Role for ribosome-associated complex and stress-seventy subfamily B (RAC-Ssb) in integral membrane protein translation.

Science.gov (United States)

Acosta-Sampson, Ligia; Döring, Kristina; Lin, Yuping; Yu, Vivian Y; Bukau, Bernd; Kramer, Günter; Cate, Jamie H D

2017-12-01

Targeting of most integral membrane proteins to the endoplasmic reticulum is controlled by the signal recognition particle, which recognizes a hydrophobic signal sequence near the protein N terminus. Proper folding of these proteins is monitored by the unfolded protein response and involves protein degradation pathways to ensure quality control. Here, we identify a new pathway for quality control of major facilitator superfamily transporters that occurs before the first transmembrane helix, the signal sequence recognized by the signal recognition particle, is made by the ribosome. Increased rates of translation elongation of the N-terminal sequence of these integral membrane proteins can divert the nascent protein chains to the ribosome-associated complex and stress-seventy subfamily B chaperones. We also show that quality control of integral membrane proteins by ribosome-associated complex-stress-seventy subfamily B couples translation rate to the unfolded protein response, which has implications for understanding mechanisms underlying human disease and protein production in biotechnology. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

The eIF4E-binding proteins are modifiers of cytoplasmic eIF4E relocalization during the heat shock response.

Science.gov (United States)

Sukarieh, R; Sonenberg, N; Pelletier, J

2009-05-01

Stress granules (SGs) arise as a consequence of cellular stress, contain stalled translation preinitiation complexes, and are associated with cell survival during environmental insults. SGs are dynamic entities with proteins relocating into and out of them during stress. Among the repertoire of proteins present in SGs is eukaryotic initiation factor 4E (eIF4E), a translation factor required for cap-dependent translation and that regulates a rate-limiting step for protein synthesis. Herein, we demonstrate that localization of eIF4E to SGs is dependent on the presence of a family of repressor proteins, eIF4E-binding proteins (4E-BPs). Our results demonstrate that 4E-BPs regulate the SG localization of eIF4E.

(−)-EPICATECHIN IMPROVES MITOCHONDRIAL RELATED PROTEIN LEVELS AND AMELIORATES OXIDATIVE STRESS IN DYSTROPHIC DELTA SARCOGLYCAN NULL MOUSE STRIATED MUSCLE

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Ramirez-Sanchez, Israel; De los Santos, Sergio; Gonzalez-Basurto, Silvia; Canto, Patricia; Mendoza-Lorenzo, Patricia; Palma-Flores, Carlos; Ceballos-Reyes, Guillermo; Villarreal, Francisco; Zentella-Dehesa, Alejandro; Coral-Vazquez, Ramon

2014-01-01

Muscular dystrophies (MD) are a group of heterogeneous genetic disorders characterized by progressive striated muscle wasting and degeneration. Although the genetic basis for many of these disorders has been identified, the exact mechanism for disease pathogenesis remains unclear. The presence of oxidative stress (OS) is known to contribute to the pathophysiology and severity of the MD. Mitochondrial dysfunction is observed in MD and likely represents an important determinant of increased OS. Experimental antioxidant therapies have been implemented with the aim of protecting against disease progression, but results from clinical trials have been disappointing. In this study, we explored the capacity of the cacao flavonoid (−)-epicatechin (Epi) to mitigate OS by acting as a positive regulator of mitochondrial structure/function endpoints and redox balance control systems in skeletal and cardiac muscles of dystrophic, δ-sarcoglycan (δ-SG) null mice. Wild type or δ-SG null 2.5 month old male mice were treated via oral gavage with either water (control animals) or Epi (1 mg/kg, twice/day) for 2 weeks. Results evidence a significant normalization of total protein carbonylation, recovery of reduced/oxidized glutathione (GSH/GSSG ratio) and enhanced superoxide dismutase 2, catalase and citrate synthase activities with Epi treatment. These effects were accompanied by increases in protein levels for thiolredoxin, glutathione peroxidase, superoxide dismutase 2, catalase and mitochondrial endpoints. Furthermore, we evidence decreases in heart and skeletal muscle fibrosis, accompanied with an improvement in skeletal muscle function with treatment. These results warrant the further investigation of Epi as a potential therapeutic agent to mitigate MD associated muscle degeneration. PMID:25284161

Cloning and expression of SgCYP450-4 from Siraitia grosvenorii

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Dongping Tu

2016-10-01

Full Text Available CYP450 plays an essential role in the development and growth of the fruits of Siraitia grosvenorii. However, little is known about the SgCYP450-4 gene in S. grosvenorii. Here, based on transcriptome data, a full-length cDNA sequence of SgCYP450-4 was cloned by reverse transcriptase-polymerase chain reaction (RT-PCR and rapid-amplification of cDNA ends (RACE strategies. SgCYP450-4 is 1677 bp in length (GenBank accession No. AEM42985.1 and contains a complete open reading frame (ORF of 1422 bp. The deduced protein was composed of 473 amino acids, the molecular weight is 54.01 kDa, the theoretical isoelectric point (PI is 8.8, and the protein was predicted to possess cytochrome P450 domains. SgCYP450-4 gene was highly expressed in root, diploid fruit and fruit treated with hormone and pollination. At 10 days after treatment with pollination and hormones, the expression of SgCYP450-4 had the highest level and then decreased over time, which was consistent with the development of fruits of S. Grosvenorii. Hormonal treatment could significantly induce the expression of SgCYP450-4. These results provide a reference for regulation of fruit development and the use of parthenocarpy to generate seedless fruit, and provide a scientific basis for the production of growth regulator application agents.

Hepatitis C virus (HCV) induces formation of stress granules whose proteins regulate HCV RNA replication and virus assembly and egress.

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Garaigorta, Urtzi; Heim, Markus H; Boyd, Bryan; Wieland, Stefan; Chisari, Francis V

2012-10-01

Stress granules (SGs) are cytoplasmic structures that are induced in response to environmental stress, including viral infections. Here we report that hepatitis C virus (HCV) triggers the appearance of SGs in a PKR- and interferon (IFN)-dependent manner. Moreover, we show an inverse correlation between the presence of stress granules and the induction of IFN-stimulated proteins, i.e., MxA and USP18, in HCV-infected cells despite high-level expression of the corresponding MxA and USP18 mRNAs, suggesting that interferon-stimulated gene translation is inhibited in stress granule-containing HCV-infected cells. Finally, in short hairpin RNA (shRNA) knockdown experiments, we found that the stress granule proteins T-cell-restricted intracellular antigen 1 (TIA-1), TIA1-related protein (TIAR), and RasGAP-SH3 domain binding protein 1 (G3BP1) are required for efficient HCV RNA and protein accumulation at early time points in the infection and that G3BP1 and TIA-1 are required for intracellular and extracellular infectious virus production late in the infection, suggesting that they are required for virus assembly. In contrast, TIAR downregulation decreases extracellular infectious virus titers with little effect on intracellular RNA content or infectivity late in the infection, suggesting that it is required for infectious particle release. Collectively, these results illustrate that HCV exploits the stress granule machinery at least two ways: by inducing the formation of SGs by triggering PKR phosphorylation, thereby downregulating the translation of antiviral interferon-stimulated genes, and by co-opting SG proteins for its replication, assembly, and egress.

More Thoughts on AG-SG Comparisons and SG Scale Factor Determinations

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Crossley, David; Calvo, Marta; Rosat, Severine; Hinderer, Jacques

2018-03-01

We revisit a number of details that arise when doing joint AG-SG (absolute gravimeter-superconducting gravimeter) calibrations, focusing on the scale factor determination and the AG mean value that derives from the offset. When fitting SG data to AG data, the choice of which time span to use for the SG data can make a difference, as well as the inclusion of a trend that might be present in the fitting. The SG time delay has only a small effect. We review a number of options discussed recently in the literature on whether drops or sets provide the most accurate scale factor, and how to reject drops and sets to get the most consistent result. Two effects are clearly indicated by our tests, one being to smooth the raw SG 1 s (or similar sampling interval) data for times that coincide with AG drops, the other being a second pass in processing to reject residual outliers after the initial fit. Although drops can usefully provide smaller SG calibration errors compared to using set data, set values are more robust to data problems but one has to use the standard error to avoid large uncertainties. When combining scale factor determinations for the same SG at the same station, the expected gradual reduction of the error with each new experiment is consistent with the method of conflation. This is valid even when the SG data acquisition system is changed, or different AG's are used. We also find a relationship between the AG mean values obtained from SG to AG fits with the traditional short-term AG (`site') measurements usually done with shorter datasets. This involves different zero levels and corrections in the AG versus SG processing. Without using the Micro-g FG5 software it is possible to use the SG-derived corrections for tides, barometric pressure, and polar motion to convert an AG-SG calibration experiment into a site measurement (and vice versa). Finally, we provide a simple method for AG users who do not have the FG5-software to find an internal FG5 parameter that

More Thoughts on AG-SG Comparisons and SG Scale Factor Determinations

Science.gov (United States)

Crossley, David; Calvo, Marta; Rosat, Severine; Hinderer, Jacques

2018-05-01

We revisit a number of details that arise when doing joint AG-SG (absolute gravimeter-superconducting gravimeter) calibrations, focusing on the scale factor determination and the AG mean value that derives from the offset. When fitting SG data to AG data, the choice of which time span to use for the SG data can make a difference, as well as the inclusion of a trend that might be present in the fitting. The SG time delay has only a small effect. We review a number of options discussed recently in the literature on whether drops or sets provide the most accurate scale factor, and how to reject drops and sets to get the most consistent result. Two effects are clearly indicated by our tests, one being to smooth the raw SG 1 s (or similar sampling interval) data for times that coincide with AG drops, the other being a second pass in processing to reject residual outliers after the initial fit. Although drops can usefully provide smaller SG calibration errors compared to using set data, set values are more robust to data problems but one has to use the standard error to avoid large uncertainties. When combining scale factor determinations for the same SG at the same station, the expected gradual reduction of the error with each new experiment is consistent with the method of conflation. This is valid even when the SG data acquisition system is changed, or different AG's are used. We also find a relationship between the AG mean values obtained from SG to AG fits with the traditional short-term AG (`site') measurements usually done with shorter datasets. This involves different zero levels and corrections in the AG versus SG processing. Without using the Micro-g FG5 software it is possible to use the SG-derived corrections for tides, barometric pressure, and polar motion to convert an AG-SG calibration experiment into a site measurement (and vice versa). Finally, we provide a simple method for AG users who do not have the FG5-software to find an internal FG5 parameter that

Comparative transcriptome profiling of chilling stress responsiveness in grafted watermelon seedlings.

