA-induced injuries, including ER stress, cytotoxicity and apoptosis. Principal Findings We found that exogenous application of sodium hydrosulfide (NaHS; an H2S donor) significantly attenuated FA-induced ER stress responses, including the upregulated levels of glucose-regulated protein 78, C/EBP homologous protein, and cleaved caspase-12 expression. We showed that NaHS upregulates the expression of SIRT-1 in PC12 cells. Moreover, the protective effects of H2S on FA-elicited ER stress, cytotoxicity and apoptosis were reversed by Sirtinol, a specific inhibitor of SIRT-1. Conclusion/Significance These data indicate that H2S exerts its protection against the neurotoxicity of FA through overcoming ER stress via upregulation of SIRT-1. Our findings provide novel insights into the protective mechanisms of H2S against FA-induced neurotoxicity. PMID:24587076

Melatonin reduces endoplasmic reticulum stress and corneal dystrophy-associated TGFBIp through activation of endoplasmic reticulum-associated protein degradation.

PubMed

Choi, Seung-Il; Lee, Eunhee; Akuzum, Begum; Jeong, Jang Bin; Maeng, Yong-Sun; Kim, Tae-Im; Kim, Eung Kweon

2017-10-01

Endoplasmic reticulum (ER) stress is emerging as a factor for the pathogenesis of granular corneal dystrophy type 2 (GCD2). This study was designed to investigate the molecular mechanisms underlying the protective effects of melatonin on ER stress in GCD2. Our results showed that GCD2 corneal fibroblasts were more susceptible to ER stress-induced death than were wild-type cells. Melatonin significantly inhibited GCD2 corneal cell death, caspase-3 activation, and poly (ADP-ribose) polymerase 1 cleavage caused by the ER stress inducer, tunicamycin. Under ER stress, melatonin significantly suppressed the induction of immunoglobulin heavy-chain-binding protein (BiP) and activation of inositol-requiring enzyme 1α (IRE1α), and their downstream target, alternative splicing of X-box binding protein 1(XBP1). Notably, the reduction in BiP and IRE1α by melatonin was suppressed by the ubiquitin-proteasome inhibitor, MG132, but not by the autophagy inhibitor, bafilomycin A1, indicating involvement of the ER-associated protein degradation (ERAD) system. Melatonin treatment reduced the levels of transforming growth factor-β-induced protein (TGFBIp) significantly, and this reduction was suppressed by MG132. We also found reduced mRNA expression of the ERAD system components HRD1 and SEL1L, and a reduced level of SEL1L protein in GCD2 cells. Interestingly, melatonin treatments enhanced SEL1L levels and suppressed the inhibition of SEL1L N-glycosylation caused by tunicamycin. In conclusion, this study provides new insights into the mechanisms by which melatonin confers its protective actions during ER stress. The results also indicate that melatonin might have potential as a therapeutic agent for ER stress-related diseases including GCD2. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Role of HERP and a HERP-related protein in HRD1-dependent protein degradation at the endoplasmic reticulum.

PubMed

Huang, Chih-Hsiang; Chu, Yue-Ru; Ye, Yihong; Chen, Xin

2014-02-14

Misfolded proteins of the endoplasmic reticulum (ER) are retrotranslocated to the cytosol and degraded by the proteasome via a process termed ER-associated degradation (ERAD). The precise mechanism of retrotranslocation is unclear. Here, we use several lumenal ERAD substrates targeted for degradation by the ubiquitin ligase HRD1 including SHH (sonic hedgehog) and NHK (null Hong Kong α1-antitrypsin) to study the geometry, organization, and regulation of the HRD1-containing ERAD machinery. We report a new HRD1-associated membrane protein named HERP2, which is homologous to the previously identified HRD1 partner HERP1. Despite sequence homology, HERP2 is constitutively expressed in cells, whereas HERP1 is highly induced by ER stress. We find that these proteins are required for efficient degradation of both glycosylated and nonglycosylated SHH proteins as well as NHK. In cells depleted of HERPs, SHH proteins are largely trapped inside the ER with a fraction of the stabilized SHH protein bound to the HRD1-SEL1L ligase complex. Ubiquitination of SHH is significantly attenuated in the absence of HERPs, suggesting a defect in retrotranslocation. Both HERP proteins interact with HRD1 through a region located in the cytosol. However, unlike its homolog in Saccharomyces cerevisiae, HERPs do not regulate HRD1 stability or oligomerization status. Instead, they help recruit DERL2 to the HRD1-SEL1L complex. Additionally, the UBL domain of HERP1 also seems to have a function independent of DERL2 recruitment in ERAD. Our studies have revealed a critical scaffolding function for mammalian HERP proteins that is required for forming an active retrotranslocation complex containing HRD1, SEL1L, and DERL2.

Cannabinoid Receptor Type 1 Agonist ACEA Protects Neurons from Death and Attenuates Endoplasmic Reticulum Stress-Related Apoptotic Pathway Signaling.

PubMed

Vrechi, Talita A; Crunfli, Fernanda; Costa, Andressa P; Torrão, Andréa S

2018-05-01

Neurodegeneration is the result of progressive destruction of neurons in the central nervous system, with unknown causes and pathological mechanisms not yet fully elucidated. Several factors contribute to neurodegenerative processes, including neuroinflammation, accumulation of neurotoxic factors, and misfolded proteins in the lumen of the endoplasmic reticulum (ER). Endocannabinoid signaling has been pointed out as an important modulatory system in several neurodegeneration-related processes, inhibiting the inflammatory response and increasing neuronal survival. Thus, we investigated the presumptive protective effect of the selective cannabinoid type 1 (CB1) receptor agonist arachidonyl-2'-chloroethylamide (ACEA) against inflammatory (lipopolysaccharide, LPS) and ER stress (tunicamycin) stimuli in an in vitro neuronal model (Neuro-2a neuroblastoma cells). Cell viability analysis revealed that ACEA was able to protect against cell death induced by LPS and tunicamycin. This neuroprotective effect occurs via the CB1 receptor in the inflammation process and via the transient receptor potential of vanilloid type-1 (TRPV1) channel in ER stress. Furthermore, the immunoblotting analyses indicated that the neuroprotective effect of ACEA seems to involve the modulation of eukaryotic initiation factor 2 (eIF2α), transcription factor C/EBP homologous protein (CHOP), and caspase 12, as well as the survival/death p44/42 MAPK, ERK1/2-related signaling pathways. Together, these data suggest that the endocannabinoid system is a potential therapeutic target in neurodegenerative processes, especially in ER-related neurodegenerative diseases.

The Endoplasmic Reticulum Is a Reservoir for WAVE/SCAR Regulatory Complex Signaling in the Arabidopsis Leaf1[W][OA

PubMed Central

Zhang, Chunhua; Mallery, Eileen; Reagan, Sara; Boyko, Vitaly P.; Kotchoni, Simeon O.; Szymanski, Daniel B.

2013-01-01

During plant cell morphogenesis, signal transduction and cytoskeletal dynamics interact to locally organize the cytoplasm and define the geometry of cell expansion. The WAVE/SCAR (for WASP family verprolin homologous/suppressor of cyclic AMP receptor) regulatory complex (W/SRC) is an evolutionarily conserved heteromeric protein complex. Within the plant kingdom W/SRC is a broadly used effector that converts Rho-of-Plants (ROP)/Rac small GTPase signals into Actin-Related Protein2/3 and actin-dependent growth responses. Although the components and biochemistry of the W/SRC pathway are well understood, a basic understanding of how cells partition W/SRC into active and inactive pools is lacking. In this paper, we report that the endoplasmic reticulum (ER) is an important organelle for W/SRC regulation. We determined that a large intracellular pool of the core W/SRC subunit NAP1, like the known positive regulator of W/SRC, the DOCK family guanine nucleotide-exchange factor SPIKE1 (SPK1), localizes to the surface of the ER. The ER-associated NAP1 is inactive because it displays little colocalization with the actin network, and ER localization requires neither activating signals from SPK1 nor a physical association with its W/SRC-binding partner, SRA1. Our results indicate that in Arabidopsis (Arabidopsis thaliana) leaf pavement cells and trichomes, the ER is a reservoir for W/SRC signaling and may have a key role in the early steps of W/SRC assembly and/or activation. PMID:23613272

C-terminal, endoplasmic reticulum-lumenal domain of prosurfactant protein C - structural features and membrane interactions.

PubMed

Casals, Cristina; Johansson, Hanna; Saenz, Alejandra; Gustafsson, Magnus; Alfonso, Carlos; Nordling, Kerstin; Johansson, Jan

2008-02-01

Surfactant protein C (SP-C) constitutes the transmembrane part of prosurfactant protein C (proSP-C) and is alpha-helical in its native state. The C-terminal part of proSP-C (CTC) is localized in the endoplasmic reticulum lumen and binds to misfolded (beta-strand) SP-C, thereby preventing its aggregation and amyloid fibril formation. In this study, we investigated the structure of recombinant human CTC and the effects of CTC-membrane interaction on protein structure. CTC forms noncovalent trimers and supratrimeric oligomers. It contains two intrachain disulfide bridges, and its secondary structure is significantly affected by urea or heat only after disulfide reduction. The postulated Brichos domain of CTC, with homologs found in proteins associated with amyloid and proliferative disease, is up to 1000-fold more protected from limited proteolysis than the rest of CTC. The protein exposes hydrophobic surfaces, as determined by CTC binding to the environment-sensitive fluorescent probe 1,1'-bis(4-anilino-5,5'-naphthalenesulfonate). Fluorescence energy transfer experiments further reveal close proximity between bound 1,1'-bis(4-anilino-5,5'-naphthalenesulfonate) and tyrosine residues in CTC, some of which are conserved in all Brichos domains. CTC binds to unilamellar phospholipid vesicles with low micromolar dissociation constants, and differential scanning calorimetry and CD analyses indicate that membrane-bound CTC is less structurally ordered than the unbound protein. The exposed hydrophobic surfaces and the structural disordering that result from interactions with phospholipid membranes suggest a mechanism whereby CTC binds to misfolded SP-C in the endoplasmic reticulum membrane.

Phase III Early Restoration Public Meetings | NOAA Gulf Spill Restoration

Science.gov Websites

Archive Home Phase III Early Restoration Public Meetings Phase III Early Restoration Public Meetings share Posted on December 6, 2013 | Assessment and Early Restoration Restoration Area Title: Phase III Early on the draft plan for the third phase of Early Restoration, which proposes more than $625 million in

Contribution of Chromosomes 1HchS and 6HchS to Fertility Restoration in the Wheat msH1 CMS System under Different Environmental Conditions

PubMed Central

Castillo, Almudena; Rodríguez-Suárez, Cristina; Martín, Azahara C.; Pistón, Fernando

2015-01-01

Exploiting hybrid wheat heterosis has been long pursued to increase crop yield, stability and uniformity. Cytoplasmic male sterility (CMS) systems based in the nuclear-cytoplasmic incompatible interactions are a classic way for hybrid seed production, but to date, no definitive system is available in wheat. The msH1 CMS system results from the incompatibility between the nuclear genome of wheat and the cytoplasmic genome of the wild barley Hordeum chilense. Fertility restoration of the CMS phenotype was first associated with the disomic addition of the short arm of chromosome 6H from H. chilense. In further studies it was observed that chromosome arm 1HchS was also implicated, and the combination of genes in both chromosome arms restored fertility more efficiently. In this work we aim to dissect the effect of each chromosome in fertility restoration when combined in different genomic backgrounds and under different environmental conditions. We propose a model to explain how restoration behaves in the msH1 system and generate valuable information necessary to develop an efficient system for hybrid wheat production. PMID:26192191

Contribution of Chromosomes 1HchS and 6HchS to Fertility Restoration in the Wheat msH1 CMS System under Different Environmental Conditions.