Science.gov (United States)

Xu, Jinhua; Zhang, Man; Liu, Guang; Yang, Xingping; Hou, Xilin

2016-12-01

Rootstock grafting may improve the resistance of watermelon plants to low temperatures. However, information regarding the molecular responses of rootstock grafted plants to chilling stress is limited. To elucidate the molecular mechanisms of chilling tolerance in grafted plants, the transcriptomic responses of grafted watermelon under chilling stress were analyzed using RNA-seq analysis. Sequencing data were used for digital gene expression (DGE) analysis to characterize the transcriptomic responses in grafted watermelon seedlings. A total of 702 differentially-expressed genes (DEGs) were found in rootstock grafted (RG) watermelon relative to self-grafted (SG) watermelon; among these genes, 522 genes were up-regulated and 180 were down-regulated. Additionally, 164 and 953 genes were found to specifically expressed in RG and SG seedlings under chilling stress, respectively. Functional annotations revealed that up-regulated DEGs are involved in protein processing, plant-pathogen interaction and the spliceosome, whereas down-regulated DEGs are associated with photosynthesis. Moreover, 13 DEGs were randomly selected for quantitative real time PCR (qRT-PCR) analysis. The expression profiles of these 13 DEGs were consistent with those detected by the DGE analysis, supporting the reliability of the DGE data. This work provides additional insight into the molecular basis of grafted watermelon responses to chilling stress. Copyright © 2016. Published by Elsevier Masson SAS.

Association of Oxidative Stress with Psychiatric Disorders.

Science.gov (United States)

Hassan, Waseem; Noreen, Hamsa; Castro-Gomes, Vitor; Mohammadzai, Imdadullah; da Rocha, Joao Batista Teixeira; Landeira-Fernandez, J

2016-01-01

When concentrations of both reactive oxygen species and reactive nitrogen species exceed the antioxidative capability of an organism, the cells undergo oxidative impairment. Impairments in membrane integrity and lipid and protein oxidation, protein mutilation, DNA damage, and neuronal dysfunction are some of the fundamental consequences of oxidative stress. The purpose of this work was to review the associations between oxidative stress and psychological disorders. The search terms were the following: "oxidative stress and affective disorders," "free radicals and neurodegenerative disorders," "oxidative stress and psychological disorders," "oxidative stress, free radicals, and psychiatric disorders," and "association of oxidative stress." These search terms were used in conjunction with each of the diagnostic categories of the American Psychiatric Association's Diagnostic and Statistical Manual of Mental Disorders and World Health Organization's International Statistical Classification of Diseases and Related Health Problems. Genetic, pharmacological, biochemical, and preclinical therapeutic studies, case reports, and clinical trials were selected to explore the molecular aspects of psychological disorders that are associated with oxidative stress. We identified a broad spectrum of 83 degenerative syndromes and psychiatric disorders that were associated with oxidative stress. The multi-dimensional information identified herein supports the role of oxidative stress in various psychiatric disorders. We discuss the results from the perspective of developing novel therapeutic interventions.

The stress granule protein Vgl1 and poly(A)-binding protein Pab1 are required for doxorubicin resistance in the fission yeast Schizosaccharomyces pombe

International Nuclear Information System (INIS)

Morita, Takahiro; Satoh, Ryosuke; Umeda, Nanae; Kita, Ayako; Sugiura, Reiko

2012-01-01

Highlights: ► Stress granules (SGs) as a mechanism of doxorubicin tolerance. ► We characterize the role of stress granules in doxorubicin tolerance. ► Deletion of components of SGs enhances doxorubicin sensitivity in fission yeast. ► Doxorubicin promotes SG formation when combined with heat shock. ► Doxorubicin regulates stress granule assembly independent of eIF2α phosphorylation. -- Abstract: Doxorubicin is an anthracycline antibiotic widely used for chemotherapy. Although doxorubicin is effective in the treatment of several cancers, including solid tumors and leukemias, the basis of its mechanism of action is not completely understood. Here, we describe the effects of doxorubicin and its relationship with stress granules formation in the fission yeast, Schizosaccharomyces pombe. We show that disruption of genes encoding the components of stress granules, including vgl1 + , which encodes a multi-KH type RNA-binding protein, and pab1 + , which encodes a poly(A)-binding protein, resulted in greater sensitivity to doxorubicin than seen in wild-type cells. Disruption of the vgl1 + and pab1 + genes did not confer sensitivity to other anti-cancer drugs such as cisplatin, 5-fluorouracil, and paclitaxel. We also showed that doxorubicin treatment promoted stress granule formation when combined with heat shock. Notably, doxorubicin treatment did not induce hyperphosphorylation of eIF2α, suggesting that doxorubicin is involved in stress granule assembly independent of eIF2α phosphorylation. Our results demonstrate the usefulness of fission yeast for elucidating the molecular targets of doxorubicin toxicity and suggest a novel drug-resistance mechanism involving stress granule assembly.

Mutant HFE H63D Protein Is Associated with Prolonged Endoplasmic Reticulum Stress and Increased Neuronal Vulnerability*

Science.gov (United States)

Liu, Yiting; Lee, Sang Y.; Neely, Elizabeth; Nandar, Wint; Moyo, Mthabisi; Simmons, Zachary; Connor, James R.

2011-01-01

A specific polymorphism in the hemochromatosis (HFE) gene, H63D, is over-represented in neurodegenerative disorders such as amyotrophic lateral sclerosis and Alzheimer disease. Mutations of HFE are best known as being associated with cellular iron overload, but the mechanism by which HFE H63D might increase the risk of neuron degeneration is unclear. Here, using an inducible expression cell model developed from a human neuronal cell line SH-SY5Y, we reported that the presence of the HFE H63D protein activated the unfolded protein response (UPR). This response was followed by a persistent endoplasmic reticulum (ER) stress, as the signals of UPR sensors attenuated and followed by up-regulation of caspase-3 cleavage and activity. Our in vitro findings were recapitulated in a transgenic mouse model carrying Hfe H67D, the mouse equivalent of the human H63D mutation. In this model, UPR activation was detected in the lumbar spinal cord at 6 months then declined at 12 months in association with increased caspase-3 cleavage. Moreover, upon the prolonged ER stress, the number of cells expressing HFE H63D in early apoptosis was increased moderately. Cell proliferation was decreased without increased cell death. Additionally, despite increased iron level in cells carrying HFE H63D, it appeared that ER stress was not responsive to the change of cellular iron status. Overall, our studies indicate that the HFE H63D mutant protein is associated with prolonged ER stress and chronically increased neuronal vulnerability. PMID:21349849

Spatio-temporal Dynamics and Mechanisms of Stress Granule Assembly.

Directory of Open Access Journals (Sweden)

Daisuke Ohshima

2015-06-01

Full Text Available Stress granules (SGs are non-membranous cytoplasmic aggregates of mRNAs and related proteins, assembled in response to environmental stresses such as heat shock, hypoxia, endoplasmic reticulum (ER stress, chemicals (e.g. arsenite, and viral infections. SGs are hypothesized as a loci of mRNA triage and/or maintenance of proper translation capacity ratio to the pool of mRNAs. In brain ischemia, hippocampal CA3 neurons, which are resilient to ischemia, assemble SGs. In contrast, CA1 neurons, which are vulnerable to ischemia, do not assemble SGs. These results suggest a critical role SG plays in regards to cell fate decisions. Thus SG assembly along with its dynamics should determine the cell fate. However, the process that exactly determines the SG assembly dynamics is largely unknown. In this paper, analyses of experimental data and computer simulations were used to approach this problem. SGs were assembled as a result of applying arsenite to HeLa cells. The number of SGs increased after a short latent period, reached a maximum, then decreased during the application of arsenite. At the same time, the size of SGs grew larger and became localized at the perinuclear region. A minimal mathematical model was constructed, and stochastic simulations were run to test the modeling. Since SGs are discrete entities as there are only several tens of them in a cell, commonly used deterministic simulations could not be employed. The stochastic simulations replicated observed dynamics of SG assembly. In addition, these stochastic simulations predicted a gamma distribution relative to the size of SGs. This same distribution was also found in our experimental data suggesting the existence of multiple fusion steps in the SG assembly. Furthermore, we found that the initial steps in the SG assembly process and microtubules were critical to the dynamics. Thus our experiments and stochastic simulations presented a possible mechanism regulating SG assembly.

Interactions of early adversity with stress-related gene polymorphisms impact regional brain structure in females.

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Gupta, Arpana; Labus, Jennifer; Kilpatrick, Lisa A; Bonyadi, Mariam; Ashe-McNalley, Cody; Heendeniya, Nuwanthi; Bradesi, Sylvie; Chang, Lin; Mayer, Emeran A

2016-04-01

Early adverse life events (EALs) have been associated with regional thinning of the subgenual cingulate cortex (sgACC), a brain region implicated in the development of disorders of mood and affect, and often comorbid functional pain disorders, such as irritable bowel syndrome (IBS). Regional neuroinflammation related to chronic stress system activation has been suggested as a possible mechanism underlying these neuroplastic changes. However, the interaction of genetic and environmental factors in these changes is poorly understood. The current study aimed to evaluate the interactions of EALs and candidate gene polymorphisms in influencing thickness of the sgACC. 210 female subjects (137 healthy controls; 73 IBS) were genotyped for stress and inflammation-related gene polymorphisms. Genetic variation with EALs, and diagnosis on sgACC thickness was examined, while controlling for race, age, and total brain volume. Compared to HCs, IBS had significantly reduced sgACC thickness (p = 0.03). Regardless of disease group (IBS vs. HC), thinning of the left sgACC was associated with a significant gene-gene environment interaction between the IL-1β genotype, the NR3C1 haplotype, and a history of EALs (p = 0.05). Reduced sgACC thickness in women with the minor IL-1β allele, was associated with EAL total scores regardless of NR3C1 haplotype status (p = 0.02). In subjects homozygous for the major IL-1β allele, reduced sgACC with increasing levels of EALs was seen only with the less common NR3C1 haplotype (p = 0.02). These findings support an interaction between polymorphisms related to stress and inflammation and early adverse life events in modulating a key region of the emotion arousal circuit.

Mindfulness meditation training alters stress-related amygdala resting state functional connectivity: a randomized controlled trial.

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Taren, Adrienne A; Gianaros, Peter J; Greco, Carol M; Lindsay, Emily K; Fairgrieve, April; Brown, Kirk Warren; Rosen, Rhonda K; Ferris, Jennifer L; Julson, Erica; Marsland, Anna L; Bursley, James K; Ramsburg, Jared; Creswell, J David

2015-12-01

Recent studies indicate that mindfulness meditation training interventions reduce stress and improve stress-related health outcomes, but the neural pathways for these effects are unknown. The present research evaluates whether mindfulness meditation training alters resting state functional connectivity (rsFC) of the amygdala, a region known to coordinate stress processing and physiological stress responses. We show in an initial discovery study that higher perceived stress over the past month is associated with greater bilateral amygdala-subgenual anterior cingulate cortex (sgACC) rsFC in a sample of community adults (n = 130). A follow-up, single-blind randomized controlled trial shows that a 3-day intensive mindfulness meditation training intervention (relative to a well-matched 3-day relaxation training intervention without a mindfulness component) reduced right amygdala-sgACC rsFC in a sample of stressed unemployed community adults (n = 35). Although stress may increase amygdala-sgACC rsFC, brief training in mindfulness meditation could reverse these effects. This work provides an initial indication that mindfulness meditation training promotes functional neuroplastic changes, suggesting an amygdala-sgACC pathway for stress reduction effects. © The Author (2015). Published by Oxford University Press. For Permissions, please email: journals.permissions@oup.com.

Protective effects of transforming growth factor β2 in intestinal epithelial cells by regulation of proteins associated with stress and endotoxin responses

DEFF Research Database (Denmark)

Nguyen, Duc Ninh; Jiang, Pingping; Jacobsen, Susanne

2015-01-01

electrophoresis-MS and LC-MS-based proteomics. TGF-β2 alone induced the differential expression of 13 proteins and the majority of the identified proteins were associated with stress responses, TGF-β and Toll-like receptor 4 signaling cascades. In particular, a series of heat shock proteins had similar...... with stress response pathways, among which five proteins were altered by LPS and restored by TGF-β2, whereas six were differentially expressed only by TGF-β2 in LPS-challenged IECs. Based on previously reported biological functions, these patterns indicate the anti-stress and anti-inflammatory effects of TGF...

The stress granule protein Vgl1 and poly(A)-binding protein Pab1 are required for doxorubicin resistance in the fission yeast Schizosaccharomyces pombe

Energy Technology Data Exchange (ETDEWEB)

Morita, Takahiro [Laboratory of Molecular Pharmacogenomics, School of Pharmaceutical Sciences, Kinki University, Kowakae 3-4-1, Higashi-Osaka 577-8502 (Japan); Satoh, Ryosuke [Laboratory of Molecular Pharmacogenomics, School of Pharmaceutical Sciences, Kinki University, Kowakae 3-4-1, Higashi-Osaka 577-8502 (Japan); Japan Society for the Promotion of Science, 1-8 Chiyoda-ku, Tokyo 102-8472 (Japan); Umeda, Nanae; Kita, Ayako [Laboratory of Molecular Pharmacogenomics, School of Pharmaceutical Sciences, Kinki University, Kowakae 3-4-1, Higashi-Osaka 577-8502 (Japan); Sugiura, Reiko, E-mail: sugiurar@phar.kindai.ac.jp [Laboratory of Molecular Pharmacogenomics, School of Pharmaceutical Sciences, Kinki University, Kowakae 3-4-1, Higashi-Osaka 577-8502 (Japan)

2012-01-06

Highlights: Black-Right-Pointing-Pointer Stress granules (SGs) as a mechanism of doxorubicin tolerance. Black-Right-Pointing-Pointer We characterize the role of stress granules in doxorubicin tolerance. Black-Right-Pointing-Pointer Deletion of components of SGs enhances doxorubicin sensitivity in fission yeast. Black-Right-Pointing-Pointer Doxorubicin promotes SG formation when combined with heat shock. Black-Right-Pointing-Pointer Doxorubicin regulates stress granule assembly independent of eIF2{alpha} phosphorylation. -- Abstract: Doxorubicin is an anthracycline antibiotic widely used for chemotherapy. Although doxorubicin is effective in the treatment of several cancers, including solid tumors and leukemias, the basis of its mechanism of action is not completely understood. Here, we describe the effects of doxorubicin and its relationship with stress granules formation in the fission yeast, Schizosaccharomyces pombe. We show that disruption of genes encoding the components of stress granules, including vgl1{sup +}, which encodes a multi-KH type RNA-binding protein, and pab1{sup +}, which encodes a poly(A)-binding protein, resulted in greater sensitivity to doxorubicin than seen in wild-type cells. Disruption of the vgl1{sup +} and pab1{sup +} genes did not confer sensitivity to other anti-cancer drugs such as cisplatin, 5-fluorouracil, and paclitaxel. We also showed that doxorubicin treatment promoted stress granule formation when combined with heat shock. Notably, doxorubicin treatment did not induce hyperphosphorylation of eIF2{alpha}, suggesting that doxorubicin is involved in stress granule assembly independent of eIF2{alpha} phosphorylation. Our results demonstrate the usefulness of fission yeast for elucidating the molecular targets of doxorubicin toxicity and suggest a novel drug-resistance mechanism involving stress granule assembly.

Genome-wide identification and characterization of stress-associated protein (SAP gene family encoding A20/AN1 zinc-finger proteins in Medicago truncatula

Directory of Open Access Journals (Sweden)

Zhou Yong

2018-01-01

Full Text Available Stress associated proteins (SAPs play important roles in developmental processes, responses to various stresses and hormone stimulation in plants. However, little is known about the SAP gene family in Medicago truncatula. In this study, a total of 17 MtSAP genes encoding A20/AN1 zinc-finger proteins were characterized. Out of these 17 genes, 15 were distributed over all 8 chromosomes at different densities, and two segmental duplication events were detected. The phylogenetic analysis of these proteins and their orthologs from Arabidopsis and rice suggested that they could be classified into five out of the seven groups of SAP family genes, with genes in the same group showing similar structures and conserved domains. The cis-elements of the MtSAP promoters were studied, and many cis-elements related to stress and plant hormone responses were identified. We also investigated the stress-responsive expression patterns of the MtSAP genes under various stresses, including drought, exposure to NaCl and cold. The qRT-PCR results showed that numerous MtSAP genes exhibited transcriptional responses to multiple abiotic stresses. These results lay the foundation for further functional characterization of SAP genes. To the best of our knowledge, this is the first report of a genome-wide analysis of the SAP gene family in M. truncatula.

STAC -- a new Swedish code for statistical analysis of cracks in SG-tubes

International Nuclear Information System (INIS)

Poern, K.

1997-01-01

Steam generator (SG) tubes in pressurized water reactor plants are exposed to various types of degradation processes, among which stress corrosion cracking in particular has been observed. To be able to evaluate the safety importance of such cracking of SG-tubes one has to have a good and empirically founded knowledge about the scope and the size of the cracks as well as the rate of their continuous growth. The basis of experience is to a large extent constituted of the annually performed SG-inspections and crack sizing procedures. On the basis of this experience one can estimate the distribution of existing crack lengths, and modify this distribution with regard to maintenance (plugging) and the predicted rate of crack propagation. Finally, one can calculate the rupture probability of SG-tubes as a function of a given critical crack length. On account of the Swedish Nuclear Power Inspectorate an introductory study has been performed in order to get a survey of what has been done elsewhere in this field. The study resulted in a proposal of a computerizable model to be able to estimate the distribution of true cracks, to modify this distribution due to the crack growth and to compute the probability of tube rupture. The model has now been implemented in a compute code, called STAC (STatistical Analysis of Cracks). This paper is aimed to give a brief outline of the model to facilitate the understanding of the possibilities and limitations associated with the model

Lipopolysaccharide-binding protein and leptin are associated with stress-induced interleukin-6 cytokine expression ex vivo in obesity.

Science.gov (United States)

Huang, Chun-Jung; Stewart, Jennifer K; Shibata, Yoshimi; Slusher, Aaron L; Acevedo, Edmund O

2015-05-01

Obesity is associated with enhanced inflammation and mental stress, but limited information has addressed the potential additive effect of psychological stress on obesity-associated inflammation. This study examined whether obese subjects would elicit a greater host immune response (IL-6 mRNA and cytokine) to lipopolysaccharide (LPS) in response to mental stress. Blood samples for LPS-stimulated IL-6 mRNA and cytokine were collected prior to and following mental stress. Results showed that obese subjects elicited a greater LPS-induced IL-6 along with its mRNA expression following mental stress compared to normal-weight subjects. Stress-induced IL-6 cytokine response to LPS was correlated with the baseline levels of plasma LPS binding protein (LBP) and leptin. These findings are consistent with the idea that endogenous inflammatory agents (e.g., LBP and leptin), often elevated with obesity, enhance inflammatory responses to psychological stress. © 2014 Society for Psychophysiological Research.

Translational control of aberrant stress responses as a hallmark of cancer.

Science.gov (United States)

El-Naggar, Amal M; Sorensen, Poul H

2018-04-01

Altered mRNA translational control is emerging as a critical factor in cancer development and progression. Targeting specific elements of the translational machinery, such as mTORC1 or eIF4E, is emerging as a new strategy for innovative cancer therapy. While translation of most mRNAs takes place through cap-dependent mechanisms, a sub-population of cellular mRNA species, particularly stress-inducible mRNAs with highly structured 5'-UTR regions, are primarily translated through cap-independent mechanisms. Intriguingly, many of these mRNAs encode proteins that are involved in tumour cell adaptation to microenvironmental stress, and thus linked to aggressive behaviour including tumour invasion and metastasis. This necessitates a rigorous search for links between microenvironmental stress and aggressive tumour phenotypes. Under stress, cells block global protein synthesis to preserve energy while maintaining selective synthesis of proteins that support cell survival. One highly conserved mechanism to regulate protein synthesis under cell stress is to sequester mRNAs into cytosolic aggregates called stress granules (SGs), where their translation is silenced. SGs confer survival advantages and chemotherapeutic resistance to tumour cells under stress. Recently, it has been shown that genetically blocking SG formation dramatically reduces tumour invasive and metastatic capacity in vivo. Therefore, targeting SG formation might represent a potential treatment strategy to block cancer metastasis. Here, we present the critical link between selective mRNA translation, stress adaptation, SGs, and tumour progression. Further, we also explain how deciphering mechanisms of selective mRNA translation occurs under cell stress holds great promise for the identification of new targets in the treatment of cancer. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2018 Pathological Society of Great Britain and Ireland

Learnings from investigations on SG divider plates: Coupling field characterizations with numerical mechanical simulation

International Nuclear Information System (INIS)

Rossillon, F.; Depradeux, L.; Miloudi, S.; Deforge, D.; Lemaire, E.; Massoud, J.P.