PubMed

Castillo, Almudena; Rodríguez-Suárez, Cristina; Martín, Azahara C; Pistón, Fernando

2015-01-01

Exploiting hybrid wheat heterosis has been long pursued to increase crop yield, stability and uniformity. Cytoplasmic male sterility (CMS) systems based in the nuclear-cytoplasmic incompatible interactions are a classic way for hybrid seed production, but to date, no definitive system is available in wheat. The msH1 CMS system results from the incompatibility between the nuclear genome of wheat and the cytoplasmic genome of the wild barley Hordeum chilense. Fertility restoration of the CMS phenotype was first associated with the disomic addition of the short arm of chromosome 6H from H. chilense. In further studies it was observed that chromosome arm 1HchS was also implicated, and the combination of genes in both chromosome arms restored fertility more efficiently. In this work we aim to dissect the effect of each chromosome in fertility restoration when combined in different genomic backgrounds and under different environmental conditions. We propose a model to explain how restoration behaves in the msH1 system and generate valuable information necessary to develop an efficient system for hybrid wheat production.

Wilderness and backcountry site restoration guide

Treesearch

Lisa Therrell; David Cole; Victor Claassen; Chris Ryan; Mary Ann Davies

2006-01-01

This comprehensive guide focuses on restoration of small-scale impact caused by human actions in wilderness and backcountry areas. The guide's goals are to: 1) Help practitioners develop plans that thoroughly address the question of whether site restoration is the best management action and, if so, develop a site-specific restoration plan that incorporates...

Adrenomedullin and angiopoietin-1 additively restore erectile function in diabetic rats: comparison with the combination therapy of vascular endothelial growth factor and angiopoietin-1.

PubMed

Nishimatsu, Hiroaki; Suzuki, Etsu; Nomiya, Akira; Niimi, Aya; Suzuki, Motofumi; Fujimura, Tetsuya; Fukuhara, Hiroshi; Homma, Yukio

2013-07-01

Erectile dysfunction (ED) is a major health problem. We have shown that adrenomedullin (AM) restores erectile function in diabetic rats. The aim of this study is to explore a better treatment for ED, we examined whether combination of AM and angiopoietin-1 (Ang-1) was more effective to treat ED than treatment with AM alone or Ang-1 alone. We also compared the effect of the combination therapy with that of treatment with vascular endothelial growth factor-A (VEGF-A). Male Wistar rats were injected with streptozotocin (STZ) to induce diabetes. Adenoviruses expressing AM (AdAM), Ang-1 (AdAng-1), and VEGF-A (AdVEGF-A) were injected into the penis 6 weeks after STZ administration. Erectile function, penile histology, and protein expression were analyzed 4 weeks after the injection of the adenoviruses. Intracavernous pressure and mean arterial pressure were measured to evaluate erectile function. The morphology of the penis was analyzed by Elastica van Gieson stain and immunohistochemistry. The expression of α-smooth muscle actin (SMA), VE-cadherin and type I collagen was assessed by Western blot analysis. Infection with AdAM plus AdAng-1 more effectively restored erectile function than infection with AdAM alone or AdAng-1 alone. This combination therapy restored erectile function to a level similar to that observed in the age-matched Wistar rats. Expression of SMA and VE-cadherin increased more significantly in the AdAM plus AdAng-1-treated group than in the AdAM- or AdAng-1-treated group. Although AdVEGF-A infection restored erectile function significantly, it also caused enlargement of the trabeculae of the cavernous body, aberrant angiogenesis, and overproduction of type I collagen. These results suggested that combination therapy with AM and Ang-1 potently restored erectile function and normal morphology of the cavernous body compared with VEGF-A administration. This combination therapy will be useful to treat ED patients with a severely damaged cavernous body. �

Synergy between Sentinel-1 radar time series and Sentinel-2 optical for the mapping of restored areas in Danube delta

NASA Astrophysics Data System (ADS)

Niculescu, Simona; Lardeux, Cédric; Hanganu, Jenica

2018-05-01

Wetlands are important and valuable ecosystems, yet, since 1900, more than 50 % of wetlands have been lost worldwide. An example of altered and partially restored coastal wetlands is the Danube Delta in Romania. Over time, human intervention has manifested itself in more than a quarter of the entire Danube surface. This intervention was brutal and has rendered ecosystem restoration very difficult. Studies for the rehabilitation / re-vegetation were started immediately after the Danube Delta was declared as a Biosphere Reservation in 1990. Remote sensing offers accurate methods for detecting and mapping change in restored wetlands. Vegetation change detection is a powerful indicator of restoration success. The restoration projects use vegetative cover as an important indicator of restoration success. To follow the evolution of the vegetation cover of the restored areas, satellite images radar and optical of last generation have been used, such as Sentinel-1 and Sentinel-2. Indeed the sensor sensitivity to the landscape depends on the wavelength what- ever radar or optical data and their polarization for radar data. Combining this kind of data is particularly relevant for the classification of wetland vegetation, which are associated with the density and size of the vegetation. In addition, the high temporal acquisition frequency of Sentinel-1 which are not sensitive to cloud cover al- low to use temporal signature of the different land cover. Thus we analyse the polarimetric and temporal signature of Sentinel-1 data in order to better understand the signature of the different study classes. In a second phase, we performed classifications based on the Random Forest supervised classification algorithm involving the entire Sentinel-1 time series, then starting from a Sentinel-2 collection and finally involving combinations of Sentinel-1 and -2 data.

The role of EDEM2 compared with EDEM1 in ricin transport from the endoplasmic reticulum to the cytosol.

PubMed

Słomińska-Wojewódzka, Monika; Pawlik, Anna; Sokołowska, Iwona; Antoniewicz, Jakub; Węgrzyn, Grzegorz; Sandvig, Kirsten

2014-02-01

EDEM1 [ER (endoplasmic reticulum)-degradation-enhancing α-mannosidase I-like protein 1] and EDEM2 are crucial regulators of ERAD (ER-associated degradation) that extracts non-native glycoproteins from the calnexin chaperone system. Ricin is a potent plant cytotoxin composed of an A-chain (RTA) connected by a disulfide bond to a cell-binding lectin B-chain (RTB). After endocytic uptake, the toxin is transported retrogradely to the ER, where the enzymatically active RTA is translocated to the cytosol in a similar manner as misfolded ER proteins. This transport is promoted by EDEM1. In the present study we report that EDEM2 is also involved in ricin retrotranslocation out of the ER. However, the role of EDEM1 and EDEM2 in ricin transport to the cytosol seems to differ. EDEM2 promotes ricin retrotranslocation irrespectively of ER translocon accessibility; moreover, co-immunoprecipitation and pull-down studies revealed that more ricin can interact with EDEM2 in comparison with EDEM1. On the other hand, interactions of both lectins with RTA are dependent on the structure of the RTA. Thus our data display a newly discovered role for EDEM2. Moreover, analysis of the involvement of EDEM1 and EDEM2 in ricin retrotranslocation to the cytosol may provide crucial information about general mechanisms of the recognition of ERAD substrates in the ER.

Grp94 Protein Delivers γ-Aminobutyric Acid Type A (GABAA) Receptors to Hrd1 Protein-mediated Endoplasmic Reticulum-associated Degradation.

PubMed

Di, Xiao-Jing; Wang, Ya-Juan; Han, Dong-Yun; Fu, Yan-Lin; Duerfeldt, Adam S; Blagg, Brian S J; Mu, Ting-Wei

2016-04-29

Proteostasis maintenance of γ-aminobutyric acid type A (GABAA) receptors dictates their function in controlling neuronal inhibition in mammalian central nervous systems. However, as a multisubunit, multispan, integral membrane protein, even wild type subunits of GABAA receptors fold and assemble inefficiently in the endoplasmic reticulum (ER). Unassembled and misfolded subunits undergo ER-associated degradation (ERAD), but this degradation process remains poorly understood for GABAA receptors. Here, using the α1 subunits of GABAA receptors as a model substrate, we demonstrated that Grp94, a metazoan-specific Hsp90 in the ER lumen, uses its middle domain to interact with the α1 subunits and positively regulates their ERAD. OS-9, an ER-resident lectin, acts downstream of Grp94 to further recognize misfolded α1 subunits in a glycan-dependent manner. This delivers misfolded α1 subunits to the Hrd1-mediated ubiquitination and the valosin-containing protein-mediated extraction pathway. Repressing the initial ERAD recognition step by inhibiting Grp94 enhances the functional surface expression of misfolding-prone α1(A322D) subunits, which causes autosomal dominant juvenile myoclonic epilepsy. This study clarifies a Grp94-mediated ERAD pathway for GABAA receptors, which provides a novel way to finely tune their function in physiological and pathophysiological conditions. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Embelin Restores Carbapenem Efficacy against NDM-1-Positive Pathogens

PubMed Central

Ning, Nian-Zhi; Liu, Xiong; Chen, Fanghong; Zhou, Peng; Hu, Lihong; Huang, Jian; Li, Zhan; Huang, Jie; Li, Tao; Wang, Hui

2018-01-01

The emergence and spread of carbapenemase in Gram-negative pathogens poses an enormous threat to global public health. New Delhi metallo-β-lactamase-1 (NDM-1) inactivates nearly every class of β-lactam antibiotics, including carbapenem; however, there is no clinically useful NDM-1 inhibitor. Embelin, an important ingredient in traditional herbal medicine, has anti-tumor effects. The current study is the first to discover and examine the inhibitory activity of embelin against β-lactamase NDM-1. The IC50 of embelin was 2.1 ± 0.2 μM when tested against NDM-1 carbapenemase. Most regions of the embelin molecule were buried within NDM-1’s active site, and the hydroxyl group of embelin interacted directly with the metal ion Zn2+, as shown by molecular dynamic simulation. Systematic analysis of the antibacterial activities of embelin and antibiotics demonstrated that embelin restored meropenem activity against a panel of NDM-positive pathogens, such as Escherichia coli, Klebsiella pneumoniae, and Acinetobacter baumannii. Based on these results, embelin could be a promising carbapenem adjuvant candidate against NDM-1-producing bacterial strains. PMID:29422890

Hyperhomocysteinemia, endoplasmic reticulum stress, and alcoholic liver injury

PubMed Central

Ji, Cheng; Kaplowitz, Neil

2004-01-01

Deficiencies in vitamins or other factors (B6, B12, folic acid, betaine) and genetic disorders for the metabolism of the non-protein amino acid-homocysteine (Hcy) lead to hyperhomocysteinemia (HHcy). HHcy is an integral component of several disorders including cardiovascular disease, neurodegeneration, diabetes and alcoholic liver disease. HHcy unleashes mediators of inflammation such as NFκB, IL-1β, IL-6, and IL-8, increases production of intracellular superoxide anion causing oxidative stress and reducing intracellular level of nitric oxide (NO), and induces endoplasmic reticulum (ER) stress which can explain many processes of Hcy-promoted cell injury such as apoptosis, fat accumulation, and inflammation. Animal models have played an important role in determining the biological effects of HHcy. ER stress may also be involved in other liver diseases such as α1-antitrypsin (α1-AT) deficiency and hepatitis C and/or B virus infection. Future research should evaluate the possible potentiative effects of alcohol and hepatic virus infection on ER stress-induced liver injury, study potentially beneficial effects of lowering Hcy and preventing ER stress in alcoholic humans, and examine polymorphism of Hcy metabolizing enzymes as potential risk-factors for the development of HHcy and liver disease. PMID:15188490

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

PubMed

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.

Reciprocal relationship between alpha1,2 mannosidase processing and reglucosylation in the rough endoplasmic reticulum of Man-P-Dol deficient cells.

PubMed

Duvet, S; Chirat, F; Mir, A M; Verbert, A; Dubuisson, J; Cacan, R

2000-02-01

The study of the glycosylation pathway of a mannosylphosphoryldolichol-deficient CHO mutant cell line (B3F7) reveals that truncated Glc(0-3)Man5GlcNAc2 oligosaccharides are transferred onto nascent proteins. Pulse-chase experiments indicate that these newly synthesized glycoproteins are retained in intracellular compartments and converted to Man4GlcNAc2 species. In this paper, we demonstrate that the alpha1,2 mannosidase, which is involved in the processing of Man5GlcNAc2 into Man4GlcNAc2, is located in the rough endoplasmic reticulum. The enzyme was shown to be inhibited by kifunensine and deoxymannojirimycin, indicating that it is a class I mannosidase. In addition, Man4GlcNAc2 species were produced at the expense of Glc1Man5GlcNAc2 species. Thus, the trimming of Man5GlcNAc2 to Man4GlcNAc2, which is catalyzed by this mannosidase, could be involved in the control of the glucose-dependent folding pathway.

Liver-Specific Deletion of Protein-Tyrosine Phosphatase 1B (PTP1B) Improves Metabolic Syndrome and Attenuates Diet-Induced Endoplasmic Reticulum Stress

PubMed Central

Delibegovic, Mirela; Zimmer, Derek; Kauffman, Caitlin; Rak, Kimberly; Hong, Eun-Gyoung; Cho, You-Ree; Kim, Jason K.; Kahn, Barbara B.; Neel, Benjamin G.; Bence, Kendra K.

2009-01-01

OBJECTIVE—The protein tyrosine phosphatase PTP1B is a negative regulator of insulin signaling; consequently, mice deficient in PTP1B are hypersensitive to insulin. Because PTP1B−/− mice have diminished fat stores, the extent to which PTP1B directly regulates glucose homeostasis is unclear. Previously, we showed that brain-specific PTP1B−/− mice are protected against high-fat diet–induced obesity and glucose intolerance, whereas muscle-specific PTP1B−/− mice have increased insulin sensitivity independent of changes in adiposity. Here we studied the role of liver PTP1B in glucose homeostasis and lipid metabolism. RESEARCH DESIGN AND METHODS—We analyzed body mass/adiposity, insulin sensitivity, glucose tolerance, and lipid metabolism in liver-specific PTP1B−/− and PTP1Bfl/fl control mice, fed a chow or high-fat diet. RESULTS—Compared with normal littermates, liver-specific PTP1B−/− mice exhibit improved glucose homeostasis and lipid profiles, independent of changes in adiposity. Liver-specific PTP1B−/− mice have increased hepatic insulin signaling, decreased expression of gluconeogenic genes PEPCK and G-6-Pase, enhanced insulin-induced suppression of hepatic glucose production, and improved glucose tolerance. Liver-specific PTP1B−/− mice exhibit decreased triglyceride and cholesterol levels and diminished expression of lipogenic genes SREBPs, FAS, and ACC. Liver-specific PTP1B deletion also protects against high-fat diet–induced endoplasmic reticulum stress response in vivo, as evidenced by decreased phosphorylation of p38MAPK, JNK, PERK, and eIF2α and lower expression of the transcription factors C/EBP homologous protein and spliced X box-binding protein 1. CONCLUSIONS—Liver PTP1B plays an important role in glucose and lipid metabolism, independent of alterations in adiposity. Inhibition of PTP1B in peripheral tissues may be useful for the treatment of metabolic syndrome and reduction of cardiovascular risk in addition to

Optical properties of composite restorations influenced by dissimilar dentin restoratives.

PubMed

Marjanovic, Jovana; Veljovic, Djordje N; Stasic, Jovana N; Savic-Stankovic, Tatjana; Trifkovic, Branka; Miletic, Vesna

2018-05-01

To evaluate optical properties (color and translucency) of 'sandwich' restorations of resin-based composites and esthetically unfavorable dentin restoratives. Cylindrical 'dentin' specimens (8mm in diameter and 2mm thick, N=5/group) were prepared using EverX Posterior (GC), Biodentine (Septodont), experimental hydroxyapatite (HAP) or conventional composites (Gradia Direct Posterior, GC; Filtek Z250 and Filtek Z500, 3M ESPE). Capping 'enamel' layers were prepared using composites (Gradia Direct Posterior, Filtek Z250 or Z550) of A1 or A3 shade and the following thickness: 0.6, 1 or 2mm. Color (ΔE) and translucency parameter (TP) were determined using a spectrophotometer (VITA Easyshade Advance 4.0, VITA Zahnfabrik). Data were statistically analyzed using analysis of variance with Tukey's post-hoc tests (α=0.05). TP was greatly affected by layer thickness, whilst ΔE depended on shade and layer thickness of the capping composite. HAP and Biodentine showed significantly lower TP and higher ΔE (deviation from 'ideal white') than composites (p<0.05). Greater TP was seen in EverX_composite groups than in corresponding control groups of the same shade and thickness. TP of composites combined with Biodentine or HAP was below 2, lower than the corresponding control groups (p<0.05). Within-group differences of ΔE were greatest in HAP_composite groups. EverX_Gradia and EverX_FiltekZ250 combinations showed the most comparable ΔE with the control groups. A 2mm thick layer of composite covering dentin restoratives with unfavorable esthetics is recommended for a final 'sandwich' restoration that is esthetically comparable to a conventional, mono-composite control restoration. Copyright © 2018 The Academy of Dental Materials. Published by Elsevier Inc. All rights reserved.

Deletion of protein tyrosine phosphatase 1B obliterates endoplasmic reticulum stress-induced myocardial dysfunction through regulation of autophagy.

PubMed

Wang, Shuyi; Chen, Xiyao; Nair, Sreejayan; Sun, Dongdong; Wang, Xiaoming; Ren, Jun

2017-12-01

Endoplasmic reticulum (ER) stress has been demonstrated to prompt various cardiovascular risks although the underlying mechanism remains elusive. Protein tyrosine phosphatase-1B (PTP1B) serves as an essential negative regulator for insulin signaling. This study examined the role of PTP1B in ER stress-induced myocardial anomalies and underlying mechanism involved with a focus on autophagy. WT and PTP1B knockout mice were subjected to the ER stress inducer tunicamycin (1mg/kg). Cardiac function was evaluated with echocardiography and an Ion-Optix MyoCam system. Western blot analysis was used to monitor the levels of ER stress, autophagy and insulin signaling including insulin receptor substrate (IRS), tribbles homolog 3 (TRIB3), Atg5/7, p62 and LC3-II. Our results showed that ER stress resulted in compromised echocardiographic and cardiomyocyte contractile function, intracellular Ca 2+ mishandling, ER stress, O 2 - production, apoptosis, the effects of which (with the exception of ER stress) were significantly attenuated or negated by PTP1B ablation. Levels of serine phosphorylation of IRS-1, TRIB3, Atg5/7, LC3B and the autophagy adaptor p62 were significantly upregulated while IRS-1 tyrosine phosphorylation was reduced by tunicamycin, the effect of which were obliterated by PTP1B ablation. In vitro study revealed that the autophagy inducer rapamycin and TRIB3 overexpression cancelled PTP1B ablation-offered beneficial effects on cardiomyocyte function or O 2 - production in murine cardiomyocytes or H9C2 myoblasts. Antioxidant or gene silencing of TRIB3 mimicked PTP1B ablation-induced protective effects. These findings collectively suggested that PTP1B ablation protects against ER stress-induced cardiac anomalies through regulation of autophagy. Copyright © 2017 Elsevier B.V. All rights reserved.

The failure to express a protein disulphide isomerase-like protein results in a floury endosperm and an endoplasmic reticulum stress response in rice.

PubMed

Han, Xiaohua; Wang, Yihua; Liu, Xi; Jiang, Ling; Ren, Yulong; Liu, Feng; Peng, Cheng; Li, Jingjing; Jin, Ximing; Wu, Fuqing; Wang, Jiulin; Guo, Xiuping; Zhang, Xin; Cheng, Zhijun; Wan, Jianmin

2012-01-01

The rice somaclonal mutant T3612 produces small grains with a floury endosperm, caused by the loose packing of starch granules. The positional cloning of the mutation revealed a deletion in a gene encoding a protein disulphide isomerase-like enzyme (PDIL1-1). In the wild type, PDIL1-1 was expressed throughout the plant, but most intensely in the developing grain. In T3612, its expression was abolished, resulting in a decrease in the activity of plastidial phosphorylase and pullulanase, and an increase in that of soluble starch synthase I and ADP-glucose pyrophosphorylase. The amylopectin in the T3612 endosperm showed an increase in chains with a degree of polymerization 8-13 compared with the wild type. The expression in the mutant's endosperm of certain endoplasmic reticulum stress-responsive genes was noticeably elevated. PDIL1-1 appears to play an important role in starch synthesis. Its absence is associated with endoplasmic reticulum stress in the endosperm, which is likely to underlie the formation of the floury endosperm in the T3612 mutant.

Endoplasmic reticulum stress is induced in the human placenta during labour.

PubMed

Veerbeek, J H W; Tissot Van Patot, M C; Burton, G J; Yung, H W

2015-01-01

Placental endoplasmic reticulum (ER) stress has been postulated in the pathophysiology of pre-eclampsia (PE) and intrauterine growth restriction (IUGR), but its activation remains elusive. Oxidative stress induced by ischaemia/hypoxia-reoxygenation activates ER stress in vitro. Here, we explored whether exposure to labour represents an in vivo model for the study of acute placental ER stress. ER stress markers, GRP78, P-eIF2α and XBP-1, were significantly higher in laboured placentas than in Caesarean-delivered controls localised mainly in the syncytiotrophoblast. The similarities to changes observed in PE/IUGR placentas suggest exposure to labour can be used to investigate induction of ER stress in pathological placentas. Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.

Endoplasmic Reticulum Stress and Oxidative Stress in Cell Fate Decision and Human Disease

PubMed Central

Cao, Stewart Siyan

2014-01-01

Abstract Significance: The endoplasmic reticulum (ER) is a specialized organelle for the folding and trafficking of proteins, which is highly sensitive to changes in intracellular homeostasis and extracellular stimuli. Alterations in the protein-folding environment cause accumulation of misfolded proteins in the ER that profoundly affect a variety of cellular signaling processes, including reduction–oxidation (redox) homeostasis, energy production, inflammation, differentiation, and apoptosis. The unfolded protein response (UPR) is a collection of adaptive signaling pathways that evolved to resolve protein misfolding and restore an efficient protein-folding environment. Recent Advances: Production of reactive oxygen species (ROS) has been linked to ER stress and the UPR. ROS play a critical role in many cellular processes and can be produced in the cytosol and several organelles, including the ER and mitochondria. Studies suggest that altered redox homeostasis in the ER is sufficient to cause ER stress, which could, in turn, induce the production of ROS in the ER and mitochondria. Critical Issues: Although ER stress and oxidative stress coexist in many pathologic states, whether and how these stresses interact is unknown. It is also unclear how changes in the protein-folding environment in the ER cause oxidative stress. In addition, how ROS production and protein misfolding commit the cell to an apoptotic death and contribute to various degenerative diseases is unknown. Future Directions: A greater fundamental understanding of the mechanisms that preserve protein folding homeostasis and redox status will provide new information toward the development of novel therapeutics for many human diseases. Antioxid. Redox Signal. 21, 396–413. PMID:24702237

Small GTPase Sar1 is crucial for proglutelin and α-globulin export from the endoplasmic reticulum in rice endosperm.