2014-01-01

Nickel based alloys stress corrosion cracking (SCC) has been a major concern for the nuclear power plant utilities since the 1970s. Since 2002, SCC indications have been found on steam generator (SG) divider plates made of alloy 600 on French PWRs (pressurized water reactors) 900 MWe units. Although integrity is not questioned, many studies have been conducted to deepen understanding of the phenomenon. Among numerous studies to investigate the SCC damage phenomena, advanced mechanical analysis has been performed to improve the knowledge of the in-service loadings of the SG 900 MWe partition stub and divider plate. Manufacturing steps are taken into account, such as welding and the first hydro-test, to have a more precise description of the mechanical states in the vicinity of the welds where SCC is likely to occur. Recently, EDF hot laboratories made destructive examinations of a decommissioned SG. To fulfil the analyses computations have been carried out on the dedicated configuration. A 3D FE model, including the simulation of the welding and hydro-test, has been set up. Comparisons with experimental investigations on the divider plate of decommissioned SG have shown a good agreement between experimental and numerical results. These results emphasize the redistribution of weld residual stresses after the first hydro-test, and the effect of hydro-testing on the plastic deformation of the stub only in some specific cases of 900 MWe SG

Learnings from investigations on SG divider plates: Coupling field characterizations with numerical mechanical simulation

Energy Technology Data Exchange (ETDEWEB)

Rossillon, F., E-mail: frederique.rossillon@edf.fr [EDF SEPTEN, 12-14 Avenue Dutrievoz, Villeurbanne (France); Depradeux, L. [EC2-MS, 66 Bd Niels Bohr, Villeurbanne (France); Miloudi, S. [EDF CEIDRE, CNPE de Chinon, Avoine (France); Deforge, D. [EDF CEIDRE, 2 Rue Ampère, Saint Denis (France); Lemaire, E. [EDF UNIE, Cap Ampère, Saint Denis (France); Massoud, J.P. [EDF SEPTEN, 12-14 Avenue Dutrievoz, Villeurbanne (France)

2014-04-01

Nickel based alloys stress corrosion cracking (SCC) has been a major concern for the nuclear power plant utilities since the 1970s. Since 2002, SCC indications have been found on steam generator (SG) divider plates made of alloy 600 on French PWRs (pressurized water reactors) 900 MWe units. Although integrity is not questioned, many studies have been conducted to deepen understanding of the phenomenon. Among numerous studies to investigate the SCC damage phenomena, advanced mechanical analysis has been performed to improve the knowledge of the in-service loadings of the SG 900 MWe partition stub and divider plate. Manufacturing steps are taken into account, such as welding and the first hydro-test, to have a more precise description of the mechanical states in the vicinity of the welds where SCC is likely to occur. Recently, EDF hot laboratories made destructive examinations of a decommissioned SG. To fulfil the analyses computations have been carried out on the dedicated configuration. A 3D FE model, including the simulation of the welding and hydro-test, has been set up. Comparisons with experimental investigations on the divider plate of decommissioned SG have shown a good agreement between experimental and numerical results. These results emphasize the redistribution of weld residual stresses after the first hydro-test, and the effect of hydro-testing on the plastic deformation of the stub only in some specific cases of 900 MWe SG.

Active Intracellular Delivery of a Cas9/sgRNA Complex Using Ultrasound-Propelled Nanomotors.

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Hansen-Bruhn, Malthe; de Ávila, Berta Esteban-Fernández; Beltrán-Gastélum, Mara; Zhao, Jing; Ramírez-Herrera, Doris E; Angsantikul, Pavimol; Vesterager Gothelf, Kurt; Zhang, Liangfang; Wang, Joseph

2018-03-01

Direct and rapid intracellular delivery of a functional Cas9/sgRNA complex using ultrasound-powered nanomotors is reported. The Cas9/sgRNA complex is loaded onto the nanomotor surface through a reversible disulfide linkage. A 5 min ultrasound treatment enables the Cas9/sgRNA-loaded nanomotors to directly penetrate through the plasma membrane of GFP-expressing B16F10 cells. The Cas9/sgRNA is released inside the cells to achieve highly effective GFP gene knockout. The acoustic Cas9/sgRNA-loaded nanomotors display more than 80 % GFP knockout within 2 h of cell incubation compared to 30 % knockout using static nanowires. More impressively, the nanomotors enable highly efficient knockout with just 0.6 nm of the Cas9/sgRNA complex. This nanomotor-based intracellular delivery method thus offers an attractive route to overcome physiological barriers for intracellular delivery of functional proteins and RNAs, thus indicating considerable promise for highly efficient therapeutic applications. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Stress proteins, autoimmunity, and autoimmune disease.

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Winfield, J B; Jarjour, W N

1991-01-01

At birth, the immune system is biased toward recognition of microbial antigens in order to protect the host from infection. Recent data suggest that an important initial line of defense in this regard involves autologous stress proteins, especially conserved peptides of hsp60, which are presented to T cells bearing gamma delta receptors by relatively nonpolymorphic class lb molecules. Natural antibodies may represent a parallel B cell mechanism. Through an evolving process of "physiological" autoreactivity and selection by immunodominant stress proteins common to all prokaryotes, B and T cell repertoires expand during life to meet the continuing challenge of infection. Because stress proteins of bacteria are homologous with stress proteins of the host, there exists in genetically susceptible individuals a constant risk of autoimmune disease due to failure of mechanisms for self-nonself discrimination. That stress proteins actually play a role in autoimmune processes is supported by a growing body of evidence which, collectively, suggests that autoreactivity in chronic inflammatory arthritis involves, at least initially, gamma delta cells which recognize epitopes of the stress protein hsp60. Alternate mechanisms for T cell stimulation by stress proteins undoubtedly also exist, e.g., molecular mimicry of the DR beta third hypervariable region susceptibility locus for rheumatoid arthritis by a DnaJ stress protein epitope in gram-negative bacteria. While there still is confusion with respect to the most relevant stress protein epitopes, a central role for stress proteins in the etiology of arthritis appears likely. Furthermore, insight derived from the work thus far in adjuvant-induced arthritis already is stimulating analyses of related phenomena in autoimmune diseases other than those involving joints. Only limited data are available in the area of humoral autoimmunity to stress proteins. Autoantibodies to a number of stress proteins have been identified in SLE and

Ebola virus VP35 blocks stress granule assembly.

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Le Sage, Valerie; Cinti, Alessandro; McCarthy, Stephen; Amorim, Raquel; Rao, Shringar; Daino, Gian Luca; Tramontano, Enzo; Branch, Donald R; Mouland, Andrew J

2017-02-01

Stress granules (SGs) are dynamic cytoplasmic aggregates of translationally silenced mRNAs that assemble in response to environmental stress. SGs appear to play an important role in antiviral innate immunity and many viruses have evolved to block or subvert SGs components for their own benefit. Here, we demonstrate that intracellular Ebola virus (EBOV) replication and transcription-competent virus like particles (trVLP) infection does not lead to SG assembly but leads to a blockade to Arsenite-induced SG assembly. Moreover we show that EBOV VP35 represses the assembly of canonical and non-canonical SGs induced by a variety of pharmacological stresses. This SG blockade requires, at least in part, the C-terminal domain of VP35. Furthermore, results from our co-immunoprecipitation studies indicate that VP35 interacts with multiple SG components, including G3BP1, eIF3 and eEF2 through a stress- and RNA-independent mechanism. These data suggest a novel function for EBOV VP35 in the repression of SG assembly. Copyright © 2016 Elsevier Inc. All rights reserved.

Study of the experimental parameters associated to the determination of residual macro stresses in stainless steel tubes, through x-ray diffraction method

International Nuclear Information System (INIS)

Guimaraes, L.R.

1990-01-01

The basic principles related to the determination of residual macro stresses by X-rays diffractometry are present, whereas different techniques associated with the respective experimental errors are discussed. The residual stresses in two 304 L stainless steel tubes were measured using three models of diffractometers, Rigaku SG-8, Jeol JDX-11PA and Rigaku Strainflex. The measured values of stresses as well as the reproducibilities are examined. The suitability of peak location method, by fitting three data points to the parabolic function, is discussed through values of position and intensity obtained by two of the above diffractometers. (author)

Study of the experimental parameters associated to the determination of residual macro stresses in stainless steel tubes through x-rays diffraction method

International Nuclear Information System (INIS)

Guimaraes, L.R.

1990-01-01

The basic principles related to the determination of residual macro stresses by X-rays diffractometry are present, whereas different techniques associated with the respective experimental errors are discussed. The residual stresses in two 304 L stainless steel tubes were measured using three models of diffractometers, Rigaku SG-8, Jeol JDX-11PA and Rigaku Strainflex. The measured values of stresses as well as the reproducibilities are examined. The suitability of peak location method, by fitting three data points to the parabolic function, is discussed through values of position and intensity obtained by two of the above diffractometers. (author)

ANGUSTIFOLIA, a Plant Homolog of CtBP/BARS Localizes to Stress Granules and Regulates Their Formation

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Hemal Bhasin

2017-06-01

Full Text Available The ANGUSTIFOLIA (AN gene in Arabidopsis is important for a plethora of morphological phenotypes. Recently, AN was also reported to be involved in responses to biotic and abiotic stresses. It encodes a homolog of the animal C-terminal binding proteins (CtBPs. In contrast to animal CtBPs, AN does not appear to function as a transcriptional co-repressor and instead functions outside nucleus where it might be involved in Golgi-associated membrane trafficking. In this study, we report a novel and unexplored role of AN as a component of stress granules (SGs. Interaction studies identified several RNA binding proteins that are associated with AN. AN co-localizes with several messenger ribonucleoprotein granule markers to SGs in a stress dependent manner. an mutants exhibit an altered SG formation. We provide evidence that the NAD(H binding domain of AN is relevant in this context as proteins carrying mutations in this domain localize to a much higher degree to SGs and strongly reduce AN dimerization and its interaction with one interactor but not the others. Finally, we show that AN is a negative regulator of salt and osmotic stress responses in Arabidopsis suggesting a functional relevance in SGs.

AAV exploits subcellular stress associated with inflammation, endoplasmic reticulum expansion, and misfolded proteins in models of cystic fibrosis.

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Jarrod S Johnson

2011-05-01

Full Text Available Barriers to infection act at multiple levels to prevent viruses, bacteria, and parasites from commandeering host cells for their own purposes. An intriguing hypothesis is that if a cell experiences stress, such as that elicited by inflammation, endoplasmic reticulum (ER expansion, or misfolded proteins, then subcellular barriers will be less effective at preventing viral infection. Here we have used models of cystic fibrosis (CF to test whether subcellular stress increases susceptibility to adeno-associated virus (AAV infection. In human airway epithelium cultured at an air/liquid interface, physiological conditions of subcellular stress and ER expansion were mimicked using supernatant from mucopurulent material derived from CF lungs. Using this inflammatory stimulus to recapitulate stress found in diseased airways, we demonstrated that AAV infection was significantly enhanced. Since over 90% of CF cases are associated with a misfolded variant of Cystic Fibrosis Transmembrane Conductance Regulator (ΔF508-CFTR, we then explored whether the presence of misfolded proteins could independently increase susceptibility to AAV infection. In these models, AAV was an order of magnitude more efficient at transducing cells expressing ΔF508-CFTR than in cells expressing wild-type CFTR. Rescue of misfolded ΔF508-CFTR under low temperature conditions restored viral transduction efficiency to that demonstrated in controls, suggesting effects related to protein misfolding were responsible for increasing susceptibility to infection. By testing other CFTR mutants, G551D, D572N, and 1410X, we have shown this phenomenon is common to other misfolded proteins and not related to loss of CFTR activity. The presence of misfolded proteins did not affect cell surface attachment of virus or influence expression levels from promoter transgene cassettes in plasmid transfection studies, indicating exploitation occurs at the level of virion trafficking or processing. Thus

Age-Related Decrease in Heat Shock 70-kDa Protein 8 in Cerebrospinal Fluid Is Associated with Increased Oxidative Stress.