PubMed

Tian, Lihong; Dai, Ling Ling; Yin, Zhi Jie; Fukuda, Masako; Kumamaru, Toshihiro; Dong, Xiang Bai; Xu, Xiu Ping; Qu, Le Qing

2013-07-01

Rice seed storage proteins glutelin and α-globulin are synthesized in the endoplasmic reticulum (ER) and deposited in protein storage vacuoles (PSVs). Sar1, a small GTPase, acts as a molecular switch to regulate the assembly of coat protein complex II, which exports secretory protein from the ER to the Golgi apparatus. To reveal the route by which glutelin and α-globulin exit the ER, four putative Sar1 genes (OsSar1a/b/c/d) were cloned from rice, and transgenic rice were generated with Sar1 overexpressed or suppressed by RNA interference (RNAi) specifically in the endosperm under the control of the rice glutelin promoter. Overexpression or suppression of any OsSar1 did not alter the phenotype. However, simultaneous knockdown of OsSar1a/b/c resulted in floury and shrunken seeds, with an increased level of glutelin precursor and decreased level of the mature α- and β-subunit. OsSar1abc RNAi endosperm generated numerous, spherical, novel protein bodies with highly electron-dense matrixes containing both glutelin and α-globulin. Notably, the novel protein bodies were surrounded by ribosomes, showing that they were derived from the ER. Some of the ER-derived dense protein bodies were attached to a blebbing structure containing prolamin. These results indicated that OsSar1a/b/c play a crucial role in storage proteins exiting from the ER, with functional redundancy in rice endosperm, and glutelin and α-globulin transported together from the ER to the Golgi apparatus by a pathway mediated by coat protein complex II.

Restoration Planning | NOAA Gulf Spill Restoration

Science.gov Websites

Mexico. Since then, we have worked with the public and BP to identify and implement early restoration restoration plan for the Gulf of Mexico and took public comments on the draft plan through December 4, 2015

Neuronal-specific endoplasmic reticulum Mg(2+)/Ca(2+) ATPase Ca(2+) sequestration in mixed primary hippocampal culture homogenates.

PubMed

Parsons, J Travis; Sun, David A; DeLorenzo, Robert J; Churn, Severn B

2004-07-01

Endoplasmic reticulum Mg(2+)/Ca(2+) ATPase Ca(2+) sequestration is crucial for maintenance of neuronal Ca(2+) homeostasis. The use of cell culture in conjunction with modern Ca(2+) imaging techniques has been invaluable in elucidating these mechanisms. While imaging protocols evaluate endoplasmic reticulum Ca(2+) loads, measurement of Mg(2+)/Ca(2+) ATPase activity is indirect, comparing cytosolic Ca(2+) levels in the presence or absence of the Mg(2+)/Ca(2+) ATPase inhibitor thapsigargin. Direct measurement of Mg(2+)/Ca(2+) ATPase by isolation of microsomes is impossible due to the minuscule amounts of protein yielded from cultures used for imaging. In the current study, endoplasmic reticulum Mg(2+)/Ca(2+) ATPase Ca(2+) sequestration was measured in mixed homogenates of neurons and glia from primary hippocampal cultures. It was demonstrated that Ca(2+) uptake was mediated by the endoplasmic reticulum Mg(2+)/Ca(2+) ATPase due to its dependence on ATP and Mg(2+), enhancement by oxalate, and inhibition by thapsigargin. It was also shown that neuronal Ca(2+) uptake, mediated by the type 2 sarco(endo)plasmic reticulum Ca(2+) ATPase isoform, could be distinguished from glial Ca(2+) uptake in homogenates composed of neurons and glia. Finally, it was revealed that Ca(2+) uptake was sensitive to incubation on ice, extremely labile in the absence of protease inhibitors, and significantly more stable under storage conditions at -80 degrees C.

Phase III of Early Restoration | NOAA Gulf Spill Restoration

Science.gov Websites

information about this phase of Early Restoration, including fact sheets on each project. The final Phase III 44 projects are documented in a final Record of Decision. Information about Phase III of Early Archive Home Phase III of Early Restoration Phase III of Early Restoration Beach habitat would be restored

[Progress and prospects on evaluation of ecological restoration: a review of the 5th World Conference on Ecological Restoration].

PubMed

Ding, Jing-Yi; Zhao, Wen-Wu

2014-09-01

The 5th World Conference on Ecological Restoration was held in Madison, Wisconsin, USA on October 6-11, 2013. About 1200 delegates from more than 50 countries attended the conference, and discussed the latest developments in different thematic areas of ecological restoration. Discussions on evaluation of ecological restoration were mainly from three aspects: The construction for evaluation indicator system of ecological restoration; the evaluation methods of ecological restoration; monitoring and dynamic evaluation of ecological restoration. The meeting stressed the importance of evaluation in the process of ecological restoration and concerned the challenges in evaluation of ecological restoration. The conference had the following enlightenments for China' s research on evaluation of ecological restoration: 1) Strengthening the construction of comprehensive evaluation indicators system and focusing on the multi-participation in the evaluation process. 2) Paying more attentions on scale effect and scale transformation in the evaluation process of ecological restoration. 3) Expanding the application of 3S technology in assessing the success of ecological restoration and promoting the dynamic monitoring of ecological restoration. 4) Carrying out international exchanges and cooperation actively, and promoting China's international influence in ecological restoration research.

Endocrown restorations: Influence of dental remnant and restorative material on stress distribution.

PubMed

Tribst, João Paulo Mendes; Dal Piva, Amanda Maria de Oliveira; Madruga, Camila Ferreira Leite; Valera, Marcia Carneiro; Borges, Alexandre Luiz Souto; Bresciani, Eduardo; de Melo, Renata Marques

2018-06-20

The goal of this study was to evaluate the stress distribution in a tooth/restoration system according to the factors "amount of dental remnant" (3 levels) and "restorative material" (2 levels). Three endodontically treated maxillary molars were modeled with CAD software for conducting non-linear finite element analysis (FEA), each with a determined amount of dental remnant of 1.5, 3, or 4.5mm. Models were duplicated, and half received restorations in lithium disilicate (IPS e.max CAD), while the other half received leucite ceramic restorations (IPS Empress CAD), both from Ivoclar Vivadent (Schaan, Liechtenstein). The solids were imported to analysis software (ANSYS 17.2, ANSYS Inc., Houston, TX, USA) in STEP format. All contacts involving the resin cement were considered no-separation, whereas between teeth and fixation cylinder, the contact was considered perfectly bonded. The mechanical properties of each structure were reported, and the materials were considered isotropic, linearly elastic, and homogeneous. An axial load (300N) was applied at the occlusal surface (triploidism area). Results were determined by colorimetric graphs of maximum principal stress (MPS) on tooth remnant, cement line, and restoration. MPS revealed that both factors influenced the stress distribution for all structures; the higher the material's elastic modulus, the higher the stress concentration on the restoration and the lower the stress concentration on the cement line. Moreover, the greater the dental crown remnant, the higher the stress concentration on the restoration. Thus, the remaining dental tissue should always be preserved. In situations in which few dental remnants are available, the thicker the restoration, the higher the concentration of stresses in its structure, protecting the adhesive interface from potential adhesive failures. Results are more promising when the endocrown is fabricated with lithium disilicate ceramic. Copyright © 2018 The Academy of Dental Materials

Silencing or inhibition of endoplasmic reticulum aminopeptidase 1 (ERAP1) suppresses free heavy chain expression and Th17 responses in ankylosing spondylitis

PubMed Central

Chen, Liye; Ridley, Anna; Hammitzsch, Ariane; Al-Mossawi, Mohammad Hussein; Bunting, Helen; Georgiadis, Dimitris; Chan, Antoni; Kollnberger, Simon; Bowness, Paul

2016-01-01

Objective Human leucocyte antigen (HLA)-B27 and endoplasmic reticulum aminopeptidase 1 (ERAP1) are strongly associated with ankylosing spondylitis (AS). ERAP1 is a key aminopeptidase in HLA class I presentation and can potentially alter surface expression of HLA-B27 free heavy chains (FHCs). We studied the effects of ERAP1 silencing/inhibition/variations on HLA-B27 FHC expression and Th17 responses in AS. Methods Flow cytometry was used to measure surface expression of HLA class I in peripheral blood mononuclear cells (PBMCs) from patients with AS carrying different ERAP1 genotypes (rs2287987, rs30187 and rs27044) and in ERAP1-silenced/inhibited/mutated HLA-B27-expressing antigen presenting cells (APCs). ERAP1-silenced/inhibited APCs were cocultured with KIR3DL2CD3ε-reporter cells or AS CD4+ T cells. Th17 responses of AS CD4+ T cells were measured by interleukin (IL)-17A ELISA and Th17 intracellular cytokine staining. FHC cell surface expression and Th17 responses were also measured in AS PBMCs following ERAP1 inhibition. Results The AS-protective ERAP1 variants, K528R and Q730E, were associated with reduced surface FHC expression by monocytes from patients with AS and HLA-B27-expressing APCs. ERAP1 silencing or inhibition in APCs downregulated HLA-B27 FHC surface expression, reduced IL-2 production by KIR3DL2CD3ε-reporter cells and suppressed the Th17 expansion and IL-17A secretion by AS CD4+ T cells. ERAP1 inhibition of AS PBMCs reduced HLA class I FHC surface expression by monocytes and B cells, and suppressed Th17 expansion. Conclusions ERAP1 activity determines surface expression of HLA-B27 FHCs and potentially promotes Th17 responses in AS through binding of HLA-B27 FHCs to KIR3DL2. Our data suggest that ERAP1 inhibition has potential for AS treatment. PMID:26130142

A physical/psychological and biological stress combine to enhance endoplasmic reticulum stress

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mondal, Tapan Kumar; Emeny, Rebecca T.; Gao, Donghong

The generation of an immune response against infectious and other foreign agents is substantially modified by allostatic load, which is increased with chemical, physical and/or psychological stressors. The physical/psychological stress from cold-restraint (CR) inhibits host defense against Listeria monocytogenes (LM), due to early effects of the catecholamine norepinephrine (NE) from sympathetic nerves on β1-adrenoceptors (β1AR) of immune cells. Although CR activates innate immunity within 2 h, host defenses against bacterial growth are suppressed 2–3 days after infection (Cao and Lawrence 2002). CR enhances inducible nitric oxide synthase (iNOS) expression and NO production. The early innate activation leads to cellular reduction-oxidationmore » (redox) changes of immune cells. Lymphocytes from CR-treated mice express fewer surface thiols. Splenic and hepatic immune cells also have fewer proteins with free thiols after CR and/or LM, and macrophages have less glutathione after the in vivo CR exposure or exposure to NE in vitro. The early induction of CR-induced oxidative stress elevates endoplasmic reticulum (ER) stress, which could interfere with keeping phagocytized LM within the phagosome or re-encapsuling LM by autophagy once they escape from the phagosome. ER stress-related proteins, such as glucose-regulated protein 78 (GRP78), have elevated expression with CR and LM. The results indicate that CR enhances the unfolded protein response (UPR), which interferes with host defenses against LM. Thus, it is postulated that increased stress, as exists with living conditions at low socioeconomic conditions, can lower host defenses against pathogens because of oxidative and ER stress processes. - Highlights: • Cold-restraint (physical/psychological stress) induces early oxidative stress. • The oxidative stress relates to catecholamine signaling beta-adrenoceptors. • Physical/psychological stress combines infection enhancing inflammation. • Endoplasmic

Sediment Budgets and Sources Inform a Novel Valley Bottom Restoration Practice Impacted by Legacy Sediment: The Big Spring Run, PA, Restoration Experiment

NASA Astrophysics Data System (ADS)

Walter, R. C.; Merritts, D.; Rahnis, M. A.; Gellis, A.; Hartranft, J.; Mayer, P. M.; Langland, M.; Forshay, K.; Weitzman, J. N.; Schwarz, E.; Bai, Y.; Blair, A.; Carter, A.; Daniels, S. S.; Lewis, E.; Ohlson, E.; Peck, E. K.; Schulte, K.; Smith, D.; Stein, Z.; Verna, D.; Wilson, E.