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Loeffler, David A; Klaver, Andrea C; Coffey, Mary P; Aasly, Jan O; LeWitt, Peter A

2016-01-01

Age-associated declines in protein homeostasis mechanisms ("proteostasis") are thought to contribute to age-related neurodegenerative disorders. The increased oxidative stress which occurs with aging can activate a key proteostatic process, chaperone-mediated autophagy. This study investigated age-related alteration in cerebrospinal fluid (CSF) concentrations of heat shock 70-kDa protein 8 (HSPA8), a molecular chaperone involved in proteostatic mechanisms including chaperone-mediated autophagy, and its associations with indicators of oxidative stress (8-hydroxy-2'-deoxyguanosine [8-OHdG] and 8-isoprostane) and total anti-oxidant capacity. We examined correlations between age, HSPA8, 8-OHdG, 8-isoprostane, and total antioxidant capacity (TAC) in CSF samples from 34 healthy subjects ranging from 20 to 75 years of age. Age was negatively associated with HSPA8 (ρ = -0.47; p = 0.005). An age-related increase in oxidative stress was indicated by a positive association between age and 8-OHdG (ρ = 0.61; p = 0.0001). HSPA8 was moderately negatively associated with 8-OHdG (ρ = -0.58; p = 0.0004). Age and HSPA8 were weakly associated with 8-isoprostane and TAC (range of ρ values: -0.15 to 0.16). Our findings in this exploratory study suggest that during healthy aging, CSF HSPA8 may decrease, perhaps due in part to an increase in oxidative stress. Our results also suggest that 8-OHdG may be more sensitive than 8-isoprostane for measuring oxidative stress in CSF. Further studies are indicated to determine if our findings can be replicated with a larger cohort, and if the age-related decrease in HSPA8 in CSF is reflected by a similar change in the brain.

Stress proteins and the immune response.

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Moseley, P

2000-07-25

The heat shock or stress response is one of the most highly conserved adaptive responses in nature. In single cell organisms, the stress response confers tolerance to a variety of stresses including hyperthermia, hyperoxia, hypoxia, and other perturbations, which alter protein synthesis. This tolerance phenomenon is also extremely important in the multicellular organism, resulting in not only thermal tolerance, but also resistance to stresses of the whole organism such as ischemia-reperfusion injury. Moreover, recent data indicates that these stress proteins have the ability to modulate the cellular immune response. Although the terms heat shock proteins (HSPs) and stress proteins are often used interchangeably, the term stress proteins includes the HSPs, the glucose-regulated proteins (GRPs) and ubiquitin. The stress proteins may be grouped by molecular weight ranging from the large 110 kDa HSP110 to ubiquitin at 8 kDa. These proteins serve as cellular chaperones, participating in protein synthesis and transport through the various cellular compartments. Because these proteins have unique cellular localizations, the chaperone function of the stress proteins often involves a transfer of peptides between stress proteins as the peptide is moved between cellular compartments. For example, HSP70 is a cytosolic and nuclear chaperone, which is critical for the transfer of cellular peptides in the mitochondrion through a hand-off that involves mitochondrial HSP60 at the inner mitochondrial membrane. Similarly, cytosolic proteins are transferred from HSP70 to gp96 as they move into the endoplasmic reticulum. The central role of the stress proteins in the transfer of peptides through the cell may be responsible for the recently recognized importance of the stress proteins in the modulation of the immune system [Feder, M.E., Hofmann, G.E., 1999. Heat-shock proteins, molecular chaperones, and the stress response: evolutionary and ecological physiology. Annu. Rev. Physiol. 61

Antagonistic action of Bacillus subtilis strain SG6 on Fusarium graminearum.

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Zhao, Yueju; Selvaraj, Jonathan Nimal; Xing, Fuguo; Zhou, Lu; Wang, Yan; Song, Huimin; Tan, Xinxin; Sun, Lichao; Sangare, Lancine; Folly, Yawa Minnie Elodie; Liu, Yang

2014-01-01

Fusarium graminearum causes Fusarium head blight (FHB), a devastating disease that leads to extensive yield and quality loss of wheat and barley. Bacteria isolated from wheat kernels and plant anthers were screened for antagonistic activity against F. graminearum. Based on its in vitro effectiveness, strain SG6 was selected for characterization and identified as Bacillus subtilis. B. subtilis SG6 exhibited a high antifungal effect on the mycelium growth, sporulation and DON production of F. graminearum with the inhibition rate of 87.9%, 95.6% and 100%, respectively. In order to gain insight into biological control effect in situ, we applied B. subtilis SG6 at anthesis through the soft dough stage of kernel development in field test. It was revealed that B. subtilis SG6 significantly reduced disease incidence (DI), FHB index and DON (P ≤ 0.05). Further, ultrastructural examination shows that B. subtilis SG6 strain induced stripping of F. graminearum hyphal surface by destroying the cellular structure. When hypha cell wall was damaged, the organelles and cytoplasm inside cell would exude, leading to cell death. The antifungal activity of SG6 could be associated with the coproduction of chitinase, fengycins and surfactins.

Antagonistic action of Bacillus subtilis strain SG6 on Fusarium graminearum.

Directory of Open Access Journals (Sweden)

Yueju Zhao

Full Text Available Fusarium graminearum causes Fusarium head blight (FHB, a devastating disease that leads to extensive yield and quality loss of wheat and barley. Bacteria isolated from wheat kernels and plant anthers were screened for antagonistic activity against F. graminearum. Based on its in vitro effectiveness, strain SG6 was selected for characterization and identified as Bacillus subtilis. B. subtilis SG6 exhibited a high antifungal effect on the mycelium growth, sporulation and DON production of F. graminearum with the inhibition rate of 87.9%, 95.6% and 100%, respectively. In order to gain insight into biological control effect in situ, we applied B. subtilis SG6 at anthesis through the soft dough stage of kernel development in field test. It was revealed that B. subtilis SG6 significantly reduced disease incidence (DI, FHB index and DON (P ≤ 0.05. Further, ultrastructural examination shows that B. subtilis SG6 strain induced stripping of F. graminearum hyphal surface by destroying the cellular structure. When hypha cell wall was damaged, the organelles and cytoplasm inside cell would exude, leading to cell death. The antifungal activity of SG6 could be associated with the coproduction of chitinase, fengycins and surfactins.

Extracellular cell stress (heat shock) proteins-immune responses and disease: an overview.

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Pockley, A Graham; Henderson, Brian

2018-01-19

Extracellular cell stress proteins are highly conserved phylogenetically and have been shown to act as powerful signalling agonists and receptors for selected ligands in several different settings. They also act as immunostimulatory 'danger signals' for the innate and adaptive immune systems. Other studies have shown that cell stress proteins and the induction of immune reactivity to self-cell stress proteins can attenuate disease processes. Some proteins (e.g. Hsp60, Hsp70, gp96) exhibit both inflammatory and anti-inflammatory properties, depending on the context in which they encounter responding immune cells. The burgeoning literature reporting the presence of stress proteins in a range of biological fluids in healthy individuals/non-diseased settings, the association of extracellular stress protein levels with a plethora of clinical and pathological conditions and the selective expression of a membrane form of Hsp70 on cancer cells now supports the concept that extracellular cell stress proteins are involved in maintaining/regulating organismal homeostasis and in disease processes and phenotype. Cell stress proteins, therefore, form a biologically complex extracellular cell stress protein network having diverse biological, homeostatic and immunomodulatory properties, the understanding of which offers exciting opportunities for delivering novel approaches to predict, identify, diagnose, manage and treat disease.This article is part of the theme issue 'Heat shock proteins as modulators and therapeutic targets of chronic disease: an integrated perspective'. © 2017 The Author(s).

Dynamic Interaction of Stress Granules, DDX3X, and IKK-α Mediates Multiple Functions in Hepatitis C Virus Infection.

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Pène, Véronique; Li, Qisheng; Sodroski, Catherine; Hsu, Ching-Sheng; Liang, T Jake

2015-05-01

The ubiquitous ATP-dependent RNA helicase DDX3X is involved in many cellular functions, including innate immunity, and is a pivotal host factor for hepatitis C virus (HCV) infection. Recently, we showed that DDX3X specifically recognizes the HCV 3' untranslated region (UTR), leading to the activation of IKK-α and a cascade of lipogenic signaling to facilitate lipid droplet biogenesis and viral assembly (Q. Li, V. Pene, S. Krishnamurthy, H. Cha, and T. J. Liang, Nat Med 19:722-729, 2013, http://dx.doi.org/10.1038/nm.3190). The interaction of DDX3X with HCV core protein seems to be dispensable for its proviral role. In this study, through systematic imaging and biochemical and virologic approaches, we identified a dynamic association between DDX3X and various cellular compartments and viral elements mediating multiple functions of DDX3X in productive HCV infection. Upon HCV infection, the HCV 3'UTR interacts with DDX3X and IKK-α, which redistribute to speckle-like cytoplasmic structures shown to be stress granules (SGs). As viral proteins accumulate in infected cells, DDX3X granules together with SG-associated proteins redistribute and colocalize with HCV core protein around lipid droplets (LDs). IKK-α, however, does not relocate to the LD but translocates to the nucleus. In HCV-infected cells, various HCV nonstructural proteins also interact or colocalize with DDX3X in close proximity to SGs and LDs, consistent with the tight juxtaposition of the replication complex and the assembly site at the surface of LDs. Short interfering RNA (siRNA)-mediated silencing of DDX3X and multiple SG components markedly inhibits HCV infection. Our data suggest that DDX3X initiates a multifaceted cellular program involving dynamic associations with HCV RNA and proteins, IKK-α, SG, and LD surfaces for its crucial role in the HCV life cycle. IMPORTANCE DDX3X is a proviral host factor for HCV infection. Recently, we showed that DDX3X binds to the HCV 3'UTR, activating IKK-α and

[In silico CRISPR-based sgRNA design].

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Wang, Yuanli; Chuai, Guohui; Yan, Jifang; Shi, Lei; Liu, Qi

2017-10-25

CRISPR-based genome editing has been widely implemented in various cell types. In-silico single guide RNA (sgRNA) design is a key step for successful gene editing using CRISPR system. Continuing efforts are made to refine in-silico sgRNA design with high on-target efficacy and reduced off-target effects. In this paper, we summarize the present sgRNA design tools, and show that efficient in-silico models can be built that integrate current heterogeneous genome-editing data to derive unbiased sgRNA design rules and identify key features for improving sgRNA design. Our review shows that systematic comparisons and evaluation of on-target and off-target effects of sgRNA will allow more precise genome editing and gene therapies using the CRISPR system.

Hypothesis: NDL proteins function in stress responses by regulating microtubule organization.

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Khatri, Nisha; Mudgil, Yashwanti

2015-01-01

N-MYC DOWNREGULATED-LIKE proteins (NDL), members of the alpha/beta hydrolase superfamily were recently rediscovered as interactors of G-protein signaling in Arabidopsis thaliana. Although the precise molecular function of NDL proteins is still elusive, in animals these proteins play protective role in hypoxia and expression is induced by hypoxia and nickel, indicating role in stress. Homology of NDL1 with animal counterpart N-MYC DOWNREGULATED GENE (NDRG) suggests similar functions in animals and plants. It is well established that stress responses leads to the microtubule depolymerization and reorganization which is crucial for stress tolerance. NDRG is a microtubule-associated protein which mediates the microtubule organization in animals by causing acetylation and increases the stability of α-tubulin. As NDL1 is highly homologous to NDRG, involvement of NDL1 in the microtubule organization during plant stress can also be expected. Discovery of interaction of NDL with protein kinesin light chain- related 1, enodomembrane family protein 70, syntaxin-23, tubulin alpha-2 chain, as a part of G protein interactome initiative encourages us to postulate microtubule stabilizing functions for NDL family in plants. Our search for NDL interactors in G protein interactome also predicts the role of NDL proteins in abiotic stress tolerance management. Based on published report in animals and predicted interacting partners for NDL in G protein interactome lead us to hypothesize involvement of NDL in the microtubule organization during abiotic stress management in plants.

Quantitative protein localization signatures reveal an association between spatial and functional divergences of proteins.

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Loo, Lit-Hsin; Laksameethanasan, Danai; Tung, Yi-Ling

2014-03-01

Protein subcellular localization is a major determinant of protein function. However, this important protein feature is often described in terms of discrete and qualitative categories of subcellular compartments, and therefore it has limited applications in quantitative protein function analyses. Here, we present Protein Localization Analysis and Search Tools (PLAST), an automated analysis framework for constructing and comparing quantitative signatures of protein subcellular localization patterns based on microscopy images. PLAST produces human-interpretable protein localization maps that quantitatively describe the similarities in the localization patterns of proteins and major subcellular compartments, without requiring manual assignment or supervised learning of these compartments. Using the budding yeast Saccharomyces cerevisiae as a model system, we show that PLAST is more accurate than existing, qualitative protein localization annotations in identifying known co-localized proteins. Furthermore, we demonstrate that PLAST can reveal protein localization-function relationships that are not obvious from these annotations. First, we identified proteins that have similar localization patterns and participate in closely-related biological processes, but do not necessarily form stable complexes with each other or localize at the same organelles. Second, we found an association between spatial and functional divergences of proteins during evolution. Surprisingly, as proteins with common ancestors evolve, they tend to develop more diverged subcellular localization patterns, but still occupy similar numbers of compartments. This suggests that divergence of protein localization might be more frequently due to the development of more specific localization patterns over ancestral compartments than the occupation of new compartments. PLAST enables systematic and quantitative analyses of protein localization-function relationships, and will be useful to elucidate protein

The Sg-1 Glycosyltransferase Locus Regulates Structural Diversity of Triterpenoid Saponins of Soybean[W][OA

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Sayama, Takashi; Ono, Eiichiro; Takagi, Kyoko; Takada, Yoshitake; Horikawa, Manabu; Nakamoto, Yumi; Hirose, Aya; Sasama, Hiroko; Ohashi, Mihoko; Hasegawa, Hisakazu; Terakawa, Teruhiko; Kikuchi, Akio; Kato, Shin; Tatsuzaki, Nana; Tsukamoto, Chigen; Ishimoto, Masao

2012-01-01

Triterpene saponins are a diverse group of biologically functional products in plants. Saponins usually are glycosylated, which gives rise to a wide diversity of structures and functions. In the group A saponins of soybean (Glycine max), differences in the terminal sugar species located on the C-22 sugar chain of an aglycone core, soyasapogenol A, were observed to be under genetic control. Further genetic analyses and mapping revealed that the structural diversity of glycosylation was determined by multiple alleles of a single locus, Sg-1, and led to identification of a UDP-sugar–dependent glycosyltransferase gene (Glyma07g38460). Although their sequences are highly similar and both glycosylate the nonacetylated saponin A0-αg, the Sg-1a allele encodes the xylosyltransferase UGT73F4, whereas Sg-1b encodes the glucosyltransferase UGT73F2. Homology models and site-directed mutagenesis analyses showed that Ser-138 in Sg-1a and Gly-138 in Sg-1b proteins are crucial residues for their respective sugar donor specificities. Transgenic complementation tests followed by recombinant enzyme assays in vitro demonstrated that sg-10 is a loss-of-function allele of Sg-1. Considering that the terminal sugar species in the group A saponins are responsible for the strong bitterness and astringent aftertastes of soybean seeds, our findings herein provide useful tools to improve commercial properties of soybean products. PMID:22611180

Poliovirus infection induces the co-localization of cellular protein SRp20 with TIA-1, a cytoplasmic stress granule protein.

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Fitzgerald, Kerry D; Semler, Bert L

2013-09-01

Different types of environmental stress cause mammalian cells to form cytoplasmic foci, termed stress granules, which contain mRNPs that are translationally silenced. These foci are transient and dynamic, and contain components of the cellular translation machinery as well as certain mRNAs and RNA binding proteins. Stress granules are known to be induced by conditions such as hypoxia, nutrient deprivation, and oxidative stress, and a number of cellular factors have been identified that are commonly associated with these foci. More recently it was discovered that poliovirus infection also induces the formation of stress granules, although these cytoplasmic foci appear to be somewhat compositionally unique. Work described here examined the punctate pattern of SRp20 (a host cell mRNA splicing protein) localization in the cytoplasm of poliovirus-infected cells, demonstrating the partial co-localization of SRp20 with the stress granule marker protein TIA-1. We determined that SRp20 does not co-localize with TIA-1, however, under conditions of oxidative stress, indicating that the close association of these two proteins during poliovirus infection is not representative of a general response to cellular stress. We confirmed that the expression of a dominant negative version of TIA-1 (TIA-1-PRD) results in the dissociation of stress granules. Finally, we demonstrated that expression of wild type TIA-1 or dominant negative TIA-1-PRD in cells during poliovirus infection does not dramatically affect viral translation. Taken together, these studies provide a new example of the unique cytoplasmic foci that form during poliovirus infection. Copyright © 2013 Elsevier B.V. All rights reserved.

Poliovirus infection induces the co-localization of cellular protein SRp20 with TIA-1, a cytoplasmic stress granule protein

Science.gov (United States)

Fitzgerald, Kerry D.; Semler, Bert L.

2013-01-01

Different types of environmental stress cause mammalian cells to form cytoplasmic foci, termed stress granules, which contain mRNPs that are translationally silenced. These foci are transient and dynamic, and contain components of the cellular translation machinery as well as certain mRNAs and RNA binding proteins. Stress granules are known to be induced by conditions such as hypoxia, nutrient deprivation, and oxidative stress, and a number of cellular factors have been identified that are commonly associated with these foci. More recently it was discovered that poliovirus infection also induces the formation of stress granules, although these cytoplasmic foci appear to be somewhat compositionally unique. Work described here examined the punctate pattern of SRp20 (a host cell mRNA splicing protein) localization in the cytoplasm of poliovirus-infected cells, demonstrating the partial co-localization of SRp20 with the stress granule marker protein TIA-1. We determined that SRp20 does not co-localize with TIA-1, however, under conditions of oxidative stress, indicating that the close association of these two proteins during poliovirus infection is not representative of a general response to cellular stress. We confirmed that the expression of a dominant negative version of TIA-1 (TIA-1-PRD) results in the dissociation of stress granules. Finally, we demonstrated that expression of wild type TIA-1 or dominant negative TIA-1-PRD in cells during poliovirus infection does not dramatically affect viral translation. Taken together, these studies provide a new example of the unique cytoplasmic foci that form during poliovirus infection. PMID:23830997

Functional classification of protein structures by local structure matching in graph representation.

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Mills, Caitlyn L; Garg, Rohan; Lee, Joslynn S; Tian, Liang; Suciu, Alexandru; Cooperman, Gene; Beuning, Penny J; Ondrechen, Mary Jo

2018-03-31

As a result of high-throughput protein structure initiatives, over 14,400 protein structures have been solved by structural genomics (SG) centers and participating research groups. While the totality of SG data represents a tremendous contribution to genomics and structural biology, reliable functional information for these proteins is generally lacking. Better functional predictions for SG proteins will add substantial value to the structural information already obtained. Our method described herein, Graph Representation of Active Sites for Prediction of Function (GRASP-Func), predicts quickly and accurately the biochemical function of proteins by representing residues at the predicted local active site as graphs rather than in Cartesian coordinates. We compare the GRASP-Func method to our previously reported method, structurally aligned local sites of activity (SALSA), using the ribulose phosphate binding barrel (RPBB), 6-hairpin glycosidase (6-HG), and Concanavalin A-like Lectins/Glucanase (CAL/G) superfamilies as test cases. In each of the superfamilies, SALSA and the much faster method GRASP-Func yield similar correct classification of previously characterized proteins, providing a validated benchmark for the new method. In addition, we analyzed SG proteins using our SALSA and GRASP-Func methods to predict function. Forty-one SG proteins in the RPBB superfamily, nine SG proteins in the 6-HG superfamily, and one SG protein in the CAL/G superfamily were successfully classified into one of the functional families in their respective superfamily by both methods. This improved, faster, validated computational method can yield more reliable predictions of function that can be used for a wide variety of applications by the community. © 2018 The Authors Protein Science published by Wiley Periodicals, Inc. on behalf of The Protein Society.

Abiotic stress responses in plants: roles of calmodulin-regulated proteins

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Virdi, Amardeep S.; Singh, Supreet; Singh, Prabhjeet

2015-01-01

Intracellular changes in calcium ions (Ca2+) in response to different biotic and abiotic stimuli are detected by various sensor proteins in the plant cell. Calmodulin (CaM) is one of the most extensively studied Ca2+-sensing proteins and has been shown to be involved in transduction of Ca2+ signals. After interacting with Ca2+, CaM undergoes conformational change and influences the activities of a diverse range of CaM-binding proteins. A number of CaM-binding proteins have also been implicated in stress responses in plants, highlighting the central role played by CaM in adaptation to adverse environmental conditions. Stress adaptation in plants is a highly complex and multigenic response. Identification and characterization of CaM-modulated proteins in relation to different abiotic stresses could, therefore, prove to be essential for a deeper understanding of the molecular mechanisms involved in abiotic stress tolerance in plants. Various studies have revealed involvement of CaM in regulation of metal ions uptake, generation of reactive oxygen species and modulation of transcription factors such as CAMTA3, GTL1, and WRKY39. Activities of several kinases and phosphatases have also been shown to be modulated by CaM, thus providing further versatility to stress-associated signal transduction pathways. The results obtained from contemporary studies are consistent with the proposed role of CaM as an integrator of different stress signaling pathways, which allows plants to maintain homeostasis between different cellular processes. In this review, we have attempted to present the current state of understanding of the role of CaM in modulating different stress-regulated proteins and its implications in augmenting abiotic stress tolerance in plants. PMID:26528296

Stress and Protein Turnover in Lemna minor1

Science.gov (United States)

Cooke, Robert J.; Oliver, Jane; Davies, David D.