2017-12-01

Big Spring Run (BSR), a small agricultural watershed in southeastern Pennsylvania, is located in the Piedmont Physiographic Province, which has the highest nutrient and sediment yields in the Chesapeake Bay watershed. To effectively reduce nutrient and sediment loading it is important to monitor the effect of management practices on pollutant reduction. Here we present results of an ongoing study, begun in 2008, to understand the impact of a new valley bottom restoration strategy for reducing surface water sediment and nutrient loads. We test the hypotheses that removing legacy sediments will reduce sediment and phosphorus loads, and that restoring eco-hydrological functions of a buried Holocene wetland (Walter & Merritts 2008) will improve surface and groundwater quality by creating accommodation space to trap sediment and process nutrients. Comparisons of pre- and post-restoration gage data show that restoration lowered the annual sediment load by at least 118 t yr-1, or >75%, from the 1000 m-long restoration reach, with the entire reduction accounted for by legacy sediment removal. Repeat RTK-GPS surveys of pre-restoration stream banks verified that >90 t yr-1 of suspended sediment was from bank erosion within the restoration reach. Mass balance calculations of 137Cs data indicate 85-100% of both the pre-restoration and post-restoration suspended sediment storm load was from stream bank sources. This is consistent with trace element data which show that 80-90 % of the pre-restoration outgoing suspended sediment load at BSR was from bank erosion. Meanwhile, an inventory of fallout 137Cs activity from two hill slope transects adjacent to BSR yields average modern upland erosion rates of 2.7 t ha-1 yr-1 and 5.1 t ha-1 yr-1, showing modest erosion on slopes and deposition at toe of slopes. We conclude that upland farm slopes contribute little soil to the suspended sediment supply within this study area, and removal of historic valley bottom sediment effectively

Early Restoration PEIS Public Meeting | NOAA Gulf Spill Restoration

Science.gov Websites

, or PEIS, to evaluate the potential environmental effects of types of early restoration actions, as Early Restoration Plan. The PEIS also will evaluate the cumulative effects of early restoration. We are

Prediction of failure to retain work 1 year after interdisciplinary functional restoration in occupational injuries.

PubMed

Brede, Emily; Mayer, Tom G; Gatchel, Robert J

2012-02-01

To identify risk factors for work retention (a patients' ability to both obtain and retain employment) at 1 year after treatment for a chronic disabling occupational musculoskeletal disorder (CDOMD). Prospective cohort study. Consecutive patients undergoing interdisciplinary functional restoration treatment in a regional rehabilitation referral center. A sample of 1850 consecutive CDOMD patients, who were admitted to and completed a functional restoration program, were subsequently classified as work retention or nonwork retention at a 1-year posttreatment evaluation. Not applicable. Measures, including medical evaluations, demographic and occupational data, psychosocial diagnostic evaluation, and validated measures of pain, disability, and depressive symptoms, were obtained at admission to, and discharge from, the program. Using a multivariate logistic regression analysis, the following variables were found to be significant predictors of failure to retain work: older age (odds ratio [OR]=1.84; 95% confidence interval [CI], 1.33-2.54), female sex (OR=1.46; 95% CI, 1.09-1.94), nonworking status at discharge (OR=1.65; 95% CI, 1.11-2.45), extreme disability at admission (OR=1.46; 95% CI, 1.06-2.00), antisocial personality disorder (OR=2.11; 95% CI, 1.09-4.08), receipt of government disability benefits at admission (OR=2.28; 95% CI, 1.06-4.89), and dependence on opiate pain medications (OR=1.43; 95% CI, 1.02-2.00). The final model improved prediction by 75% over assigning all patients to the larger (work retention) group. This study identified demographic, psychosocial, and occupational factors that were predictive of failure to retain work. These risk factors may be used to individualize treatment plans for CDOMD patients in order to provide optimal functional restoration. Copyright © 2012 American Congress of Rehabilitation Medicine. Published by Elsevier Inc. All rights reserved.

Endoplasmic Reticulum-Plasma Membrane Contact Sites.

PubMed

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.

Ovalbumin-derived precursor peptides are transferred sequentially from gp96 and calreticulin to MHC I in the endoplasmic reticulum

PubMed Central

Kropp, Laura E.; Garg, Manish; Binder, Robert J.

2010-01-01

Cellular peptides generated by proteasomal degradation of proteins in the cytosol and destined for presentation by MHC I are associated with several chaperones. Hsp70, hsp90 and the TCP1-ring complex have been implicated as important cytosolic players for chaperoning these peptides. In this study we report that gp96 and calreticulin are essential for chaperoning peptides in the endoplasmic reticulum. Importantly we demonstrate that cellular peptides are transferred sequentially from gp96 to calreticulin and then to MHC I forming a relay line. Disruption of this relay line by removal of gp96 or calreticulin prevents the binding of peptides by MHC I and hence presentation of the MHC I-peptide complex on the cell surface. Our results are important for understanding how peptides are processed and trafficked within the endoplasmic reticulum before exiting in association with MHC I heavy chains and β2-microglobulin as a trimolecular complex. PMID:20410492

Estrogen receptor α protects pancreatic β-cells from apoptosis by preserving mitochondrial function and suppressing endoplasmic reticulum stress.

PubMed

Zhou, Zhenqi; Ribas, Vicent; Rajbhandari, Prashant; Drew, Brian G; Moore, Timothy M; Fluitt, Amy H; Reddish, Britany R; Whitney, Kate A; Georgia, Senta; Vergnes, Laurent; Reue, Karen; Liesa, Marc; Shirihai, Orian; van der Bliek, Alexander M; Chi, Nai-Wen; Mahata, Sushil K; Tiano, Joseph P; Hewitt, Sylvia C; Tontonoz, Peter; Korach, Kenneth S; Mauvais-Jarvis, Franck; Hevener, Andrea L

2018-03-30

Estrogen receptor α (ERα) action plays an important role in pancreatic β-cell function and survival; thus, it is considered a potential therapeutic target for the treatment of type 2 diabetes in women. However, the mechanisms underlying the protective effects of ERα remain unclear. Because ERα regulates mitochondrial metabolism in other cell types, we hypothesized that ERα may act to preserve insulin secretion and promote β-cell survival by regulating mitochondrial-endoplasmic reticulum (EndoRetic) function. We tested this hypothesis using pancreatic islet-specific ERα knockout (PERαKO) mice and Min6 β-cells in culture with Esr1 knockdown (KD). We found that Esr1-KD promoted reactive oxygen species production that associated with reduced fission/fusion dynamics and impaired mitophagy. Electron microscopy showed mitochondrial enlargement and a pro-fusion phenotype. Mitochondrial cristae and endoplasmic reticulum were dilated in Esr1-KD compared with ERα replete Min6 β-cells. Increased expression of Oma1 and Chop was paralleled by increased oxygen consumption and apoptosis susceptibility in ERα-KD cells. In contrast, ERα overexpression and ligand activation reduced both Chop and Oma1 expression, likely by ERα binding to consensus estrogen-response element sites in the Oma1 and Chop promoters. Together, our findings suggest that ERα promotes β-cell survival and insulin secretion through maintenance of mitochondrial fission/fusion-mitophagy dynamics and EndoRetic function, in part by Oma1 and Chop repression.

Clinical decisions for anterior restorations: the concept of restorative volume.

PubMed

Cardoso, Jorge André; Almeida, Paulo Júlio; Fischer, Alex; Phaxay, Somano Luang

2012-12-01

The choice of the most appropriate restoration for anterior teeth is often a difficult decision. Numerous clinical and technical factors play an important role in selecting the treatment option that best suits the patient and the restorative team. Experienced clinicians have developed decision processes that are often more complex than may seem. Less experienced professionals may find difficulties making treatment decisions because of the widely varied restorative materials available and often numerous similar products offered by different manufacturers. The authors reviewed available evidence and integrated their clinical experience to select relevant factors that could provide a logical and practical guideline for restorative decisions in anterior teeth. The presented concept of restorative volume is based on structural, optical, and periodontal factors. Each of these factors will influence the short- and long-term behavior of restorations in terms of esthetics, biology, and function. Despite the marked evolution of esthetic restorative techniques and materials, significant limitations still exist, which should be addressed by researchers. The presented guidelines must be regarded as a mere orientation for risk analysis. A comprehensive individual approach should always be the core of restorative esthetic treatments. The complex decision process for anterior esthetic restorations can be clarified by a systematized examination of structural, optical, and periodontal factors. The basis for the proposed thought process is the concept of restorative volume that is a contemporary interpretation of restoration categories and their application. © 2012 Wiley Periodicals, Inc.

Restoration in Its Natural Context: How Ecological Momentary Assessment Can Advance Restoration Research

PubMed Central

Beute, Femke; de Kort, Yvonne; IJsselsteijn, Wijnand

2016-01-01

More and more people use self-tracking technologies to track their psychological states, physiology, and behaviors to gain a better understanding of themselves or to achieve a certain goal. Ecological Momentary Assessment (EMA) also offers an excellent opportunity for restorative environments research, which examines how our physical environment (especially nature) can positively influence health and wellbeing. It enables investigating restorative health effects in everyday life, providing not only high ecological validity but also opportunities to study in more detail the dynamic processes playing out over time on recovery, thereby bridging the gap between laboratory (i.e., short-term effects) and epidemiological (long-term effects) research. We have identified four main areas in which self-tracking could help advance restoration research: (1) capturing a rich set of environment types and restorative characteristics; (2) distinguishing intra-individual from inter-individual effects; (3) bridging the gap between laboratory and epidemiological research; and (4) advancing theoretical insights by measuring a more broad range of effects in everyday life. This paper briefly introduces restorative environments research, then reviews the state of the art of self-tracking technologies and methodologies, discusses how these can be implemented to advance restoration research, and presents some examples of pioneering work in this area. PMID:27089352

Deficiency of Rac1 Blocks NADPH Oxidase Activation, Inhibits Endoplasmic Reticulum Stress, and Reduces Myocardial Remodeling in a Mouse Model of Type 1 Diabetes

PubMed Central

Li, Jianmin; Zhu, Huaqing; Shen, E; Wan, Li; Arnold, J. Malcolm O.; Peng, Tianqing

2010-01-01

OBJECTIVE Our recent study demonstrated that Rac1 and NADPH oxidase activation contributes to cardiomyocyte apoptosis in short-term diabetes. This study was undertaken to investigate if disruption of Rac1 and inhibition of NADPH oxidase would prevent myocardial remodeling in chronic diabetes. RESEARCH DESIGN AND METHODS Diabetes was induced by injection of streptozotocin in mice with cardiomyocyte-specific Rac1 knockout and their wild-type littermates. In a separate experiment, wild-type diabetic mice were treated with vehicle or apocynin in drinking water. Myocardial hypertrophy, fibrosis, endoplasmic reticulum (ER) stress, inflammatory response, and myocardial function were investigated after 2 months of diabetes. Isolated adult rat cardiomyocytes were cultured and stimulated with high glucose. RESULTS In diabetic hearts, NADPH oxidase activation, its subunits' expression, and reactive oxygen species production were inhibited by Rac1 knockout or apocynin treatment. Myocardial collagen deposition and cardiomyocyte cross-sectional areas were significantly increased in diabetic mice, which were accompanied by elevated expression of pro-fibrotic genes and hypertrophic genes. Deficiency of Rac1 or apocynin administration reduced myocardial fibrosis and hypertrophy, resulting in improved myocardial function. These effects were associated with a normalization of ER stress markers' expression and inflammatory response in diabetic hearts. In cultured cardiomyocytes, high glucose–induced ER stress was inhibited by blocking Rac1 or NADPH oxidase. CONCLUSIONS Rac1 via NADPH oxidase activation induces myocardial remodeling and dysfunction in diabetic mice. The role of Rac1 signaling may be associated with ER stress and inflammation. Thus, targeting inhibition of Rac1 and NADPH oxidase may be a therapeutic approach for diabetic cardiomyopathy. PMID:20522592

Enhancement of endoplasmic reticulum (ER) degradation of misfolded Null Hong Kong alpha1-antitrypsin by human ER mannosidase I.