1979-01-01

Transfer of fronds of Lemna minor L. to adverse growth conditions or stress situations causes a lowering of the growth rate and a loss of soluble protein per frond, the extent of the loss being dependent on the nature of the stress. The loss or protein is due to two factors: (a) a decrease in the rate constant of protein synthesis (ks); (b) an increase in the rate constant of protein degradation (kd). In plants adapted to the stresses, protein synthesis increases and the initially rapid rate of proteolysis is reduced. Addition of abscisic acid both lowers ks and increases kd, whereas benzyladenine seems to alleviate the effects of stress on protein content by decreasing kd rather than by altering ks. Based on the measurement of enzyme activities, stress-induced protein degradation appears to be a general phenomenon, affecting many soluble proteins. The adaptive significance of stress-induced proteolysis is discussed. PMID:16661102

Life management of SG for WWER plants

International Nuclear Information System (INIS)

Trunov, N. B.; Dragunov, Yu. G.; Banyuk, G. F.

2004-01-01

Nowadays, 252 steam generators (SG) of horizontal type are in operation at WWER plants constructed by the Russian designs. In connection with end of the specified service life of the reactor plant equal to 30 years the activities are performed on service life extension of the main equipment including the SG. At some Units, throughout the design service life of SG there were problems resulting in necessity of SG replacement. At the same time the SGs at some Units are in successful operation above the design service life. This report deals with the peculiarities of operation of the horizontal SGs and the problems to be highlighted as the most important for service life extension. The main component to determine possibility for SG service life extension is the SG tubing. As the operating experience shows it is water chemistry of the secondary circuit that is the main factor influencing operability of the SG tubing. Therefore, differences in water chemistry organization leads to significant differences in operability of the SG tubing at various Units and in some cases within one Unit. Owing to the fact that the cases of water chemistry disturbance and the process of tubes fouling with the corrosion products of the main condensate system are not excluded, the damages continue to occur. Tube integrity shall be inspected by eddy current method using the various instrument complexes. This method has certain disadvantages but allows to estimate the degree and direction of degradation processes. The results of eddy current test (ECT) can be used to determine the plugging criterion for defective tubes. The significant number of defective tubes at some Units makes a choice of the plugging criterion to be an important problem, on which solution the SG safety, reliability and service life depends. The report deals with directions of activities in service life management for the SG at WWER plants. Main activities are improvement of water chemistry and non-destructive tests.(author)

Reduced Insulin/IGF-1 Signaling Restores the Dynamic Properties of Key Stress Granule Proteins during Aging

Directory of Open Access Journals (Sweden)

Marie C. Lechler

2017-01-01

Full Text Available Summary: Low-complexity “prion-like” domains in key RNA-binding proteins (RBPs mediate the reversible assembly of RNA granules. Individual RBPs harboring these domains have been linked to specific neurodegenerative diseases. Although their aggregation in neurodegeneration has been extensively characterized, it remains unknown how the process of aging disturbs RBP dynamics. We show that a wide variety of RNA granule components, including stress granule proteins, become highly insoluble with age in C. elegans and that reduced insulin/insulin-like growth factor 1 (IGF-1 daf-2 receptor signaling efficiently prevents their aggregation. Importantly, stress-granule-related RBP aggregates are associated with reduced fitness. We show that heat shock transcription factor 1 (HSF-1 is a main regulator of stress-granule-related RBP aggregation in both young and aged animals. During aging, increasing DAF-16 activity restores dynamic stress-granule-related RBPs, partly by decreasing the buildup of other misfolded proteins that seed RBP aggregation. Longevity-associated mechanisms found to maintain dynamic RBPs during aging could be relevant for neurodegenerative diseases. : Lechler et al. show that RNA-binding proteins (RBPs including stress granule proteins are prone to aggregate with age in C. elegans. Aggregation of stress granule RBPs with “prion-like” domains is associated with reduced fitness. Their aggregation is prevented by longevity pathways and promoted by the aggregation of other misfolded proteins. Keywords: neurodegenerative diseases, Caenorhabditis elegans, protein aggregation, aging, RNA-binding proteins, stress granules, HSF-1, DAF-2, longevity

RPA accumulation during class switch recombination represents 5'-3' DNA-end resection during the S-G2/M phase of the cell cycle.

Science.gov (United States)

Yamane, Arito; Robbiani, Davide F; Resch, Wolfgang; Bothmer, Anne; Nakahashi, Hirotaka; Oliveira, Thiago; Rommel, Philipp C; Brown, Eric J; Nussenzweig, Andre; Nussenzweig, Michel C; Casellas, Rafael

2013-01-31

Activation-induced cytidine deaminase (AID) promotes chromosomal translocations by inducing DNA double-strand breaks (DSBs) at immunoglobulin (Ig) genes and oncogenes in the G1 phase. RPA is a single-stranded DNA (ssDNA)-binding protein that associates with resected DSBs in the S phase and facilitates the assembly of factors involved in homologous repair (HR), such as Rad51. Notably, RPA deposition also marks sites of AID-mediated damage, but its role in Ig gene recombination remains unclear. Here, we demonstrate that RPA associates asymmetrically with resected ssDNA in response to lesions created by AID, recombination-activating genes (RAG), or other nucleases. Small amounts of RPA are deposited at AID targets in G1 in an ATM-dependent manner. In contrast, recruitment in the S-G2/M phase is extensive, ATM independent, and associated with Rad51 accumulation. In the S-G2/M phase, RPA increases in nonhomologous-end-joining-deficient lymphocytes, where there is more extensive DNA-end resection. Thus, most RPA recruitment during class switch recombination represents salvage of unrepaired breaks by homology-based pathways during the S-G2/M phase of the cell cycle. Copyright © 2013 The Authors. Published by Elsevier Inc. All rights reserved.

The heat-shock protein/chaperone network and multiple stress resistance.

Science.gov (United States)

Jacob, Pierre; Hirt, Heribert; Bendahmane, Abdelhafid

2017-04-01

Crop yield has been greatly enhanced during the last century. However, most elite cultivars are adapted to temperate climates and are not well suited to more stressful conditions. In the context of climate change, stress resistance is a major concern. To overcome these difficulties, scientists may help breeders by providing genetic markers associated with stress resistance. However, multistress resistance cannot be obtained from the simple addition of single stress resistance traits. In the field, stresses are unpredictable and several may occur at once. Consequently, the use of single stress resistance traits is often inadequate. Although it has been historically linked with the heat stress response, the heat-shock protein (HSP)/chaperone network is a major component of multiple stress responses. Among the HSP/chaperone 'client proteins', many are primary metabolism enzymes and signal transduction components with essential roles for the proper functioning of a cell. HSPs/chaperones are controlled by the action of diverse heat-shock factors, which are recruited under stress conditions. In this review, we give an overview of the regulation of the HSP/chaperone network with a focus on Arabidopsis thaliana. We illustrate the role of HSPs/chaperones in regulating diverse signalling pathways and discuss several basic principles that should be considered for engineering multiple stress resistance in crops through the HSP/chaperone network. © 2016 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.

Protein stress and stress proteins: implications in aging and disease

Indian Academy of Sciences (India)

Madhu Sudhan

2007-04-02

Apr 2, 2007 ... (iii) modulating protein activity via stabilization and/or maturation to ... Resistance to any physical stress is correlated with longevity in many, if not all .... range of pathologies including cancer, diabetes, immune- problems and ...

ER stress proteins in autoimmune and inflammatory diseases

Directory of Open Access Journals (Sweden)

Daisuke eMorito

2012-03-01

Full Text Available Over the past two decades, heat shock proteins (HSPs have been implicated in inflammatory responses and autoimmunity. HSPs were originally believed to maintain protein quality control in the cytosol. However, they also exist extracellularly and appear to act as inflammatory factors. Recently, a growing body of evidence suggested that the other class of stress proteins such as, endoplasmic reticulum (ER stress proteins, which originally act as protein quality control factors in the secretory pathway and are induced by ER stress in inflammatory lesions, also participate in inflammation and autoimmunity. The immunoglobulin heavy-chain binding protein (Bip/glucose-regulated protein 78 (Grp78, homocysteine-induced ER protein (Herp, calnexin, calreticulin, glucose-regulated protein 94 (Grp94/gp96, oxygen-regulated protein 150 (ORP150 and heat shock protein 47 (Hsp47/Serpin H1, which are expressed not only in the ER but also occasionally at the cell surface play pathophysiological roles in autoimmune and inflammatory diseases as pro- or anti-inflammatory factors. Here we describe the accumulating evidence of the participation of ER stress proteins in autoimmunity and inflammation and discuss the critical differences between the two classes of stress proteins.

Coping strategy and social support modify the association between perceived stress and C-reactive protein: a longitudinal study of healthy men and women.

Science.gov (United States)

Shimanoe, Chisato; Hara, Megumi; Nishida, Yuichiro; Nanri, Hinako; Otsuka, Yasuko; Horita, Mikako; Yasukata, Jun; Miyoshi, Nobuyuki; Yamada, Yosuke; Higaki, Yasuki; Tanaka, Keitaro

2018-05-01

Inconsistent associations have been reported between perceived stress and C-reactive protein (CRP), a marker of systemic inflammation. We previously observed a male-specific inverse relationship between perceived stress and CRP in a cross-sectional study. In the present study, we examined the longitudinal association between changes in perceived stress and CRP, and further analyzed whether changes in coping strategies and social support modify this association. This study included 8454 participants in both a baseline survey and a follow-up survey 5 years later. Psychosocial measures (i.e. perceived stress, coping strategies, and social support) and CRP concentrations were measured by identical means in both surveys. Consistent with our previous findings, increased perceived stress was significantly associated with lower CRP in men (p trend  = .037), but not in women. Increased "emotional expression," a coping strategy, was also associated with lower CRP in women (p trend  = .024). Furthermore, interactions between perceived stress and a coping strategy (positive reappraisal) or social support on CRP were found in men (p interaction  = .007 and .038, respectively); the above inverse association between stress and CRP was not detected for participants with diminished positive reappraisal or social support. In conclusion, increases in perceived stress during a 5-year period were associated with decreases in CRP among healthy men, and the observed association was possibly modified by coping strategy or social support.

Protective effects of transforming growth factor β2 in intestinal epithelial cells by regulation of proteins associated with stress and endotoxin responses.

Directory of Open Access Journals (Sweden)

Duc Ninh Nguyen

Full Text Available Transforming growth factor (TGF-β2 is an important anti-inflammatory protein in milk and colostrum. TGF-β2 supplementation appears to reduce gut inflammatory diseases in early life, such as necrotizing enterocolitis (NEC in young mice. However, the molecular mechanisms by which TGF-β2 protects immature intestinal epithelial cells (IECs remain to be more clearly elucidated before interventions in infants can be considered. Porcine IECs PsIc1 were treated with TGF-β2 and/or lipopolysaccharide (LPS, and changes in the cellular proteome were subsequently analyzed using two-dimensional gel electrophoresis-MS and LC-MS-based proteomics. TGF-β2 alone induced the differential expression of 13 proteins and the majority of the identified proteins were associated with stress responses, TGF-β and Toll-like receptor 4 signaling cascades. In particular, a series of heat shock proteins had similar differential trends as previously shown in the intestine of NEC-resistant preterm pigs and young mice. Furthermore, LC-MS-based proteomics and Western blot analyses revealed 20 differentially expressed proteins following treatment with TGF-β2 in LPS-challenged IECs. Thirteen of these proteins were associated with stress response pathways, among which five proteins were altered by LPS and restored by TGF-β2, whereas six were differentially expressed only by TGF-β2 in LPS-challenged IECs. Based on previously reported biological functions, these patterns indicate the anti-stress and anti-inflammatory effects of TGF-β2 in IECs. We conclude that TGF-β2 of dietary or endogenous origin may regulate the IEC responses against LPS stimuli, thereby supporting cellular homeostasis and innate immunity in response to bacterial colonization, and the first enteral feeding in early life.