PubMed

Hosokawa, Nobuko; Tremblay, Linda O; You, Zhipeng; Herscovics, Annette; Wada, Ikuo; Nagata, Kazuhiro

2003-07-11

Misfolded glycoproteins synthesized in the endoplasmic reticulum (ER) are degraded by cytoplasmic proteasomes, a mechanism known as ERAD (ER-associated degradation). In the present study, we demonstrate that ERAD of the misfolded genetic variant-null Hong Kong alpha1-antitrypsin is enhanced by overexpression of the ER processing alpha1,2-mannosidase (ER ManI) in HEK 293 cells, indicating the importance of ER ManI in glycoprotein quality control. We showed previously that EDEM, an enzymatically inactive mannosidase homolog, interacts with misfolded alpha1-antitrypsin and accelerates its degradation (Hosokawa, N., Wada, I., Hasegawa, K., Yorihuzi, T., Tremblay, L. O., Herscovics, A., and Nagata, K. (2001) EMBO Rep. 2, 415-422). Herein we demonstrate a combined effect of ER ManI and EDEM on ERAD of misfolded alpha1-antitrypsin. We also show that misfolded alpha1-antitrypsin NHK contains labeled Glc1Man9GlcNAc and Man5-9GlcNAc released by endo-beta-N-acetylglucosaminidase H in pulse-chase experiments with [2-3H]mannose. Overexpression of ER ManI greatly increases the formation of Man8GlcNAc, induces the formation of Glc1Man8GlcNAc and increases trimming to Man5-7GlcNAc. We propose a model whereby the misfolded glycoprotein interacts with ER ManI and with EDEM, before being recognized by downstream ERAD components. This detailed characterization of oligosaccharides associated with a misfolded glycoprotein raises the possibility that the carbohydrate recognition determinant triggering ERAD may not be restricted to Man8GlcNAc2 isomer B as previous studies have suggested.

Lack of an Efficient Endoplasmic Reticulum-localized Recycling System Protects Peroxiredoxin IV from Hyperoxidation*

PubMed Central

Cao, Zhenbo; Subramaniam, Suraj; Bulleid, Neil J.

2014-01-01

Typical 2-Cys peroxiredoxins are required to remove hydrogen peroxide from several different cellular compartments. Their activity can be regulated by hyperoxidation and consequent inactivation of the active-site peroxidatic cysteine. Here we developed a simple assay to quantify the hyperoxidation of peroxiredoxins. Hyperoxidation of peroxiredoxins can only occur efficiently in the presence of a recycling system, usually involving thioredoxin and thioredoxin reductase. We demonstrate that there is a marked difference in the sensitivity of the endoplasmic reticulum-localized peroxiredoxin to hyperoxidation compared with either the cytosolic or mitochondrial enzymes. Each enzyme is equally sensitive to hyperoxidation in the presence of a robust recycling system. Our results demonstrate that peroxiredoxin IV recycling in the endoplasmic reticulum is much less efficient than in the cytosol or mitochondria, leading to the protection of peroxiredoxin IV from hyperoxidation. PMID:24403061

Euglycemia Restoration by Central Leptin in Type 1 Diabetes Requires STAT3 Signaling but Not Fast-Acting Neurotransmitter Release.

PubMed

Xu, Yuanzhong; Chang, Jeffrey T; Myers, Martin G; Xu, Yong; Tong, Qingchun

2016-04-01

Central leptin action is sufficient to restore euglycemia in insulinopenic type 1 diabetes (T1D); however, the underlying mechanism remains poorly understood. To examine the role of intracellular signal transducer and activator of transcription 3 (STAT3) pathways, we used LepRs/s mice with disrupted leptin-phosphorylated STAT3 signaling to test the effect of central leptin on euglycemia restoration. These mice developed streptozocin-induced T1D, which was surprisingly not associated with hyperglucagonemia, a typical manifestation in T1D. Further, leptin action on euglycemia restoration was abrogated in these mice, which was associated with refractory hypercorticosteronemia. To examine the role of fast-acting neurotransmitters glutamate and γ-aminobutyric acid (GABA), two major neurotransmitters in the brain, from leptin receptor (LepR) neurons, we used mice with disrupted release of glutamate, GABA, or both from LepR neurons. Surprisingly, all mice responded normally to leptin-mediated euglycemia restoration, which was associated with expected correction from hyperglucagonemia and hyperphagia. In contrast, mice with loss of glutamate and GABA appeared to develop an additive obesity effect over those with loss of single neurotransmitter release. Thus, our study reveals that STAT3 signaling, but not fast-acting neurotransmitter release, is required for leptin action on euglycemia restoration and that hyperglucagonemia is not required for T1D. © 2016 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.

Phytosphingosine degradation pathway includes fatty acid α-oxidation reactions in the endoplasmic reticulum.

PubMed

Kitamura, Takuya; Seki, Naoya; Kihara, Akio

2017-03-28

Although normal fatty acids (FAs) are degraded via β-oxidation, unusual FAs such as 2-hydroxy (2-OH) FAs and 3-methyl-branched FAs are degraded via α-oxidation. Phytosphingosine (PHS) is one of the long-chain bases (the sphingolipid components) and exists in specific tissues, including the epidermis and small intestine in mammals. In the degradation pathway, PHS is converted to 2-OH palmitic acid and then to pentadecanoic acid (C15:0-COOH) via FA α-oxidation. However, the detailed reactions and genes involved in the α-oxidation reactions of the PHS degradation pathway have yet to be determined. In the present study, we reveal the entire PHS degradation pathway: PHS is converted to C15:0-COOH via six reactions [phosphorylation, cleavage, oxidation, CoA addition, cleavage (C1 removal), and oxidation], in which the last three reactions correspond to the α-oxidation. The aldehyde dehydrogenase ALDH3A2 catalyzes both the first and second oxidation reactions (fatty aldehydes to FAs). In Aldh3a2 -deficient cells, the unmetabolized fatty aldehydes are reduced to fatty alcohols and are incorporated into ether-linked glycerolipids. We also identify HACL2 (2-hydroxyacyl-CoA lyase 2) [previous name, ILVBL; ilvB (bacterial acetolactate synthase)-like] as the major 2-OH acyl-CoA lyase involved in the cleavage (C1 removal) reaction in the FA α-oxidation of the PHS degradation pathway. HACL2 is localized in the endoplasmic reticulum. Thus, in addition to the already-known FA α-oxidation in the peroxisomes, we have revealed the existence of FA α-oxidation in the endoplasmic reticulum in mammals.

An additional function of the rough endoplasmic reticulum protein complex prolyl 3-hydroxylase 1·cartilage-associated protein·cyclophilin B: the CXXXC motif reveals disulfide isomerase activity in vitro.

PubMed

Ishikawa, Yoshihiro; Bächinger, Hans Peter

2013-11-01

Collagen biosynthesis occurs in the rough endoplasmic reticulum, and many molecular chaperones and folding enzymes are involved in this process. The folding mechanism of type I procollagen has been well characterized, and protein disulfide isomerase (PDI) has been suggested as a key player in the formation of the correct disulfide bonds in the noncollagenous carboxyl-terminal and amino-terminal propeptides. Prolyl 3-hydroxylase 1 (P3H1) forms a hetero-trimeric complex with cartilage-associated protein and cyclophilin B (CypB). This complex is a multifunctional complex acting as a prolyl 3-hydroxylase, a peptidyl prolyl cis-trans isomerase, and a molecular chaperone. Two major domains are predicted from the primary sequence of P3H1: an amino-terminal domain and a carboxyl-terminal domain corresponding to the 2-oxoglutarate- and iron-dependent dioxygenase domains similar to the α-subunit of prolyl 4-hydroxylase and lysyl hydroxylases. The amino-terminal domain contains four CXXXC sequence repeats. The primary sequence of cartilage-associated protein is homologous to the amino-terminal domain of P3H1 and also contains four CXXXC sequence repeats. However, the function of the CXXXC sequence repeats is not known. Several publications have reported that short peptides containing a CXC or a CXXC sequence show oxido-reductase activity similar to PDI in vitro. We hypothesize that CXXXC motifs have oxido-reductase activity similar to the CXXC motif in PDI. We have tested the enzyme activities on model substrates in vitro using a GCRALCG peptide and the P3H1 complex. Our results suggest that this complex could function as a disulfide isomerase in the rough endoplasmic reticulum.

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

PubMed

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.

Lysosomes shape Ins(1,4,5)P3-evoked Ca2+ signals by selectively sequestering Ca2+ released from the endoplasmic reticulum

PubMed Central

López-Sanjurjo, Cristina I.; Tovey, Stephen C.; Prole, David L.; Taylor, Colin W.

2013-01-01

Summary Most intracellular Ca2+ signals result from opening of Ca2+ channels in the plasma membrane or endoplasmic reticulum (ER), and they are reversed by active transport across these membranes or by shuttling Ca2+ into mitochondria. Ca2+ channels in lysosomes contribute to endo-lysosomal trafficking and Ca2+ signalling, but the role of lysosomal Ca2+ uptake in Ca2+ signalling is unexplored. Inhibition of lysosomal Ca2+ uptake by dissipating the H+ gradient (using bafilomycin A1), perforating lysosomal membranes (using glycyl-L-phenylalanine 2-naphthylamide) or lysosome fusion (using vacuolin) increased the Ca2+ signals evoked by receptors that stimulate inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] formation. Bafilomycin A1 amplified the Ca2+ signals evoked by photolysis of caged Ins(1,4,5)P3 or by inhibition of ER Ca2+ pumps, and it slowed recovery from them. Ca2+ signals evoked by store-operated Ca2+ entry were unaffected by bafilomycin A1. Video-imaging with total internal reflection fluorescence microscopy revealed that lysosomes were motile and remained intimately associated with the ER. Close association of lysosomes with the ER allows them selectively to accumulate Ca2+ released by Ins(1,4,5)P3 receptors. PMID:23097044

Cytochrome P450 Organization and Function Are Modulated by Endoplasmic Reticulum Phospholipid Heterogeneity.

PubMed

Brignac-Huber, Lauren M; Park, Ji Won; Reed, James R; Backes, Wayne L

2016-12-01

Cytochrome P450s (P450s) comprise a superfamily of proteins that catalyze numerous monooxygenase reactions in animals, plants, and bacteria. In eukaryotic organisms, these proteins not only carry out reactions necessary for the metabolism of endogenous compounds, but they are also important in the oxidation of exogenous drugs and other foreign compounds. Eukaryotic P450 system proteins generally reside in membranes, primarily the endoplasmic reticulum or the mitochondrial membrane. These membranes provide a scaffold for the P450 system proteins that facilitate interactions with their redox partners as well as other P450s. This review focuses on the ability of specific lipid components to influence P450 activities, as well as the role of the membrane in P450 function. These studies have shown that P450s and NADPH-cytochrome P450 reductase appear to selectively associate with specific phospholipids and that these lipid-protein interactions influence P450 activities. Finally, because of the heterogeneous nature of the endoplasmic reticulum as well as other biologic membranes, the phospholipids are not arranged randomly but associate to generate lipid microdomains. Together, these characteristics can affect P450 function by 1) altering the conformation of the proteins, 2) influencing the P450 interactions with their redox partners, and 3) affecting the localization of the proteins into specific membrane microdomains. Copyright © 2016 by The American Society for Pharmacology and Experimental Therapeutics.

Restored PB1-F2 in the 2009 Pandemic H1N1 Influenza Virus Has Minimal Effects in Swine

PubMed Central

Pena, Lindomar; Loving, Crystal L.; Henningson, Jamie N.; Lager, Kelly M.; Lorusso, Alessio

2012-01-01

PB1-F2 is an 87- to 90-amino-acid-long protein expressed by certain influenza A viruses. Previous studies have shown that PB1-F2 contributes to virulence in the mouse model; however, its role in natural hosts—pigs, humans, or birds—remains largely unknown. Outbreaks of domestic pigs infected with the 2009 pandemic H1N1 influenza virus (pH1N1) have been detected worldwide. Unlike previous pandemic strains, pH1N1 viruses do not encode a functional PB1-F2 due to the presence of three stop codons resulting in premature truncation after codon 11. However, pH1N1s have the potential to acquire the full-length form of PB1-F2 through mutation or reassortment. In this study, we assessed whether restoring the full-length PB1-F2 open reading frame (ORF) in the pH1N1 background would have an effect on virus replication and virulence in pigs. Restoring the PB1-F2 ORF resulted in upregulation of viral polymerase activity at early time points in vitro and enhanced virus yields in porcine respiratory explants and in the lungs of infected pigs. There was an increase in the severity of pneumonia in pigs infected with isogenic virus expressing PB1-F2 compared to the wild-type (WT) pH1N1. The extent of microscopic pneumonia correlated with increased pulmonary levels of alpha interferon and interleukin-1β in pigs infected with pH1N1 encoding a functional PB1-F2 but only early in the infection. Together, our results indicate that PB1-F2 in the context of pH1N1 moderately modulates viral replication, lung histopathology, and local cytokine response in pigs. PMID:22379102

Plant Endoplasmic Reticulum-Plasma Membrane Contact Sites.

PubMed

Wang, Pengwei; Hawes, Chris; Hussey, Patrick J

2017-04-01

The endoplasmic reticulum (ER) acts as a superhighway with multiple sideroads that connects the different membrane compartments including the ER to the plasma membrane (PM). ER-PM contact sites (EPCSs) are a common feature in eukaryotic organisms, but have not been studied well in plants owing to the lack of molecular markers and to the difficulty in resolving the EPCS structure using conventional microscopy. Recently, however, plant protein complexes required for linking the ER and PM have been identified. This is a further step towards understanding the structure and function of plant EPCSs. We highlight some recent studies in this field and suggest several hypotheses that relate to the possible function of EPCSs in plants. Copyright © 2016. Published by Elsevier Ltd.

Endoplasmic reticulum: ER stress regulates mitochondrial bioenergetics

PubMed Central

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

A restoration practitioner's guide to the restoration gene pool concept

Treesearch

Thomas A. Jones; Thomas A. Monaco

2007-01-01

Choosing plant materials for each desired species is often one of the most difficult steps in developing a restoration plan. The Restoration Gene Pool concept was developed to clarify the options available to the ecological restoration practitioner in terms of plant materials. We present a decision-making flowchart incorporating the issues delineated in the Restoration...

Flow Restoration in the Columbia River Basin: An Evaluation of a Flow Restoration Accounting Framework

NASA Astrophysics Data System (ADS)

McCoy, Amy L.; Holmes, S. Rankin; Boisjolie, Brett A.

2018-03-01

Securing environmental flows in support of freshwater biodiversity is an evolving field of practice. An example of a large-scale program dedicated to restoring environmental flows is the Columbia Basin Water Transactions Program in the Pacific Northwest region of North America, which has been restoring flows in dewatered tributary habitats for imperiled salmon species over the past decade. This paper discusses a four-tiered flow restoration accounting framework for tracking the implementation and impacts of water transactions as an effective tool for adaptive management. The flow restoration accounting framework provides compliance and flow accounting information to monitor transaction efficacy. We review the implementation of the flow restoration accounting framework monitoring framework to demonstrate (a) the extent of water transactions that have been implemented over the past decade, (b) the volumes of restored flow in meeting flow targets for restoring habitat for anadromous fish species, and (c) an example of aquatic habitat enhancement that resulted from Columbia Basin Water Transactions Program investments. Project results show that from 2002 to 2015, the Columbia Basin Water Transactions Program has completed more than 450 water rights transactions, restoring approximately 1.59 million megaliters to date, with an additional 10.98 million megaliters of flow protected for use over the next 100 years. This has resulted in the watering of over 2414 stream kilometers within the Columbia Basin. We conclude with a discussion of the insights gained through the implementation of the flow restoration accounting framework. Understanding the approach and efficacy of a monitoring framework applied across a large river basin can be informative to emerging flow-restoration and adaptive management efforts in areas of conservation concern.

Flow Restoration in the Columbia River Basin: An Evaluation of a Flow Restoration Accounting Framework.

PubMed

McCoy, Amy L; Holmes, S Rankin; Boisjolie, Brett A

2018-03-01

Securing environmental flows in support of freshwater biodiversity is an evolving field of practice. An example of a large-scale program dedicated to restoring environmental flows is the Columbia Basin Water Transactions Program in the Pacific Northwest region of North America, which has been restoring flows in dewatered tributary habitats for imperiled salmon species over the past decade. This paper discusses a four-tiered flow restoration accounting framework for tracking the implementation and impacts of water transactions as an effective tool for adaptive management. The flow restoration accounting framework provides compliance and flow accounting information to monitor transaction efficacy. We review the implementation of the flow restoration accounting framework monitoring framework to demonstrate (a) the extent of water transactions that have been implemented over the past decade, (b) the volumes of restored flow in meeting flow targets for restoring habitat for anadromous fish species, and (c) an example of aquatic habitat enhancement that resulted from Columbia Basin Water Transactions Program investments. Project results show that from 2002 to 2015, the Columbia Basin Water Transactions Program has completed more than 450 water rights transactions, restoring approximately 1.59 million megaliters to date, with an additional 10.98 million megaliters of flow protected for use over the next 100 years. This has resulted in the watering of over 2414 stream kilometers within the Columbia Basin. We conclude with a discussion of the insights gained through the implementation of the flow restoration accounting framework. Understanding the approach and efficacy of a monitoring framework applied across a large river basin can be informative to emerging flow-restoration and adaptive management efforts in areas of conservation concern.

Resetting translational homeostasis restores myelination in Charcot-Marie-Tooth disease type 1B mice.

PubMed

D'Antonio, Maurizio; Musner, Nicolò; Scapin, Cristina; Ungaro, Daniela; Del Carro, Ubaldo; Ron, David; Feltri, M Laura; Wrabetz, Lawrence

2013-04-08

P0 glycoprotein is an abundant product of terminal differentiation in myelinating Schwann cells. The mutant P0S63del causes Charcot-Marie-Tooth 1B neuropathy in humans, and a very similar demyelinating neuropathy in transgenic mice. P0S63del is retained in the endoplasmic reticulum of Schwann cells, where it promotes unfolded protein stress and elicits an unfolded protein response (UPR) associated with translational attenuation. Ablation of Chop, a UPR mediator, from S63del mice completely rescues their motor deficit and reduces active demyelination by half. Here, we show that Gadd34 is a detrimental effector of CHOP that reactivates translation too aggressively in myelinating Schwann cells. Genetic or pharmacological limitation of Gadd34 function moderates translational reactivation, improves myelination in S63del nerves, and reduces accumulation of P0S63del in the ER. Resetting translational homeostasis may provide a therapeutic strategy in tissues impaired by misfolded proteins that are synthesized during terminal differentiation.

Seismic data restoration with a fast L1 norm trust region method

NASA Astrophysics Data System (ADS)

Cao, Jingjie; Wang, Yanfei

2014-08-01

Seismic data restoration is a major strategy to provide reliable wavefield when field data dissatisfy the Shannon sampling theorem. Recovery by sparsity-promoting inversion often get sparse solutions of seismic data in a transformed domains, however, most methods for sparsity-promoting inversion are line-searching methods which are efficient but are inclined to obtain local solutions. Using trust region method which can provide globally convergent solutions is a good choice to overcome this shortcoming. A trust region method for sparse inversion has been proposed, however, the efficiency should be improved to suitable for large-scale computation. In this paper, a new L1 norm trust region model is proposed for seismic data restoration and a robust gradient projection method for solving the sub-problem is utilized. Numerical results of synthetic and field data demonstrate that the proposed trust region method can get excellent computation speed and is a viable alternative for large-scale computation.

Hepatitis C virus envelope components alter localization of hepatocyte tight junction-associated proteins and promote occludin retention in the endoplasmic reticulum.

PubMed

Benedicto, Ignacio; Molina-Jiménez, Francisca; Barreiro, Olga; Maldonado-Rodríguez, Alejandra; Prieto, Jesús; Moreno-Otero, Ricardo; Aldabe, Rafael; López-Cabrera, Manuel; Majano, Pedro L

2008-10-01

Hepatocyte tight junctions (TJ) play key roles in characteristic liver functions, including bile formation and secretion. Infection by hepatitis C virus (HCV) may cause alterations of the liver architecture and disruption of the bile duct, which ultimately can lead to cholestasis. Herein, we employed the HCV replicon system to analyze the effect of HCV on TJ organization. TJ-associated proteins occludin, claudin-1, and Zonula Occludens protein-1 (ZO-1) disappeared from their normal localization at the border of adjacent cells in Huh7 clones harboring genomic but not subgenomic replicons expressing only the nonstructural proteins. Furthermore, cells containing genomic replicons showed a cytoplasmic accumulation of occludin in the endoplasmic reticulum (ER). TJ-associated function, measured as FITC-dextran paracellular permeability, of genomic replicon-containing cells, was also altered. Interestingly, clearance of the HCV replicon by interferon-alpha (IFN-alpha) treatment and by short hairpin RNA (shRNA) significantly restored the localization of TJ-associated proteins. Transient expression of all HCV structural proteins, but not core protein alone, altered the localization of TJ-associated proteins in Huh7 cells and in clones with subgenomic replicons. Confocal analysis showed that accumulation of occludin in the ER partially co-localized with HCV envelope glycoprotein E2. E2/occludin association was further confirmed by co-immunoprecipitation and pull-down assays. Additionally, using a cell culture model of HCV infection, we observed the cytoplasmic dot-like accumulation of occludin in infected Huh7 cells. We propose that HCV structural proteins, most likely those of the viral envelope, promote alterations of TJ-associated proteins, which may provide new insights for HCV-related pathogenesis.

Aerobic exercise training rescues cardiac protein quality control and blunts endoplasmic reticulum stress in heart failure rats.

PubMed

Bozi, Luiz H M; Jannig, Paulo R; Rolim, Natale; Voltarelli, Vanessa A; Dourado, Paulo M M; Wisløff, Ulrik; Brum, Patricia C

2016-11-01

Cardiac endoplasmic reticulum (ER) stress through accumulation of misfolded proteins plays a pivotal role in cardiovascular diseases. In an attempt to reestablish ER homoeostasis, the unfolded protein response (UPR) is activated. However, if ER stress persists, sustained UPR activation leads to apoptosis. There is no available therapy for ER stress relief. Considering that aerobic exercise training (AET) attenuates oxidative stress, mitochondrial dysfunction and calcium imbalance, it may be a potential strategy to reestablish cardiac ER homoeostasis. We test the hypothesis that AET would attenuate impaired cardiac ER stress after myocardial infarction (MI). Wistar rats underwent to either MI or sham surgeries. Four weeks later, rats underwent to 8 weeks of moderate-intensity AET. Myocardial infarction rats displayed cardiac dysfunction and lung oedema, suggesting heart failure. Cardiac dysfunction in MI rats was paralleled by increased protein levels of UPR markers (GRP78, DERLIN-1 and CHOP), accumulation of misfolded and polyubiquitinated proteins, and reduced chymotrypsin-like proteasome activity. These results suggest an impaired cardiac protein quality control. Aerobic exercise training improved exercise capacity and cardiac function of MI animals. Interestingly, AET blunted MI-induced ER stress by reducing protein levels of UPR markers, and accumulation of both misfolded and polyubiquinated proteins, which was associated with restored proteasome activity. Taken together, our study provide evidence for AET attenuation of ER stress through the reestablishment of cardiac protein quality control, which contributes to better cardiac function in post-MI heart failure rats. These results reinforce the importance of AET as primary non-pharmacological therapy to cardiovascular disease. © 2016 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

Influence of restorative material and proximal cavity design on the fracture resistance of MOD inlay restoration.

PubMed

Liu, Xiaozhou; Fok, Alex; Li, Haiyan

2014-03-01

This study aimed to evaluate the effects of the restorative material and cavity design on the facture resistance of inlay restorations under a compressive load using acoustic emission (AE) measurement. Two restorative materials, a composite resin (MZ100, 3M ESPE) and a ceramic (IPS Empress CAD, Ivoclar Vivadent), and two cavity designs, non-proximal box and proximal box, were studied. Thirty-two extracted human third molars were selected and divided into 4 groups. The restorative materials and cavity designs used for the four groups were: (1) composite and non-proximal box; (2) ceramic and non-proximal box; (3) composite and proximal box; (4) ceramic and proximal box. The restored molars were loaded in a MTS machine via a loading head of diameter 10mm. The rate of loading was 0.1mm/min. During loading, an AE system was used to monitor the debonding and fracture of the specimens. The load corresponding to the first AE event, the final maximum load sustained, as well as the total number of AE events recorded were used to evaluate the fracture resistance of the restored teeth. For the initial fracture load, Group 2 (236.15N)1 (428.14N)1 (2003.82N)Group 2 (1685)>Group 3 (239)>Group 1 (221). The differences from pairwise comparisons in the initial fracture load and final load were mostly insignificant statistically (p>0.05), the only exception being that between Groups 2 and 3 in the initial fracture load (p=0.039). For the total number of AE events, statistically significant differences (p<0.05) were found between all group pairs that involved different materials, with the composite groups giving much fewer AE events than the ceramic groups. Conversely, no statistically significant difference in the AE results was found between groups with the same material

Inhibition of PTP1B Restores IRS1-Mediated Hepatic Insulin Signaling in IRS2-Deficient Mice

PubMed Central

González-Rodríguez, Águeda; Gutierrez, Jose A. Mas; Sanz-González, Silvia; Ros, Manuel; Burks, Deborah J.; Valverde, Ángela M.

2010-01-01

OBJECTIVE Mice with complete deletion of insulin receptor substrate 2 (IRS2) develop hyperglycemia, impaired hepatic insulin signaling, and elevated gluconeogenesis, whereas mice deficient for protein tyrosine phosphatase (PTP)1B display an opposing hepatic phenotype characterized by increased sensitivity to insulin. To define the relationship between these two signaling pathways in the regulation of liver metabolism, we used genetic and pharmacological approaches to study the effects of inhibiting PTP1B on hepatic insulin signaling and expression of gluconeogenic enzymes in IRS2−/− mice. RESEARCH DESIGN AND METHODS We analyzed glucose homeostasis and insulin signaling in liver and isolated hepatocytes from IRS2−/− and IRS2−/−/PTP1B−/− mice. Additionally, hepatic insulin signaling was assessed in control and IRS2−/− mice treated with resveratrol, an antioxidant present in red wine. RESULTS In livers of hyperglycemic IRS2−/− mice, the expression levels of PTP1B and its association with the insulin receptor (IR) were increased. The absence of PTP1B in the double-mutant mice restored hepatic IRS1-mediated phosphatidylinositol (PI) 3-kinase/Akt/Foxo1 signaling. Moreover, resveratrol treatment of hyperglycemic IRS2−/− mice decreased hepatic PTP1B mRNA and inhibited PTP1B activity, thereby restoring IRS1-mediated PI 3-kinase/Akt/Foxo1 signaling and peripheral insulin sensitivity. CONCLUSIONS By regulating the phosphorylation state of IR, PTB1B determines sensitivity to insulin in liver and exerts a unique role in the interplay between IRS1 and IRS2 in the modulation of hepatic insulin action. PMID:20028942

Dietary toxins, endoplasmic reticulum (ER) stress and diabetes.

PubMed

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.

Patterns and partners for chiral symmetry restoration

NASA Astrophysics Data System (ADS)

Gómez Nicola, A.; Ruiz de Elvira, J.

2018-04-01

We present and analyze a new set of Ward Identities which shed light on the distinction between different patterns of chiral symmetry restoration in QCD, namely O (4 ) vs O (4 )×U (1 )A. The degeneracy of chiral partners for all scalar and pseudoscalar meson nonet members is studied through their corresponding correlators. Around chiral symmetry degeneration of O (4 ) partners, our analysis predicts that U (1 )A partners are also degenerated. Our analysis also leads to I =1 /2 scalar-pseudoscalar partner degeneration at exact chiral restoration and supports ideal mixing between the η - η' and the f0(500 )- f0(980 ) mesons at O (4 )×U (1 )A restoration, with a possible range where the pseudoscalar mixing vanishes if the two transitions are well separated. We test our results with lattice data and provide further relevant observables regarding chiral and U (1 )A restoration for future lattice and model analyses.

Further Treatments of Root-filled Teeth in the Swedish Adult Population: A Comparison of Teeth Restored with Direct and Indirect Coronal Restorations.

PubMed

Dawson, Victoria S; Isberg, Per-Erik; Kvist, Thomas; Fransson, Helena

2017-09-01

The aim of this study was to evaluate the frequencies of nonsurgical retreatment, root-end surgery, extraction, and further restoration of root-filled teeth in Sweden in 2009 during a follow-up period of 5 years and to compare the outcomes in teeth restored with direct or indirect restorations. Data from the Swedish Social Insurance Agency were analyzed, and the frequencies of nonsurgical retreatment, root-end surgery, extractions, and further restoration were calculated for all teeth registered as root filled during 2009. Chi-square tests were applied to detect any significant differences in the frequency of further treatment in teeth registered as restored with either a direct or an indirect restoration within 6 months of root filling. Of the 248,299 teeth reported root filled in Sweden in 2009, nonsurgical retreatment was registered in 2.2%, root-end surgery in 1.0%, and extractions in 9.2% during the follow-up period. Of the teeth restored with a direct restoration within 6 months after the root filling, 30.3% were registered as having undergone at least 1 further direct restoration; the corresponding percentage of teeth with indirect restorations was 6.4%. A statistically significant difference in the frequency of nonsurgical retreatment, extraction, and further restoration was found; teeth restored with an indirect restoration within 6 months of root filling had fewer of these treatments than those restored by direct restoration. Low frequencies of nonsurgical retreatment and root-end surgery were reported 5 years after root filling, whereas extraction was more common. Fewer additional treatment procedures were registered for teeth with indirect restorations than for those with direct restorations. Copyright © 2017 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

An Arabidopsis Prenylated Rab Acceptor 1 Isoform, AtPRA1.B6, Displays Differential Inhibitory Effects on Anterograde Trafficking of Proteins at the Endoplasmic Reticulum1[W][OA

PubMed Central

Lee, Myoung Hui; Jung, Chanjin; Lee, Junho; Kim, Soo Youn; Lee, Yongjik; Hwang, Inhwan

2011-01-01

Prenylated Rab acceptors (PRAs), members of the Ypt-interacting protein family of small membrane proteins, are thought to aid the targeting of prenylated Rabs to their respective endomembrane compartments. In plants, the Arabidopsis (Arabidopsis thaliana) PRA1 family contains 19 members that display varying degrees of sequence homology to animal PRA1 and localize to the endoplasmic reticulum (ER) and/or endosomes. However, the exact role of these proteins remains to be fully characterized. In this study, the effect of AtPRA1.B6, a member of the AtPRA1 family, on the anterograde trafficking of proteins targeted to various endomembrane compartments was investigated. High levels of AtPRA1.B6 resulted in differential inhibition of coat protein complex II vesicle-mediated anterograde trafficking. The trafficking of the vacuolar proteins sporamin:GFP (for green fluorescent protein) and AALP:GFP, the secretory protein invertase:GFP, and the plasma membrane proteins PMP:GFP and H+-ATPase:GFP was inhibited in a dose-dependent manner, while the trafficking of the Golgi-localized proteins ST:GFP and KAM1(ΔC):mRFP was not affected. Conversely, in RNA interference plants displaying lower levels of AtPRA1.B6 transcripts, the trafficking efficiency of sporamin:GFP and AALP:GFP to the vacuole was increased. Localization and N-glycan pattern analyses of cargo proteins revealed that AtPRA1.B6-mediated inhibition of anterograde trafficking occurs at the ER. In addition, AtPRA1.B6 levels were controlled by cellular processes, including 26S proteasome-mediated proteolysis. Based on these results, we propose that AtPRA1.B6 is a negative regulator of coat protein complex II vesicle-mediated anterograde trafficking for a subset of proteins at the ER. PMID:21828250

The RESTORE program of restorative justice for sex crimes: vision, process, and outcomes.

PubMed

Koss, Mary P

2014-06-01

The article reports empirical evaluation of RESTORE, a restorative justice (RJ) conferencing program adapted to prosecutor-referred adult misdemeanor and felony sexual assaults. RESTORE conferences included voluntary enrollment, preparation, and a face-to-face meeting where primary and secondary victims voice impacts, and responsible persons acknowledge their acts and together develop a re-dress plan that is supervised for 1 year. Process data included referral and consent rates, participant characteristics, observational ratings of conferences compared with program design, services delivered, and safety monitoring. Outcome evaluation used 22 cases to assess (a) pre-post reasons for choosing RESTORE, (b) preparation and conference experiences, (c) overall program and justice satisfaction, and (d) completion rates. This is the first peer-reviewed quantitative evaluation of RJ conferencing for adult sexual assault. Although the data have limitations, the results support cautious optimism regarding feasibility, safety, and satisfactory outcomes. They help envision how conferencing could expand and individualize justice options for sexual assault.

Restoring and rehabilitating sagebrush habitats

USGS Publications Warehouse

Pyke, David A.; Knick, S.T.; Connelly, J.W.

2011-01-01

Less than half of the original habitat of the Greater Sage-Grouse (Centrocercus uropha-sianus) currently exists. Some has been perma-nently lost to farms and urban areas, but the remaining varies in condition from high quality to no longer adequate. Restoration of sagebrush (Artemisia spp.) grassland ecosystems may be pos-sible for resilient lands. However, Greater Sage-Grouse require a wide variety of habitats over large areas to complete their life cycle. Effective restoration will require a regional approach for prioritizing and identifying appropriate options across the landscape. A landscape triage method is recommended for prioritizing lands for restora-tion. Spatial models can indicate where to protect and connect intact quality habitat with other simi-lar habitat via restoration. The ecological site con-cept of land classification is recommended for characterizing potential habitat across the region along with their accompanying state and transi-tion models of plant community dynamics. These models assist in identifying if passive, manage-ment-based or active, vegetation manipulation?based restoration might accomplish the goals of improved Greater Sage-Grouse habitat. A series of guidelines help formulate questions that manag-ers might consider when developing restoration plans: (1) site prioritization through a landscape triage; (2) soil verification and the implications of soil features on plant establishment success; (3) a comparison of the existing plant community to the potential for the site using ecological site descriptions; (4) a determination of the current successional status of the site using state and transition models to aid in predicting if passive or active restoration is necessary; and (5) implemen-tation of post-treatment monitoring to evaluate restoration effectiveness and post-treatment man-agement implications to restoration success.

Phase IV of Early Restoration | NOAA Gulf Spill Restoration

Science.gov Websites

Trustees published the Final Phase IV Early Restoration Plan and Environmental Assessments. The plan habitats. Useful Links: Final Phase IV Early Restoration Plan and Environmental Assessments (pdf, 4.8 MB ) Final Phase IV Early Restoration Plan and Environmental Assessments Executive Summary (pdf, 729 KB

Environmental Factors Contribute to β Cell Endoplasmic Reticulum Stress and Neo-Antigen Formation in Type 1 Diabetes