ER stress affects processing of MHC class I-associated peptides

Directory of Open Access Journals (Sweden)

Meloche Sylvain

2009-02-01

Full Text Available Abstract Background Viral infection and neoplastic transformation trigger endoplasmic reticulum (ER stress. Thus, a large proportion of the cells that must be recognized by the immune system are stressed cells. Cells respond to ER stress by launching the unfolded protein response (UPR. The UPR regulates the two key processes that control major histocompatibility complex class I (MHC I-peptide presentation: protein synthesis and degradation. We therefore asked whether and how the UPR impinges on MHC I-peptide presentation. Results We evaluated the impact of the UPR on global MHC I expression and on presentation of the H2Kb-associated SIINFEKL peptide. EL4 cells stably transfected with vectors coding hen egg lysozyme (HEL-SIINFEKL protein variants were stressed with palmitate or exposed to glucose deprivation. UPR decreased surface expression of MHC I but did not affect MHC I mRNA level nor the total amount of intracellular MHC I proteins. Impaired MHC I-peptide presentation was due mainly to reduced supply of peptides owing to an inhibition of overall protein synthesis. Consequently, generation of H2Kb-SIINFEKL complexes was curtailed during ER stress, illustrating how generation of MHC I peptide ligands is tightly coupled to ongoing protein synthesis. Notably, the UPR-induced decline of MHC I-peptide presentation was more severe when the protein source of peptides was localized in the cytosol than in the ER. This difference was not due to changes in the translation rates of the precursor proteins but to increased stability of the cytosolic protein during ER stress. Conclusion Our results demonstrate that ER stress impairs MHC I-peptide presentation, and that it differentially regulates expression of ER- vs. cytosol-derived peptides. Furthermore, this work illustrates how ER stress, a typical feature of infected and malignant cells, can impinge on cues for adaptive immune recognition.

FMDV-induced stress granules are disrupted by the viral L-protease

DEFF Research Database (Denmark)

Polacek, Charlotta; Belsham, Graham; McInerney, Gerald

2014-01-01

Eukaryotic cells respond to environmental stress by entering a state of reduced protein synthesis, redirecting resources to damage control and defense. This reduced translation is closely linked to the formation of cytoplasmic stress granules (SGs). SGs are multicomponent foci, which contain...... stalled translation preinitiation complexes, including polyadenylated mRNAs, and several aggregation-prone RNA binding factors, such as the Ras-GAP SH3 domain-binding protein (G3BP) that enable their formation. Once the stress is lifted, the stalled complexes from the SGs are believed to re......-engage in translation, facilitating cellular recovery. A growing body of evidence shows that various viruses can trigger SG formation. However, the presence of SGs may not be beneficial to the virus and many viruses have found ways to circumvent, disrupt or even utilize these granules, suggesting a role for SGs...

Overexpression of heat shock GroEL stress protein in leptospiral biofilm.

Science.gov (United States)

Vinod Kumar, K; Lall, Chandan; Vimal Raj, R; Vedhagiri, K; Kartick, C; Surya, P; Natarajaseenivasan, K; Vijayachari, P

2017-01-01

Leptospira is the causative agent of leptospirosis, which is an emerging zoonotic disease. Recent studies on Leptospira have demonstrated biofilm formation on abiotic surfaces. The protein expressed in the biofilm was investigated by using SDS-PAGE and immunoblotting in combination with MALDI-TOF mass spectrometry. The proteins expressed in Leptospira biofilm and planktonic cells was analyzed and compared. Among these proteins, one (60 kDa) was found to overexpress in biofilm as compared to the planktonic cells. MALDI-TOF analysis identified this protein as stress and heat shock chaperone GroEL. Our findings demonstrate that GroEL is associated with Leptospira biofilm. GroEL is conserved, highly immunogenic and a prominent stress response protein in pathogenic Leptospira spp., which may have clinical relevance. Copyright © 2016 Elsevier Ltd. All rights reserved.

Fanconi anemia proteins and endogenous stresses

Energy Technology Data Exchange (ETDEWEB)

Pang Qishen [Division of Experimental Hematology and Cancer Biology, Cincinnati Children' s Research Foundation, Cincinnati, OH (United States); Department of Pediatrics, University of Cincinnati College of Medicine, Cincinnati, OH (United States); Andreassen, Paul R., E-mail: Paul.Andreassen@cchmc.org [Division of Experimental Hematology and Cancer Biology, Cincinnati Children' s Research Foundation, Cincinnati, OH (United States); Department of Pediatrics, University of Cincinnati College of Medicine, Cincinnati, OH (United States)

2009-07-31

Each of the thirteen identified Fanconi anemia (FA) genes is required for resistance to DNA interstrand crosslinking agents, such as mitomycin C, cisplatin, and melphalan. While these agents are excellent tools for understanding the function of FA proteins in DNA repair, it is uncertain whether a defect in the removal of DNA interstrand crosslinks (ICLs) is the basis for the pathophysiology of FA. For example, DNA interstrand crosslinking agents induce other types of DNA damage, in addition to ICLs. Further, other DNA-damaging agents, such as ionizing or ultraviolet radiation, activate the FA pathway, leading to monoubiquitination of FANCD2 and FANCI. Also, FA patients display congenital abnormalities, hematologic deficiencies, and a predisposition to cancer in the absence of an environmental source of ICLs that is external to cells. Here we consider potential sources of endogenous DNA damage, or endogenous stresses, to which FA proteins may respond. These include ICLs formed by products of lipid peroxidation, and other forms of oxidative DNA damage. FA proteins may also potentially respond to telomere shortening or replication stress. Defining these endogenous sources of DNA damage or stresses is critical for understanding the pathogenesis of deficiencies for FA proteins. We propose that FA proteins are centrally involved in the response to replication stress, including replication stress arising from oxidative DNA damage.

Fanconi anemia proteins and endogenous stresses

International Nuclear Information System (INIS)

Pang Qishen; Andreassen, Paul R.

2009-01-01

Each of the thirteen identified Fanconi anemia (FA) genes is required for resistance to DNA interstrand crosslinking agents, such as mitomycin C, cisplatin, and melphalan. While these agents are excellent tools for understanding the function of FA proteins in DNA repair, it is uncertain whether a defect in the removal of DNA interstrand crosslinks (ICLs) is the basis for the pathophysiology of FA. For example, DNA interstrand crosslinking agents induce other types of DNA damage, in addition to ICLs. Further, other DNA-damaging agents, such as ionizing or ultraviolet radiation, activate the FA pathway, leading to monoubiquitination of FANCD2 and FANCI. Also, FA patients display congenital abnormalities, hematologic deficiencies, and a predisposition to cancer in the absence of an environmental source of ICLs that is external to cells. Here we consider potential sources of endogenous DNA damage, or endogenous stresses, to which FA proteins may respond. These include ICLs formed by products of lipid peroxidation, and other forms of oxidative DNA damage. FA proteins may also potentially respond to telomere shortening or replication stress. Defining these endogenous sources of DNA damage or stresses is critical for understanding the pathogenesis of deficiencies for FA proteins. We propose that FA proteins are centrally involved in the response to replication stress, including replication stress arising from oxidative DNA damage.

Overexpression of a Medicago truncatula stress-associated protein gene (MtSAP1) leads to nitric oxide accumulation and confers osmotic and salt stress tolerance in transgenic tobacco.

Science.gov (United States)

Charrier, Aurélie; Planchet, Elisabeth; Cerveau, Delphine; Gimeno-Gilles, Christine; Verdu, Isabelle; Limami, Anis M; Lelièvre, Eric

2012-08-01

The impact of Medicago truncatula stress-associated protein gene (MtSAP1) overexpression has been investigated in Nicotiana tabacum transgenic seedlings. Under optimal conditions, transgenic lines overexpressing MtSAP1 revealed better plant development and higher chlorophyll content as compared to wild type seedlings. Interestingly, transgenic lines showed a stronger accumulation of nitric oxide (NO), a signaling molecule involved in growth and development processes. This NO production seemed to be partially nitrate reductase dependent. Due to the fact that NO has been also reported to play a role in tolerance acquisition of plants to abiotic stresses, the responses of MtSAP1 overexpressors to osmotic and salt stress have been studied. Compared to the wild type, transgenic lines were less affected in their growth and development. Moreover, NO content in MtSAP1 overexpressors was always higher than that detected in wild seedlings under stress conditions. It seems that this better tolerance induced by MtSAP1 overexpression could be associated with this higher NO production that would enable seedlings to reach a high protection level to prepare them to cope with abiotic stresses.

The heat shock protein/chaperone network and multiple stress resistance

KAUST Repository

Jacob, Pierre; Hirt, Heribert; Bendahmane, Abdelhafid

2016-01-01

Crop yield has been greatly enhanced during the last century. However, most elite cultivars are adapted to temperate climates and are not well suited to more stressful conditions. In the context of climate change, stress resistance is a major concern. To overcome these difficulties, scientists may help breeders by providing genetic markers associated with stress resistance. However, multi-stress resistance cannot be obtained from the simple addition of single stress resistance traits. In the field, stresses are unpredictable and several may occur at once. Consequently, the use of single stress resistance traits is often inadequate. Although it has been historically linked with the heat stress response, the heat shock protein (HSP)/chaperone network is a major component of multiple stress responses. Among the HSP/chaperone

The heat shock protein/chaperone network and multiple stress resistance

KAUST Repository

Jacob, Pierre

2016-11-15

Crop yield has been greatly enhanced during the last century. However, most elite cultivars are adapted to temperate climates and are not well suited to more stressful conditions. In the context of climate change, stress resistance is a major concern. To overcome these difficulties, scientists may help breeders by providing genetic markers associated with stress resistance. However, multi-stress resistance cannot be obtained from the simple addition of single stress resistance traits. In the field, stresses are unpredictable and several may occur at once. Consequently, the use of single stress resistance traits is often inadequate. Although it has been historically linked with the heat stress response, the heat shock protein (HSP)/chaperone network is a major component of multiple stress responses. Among the HSP/chaperone

[Knocking-out extra domain A alternative splice fragment of fibronectin using a clustered regularly interspaced short palindromic repeats/associated proteins 9 system].

Science.gov (United States)

Yang, Yue; Wang, Haicheng; Xu, Shuyu; Peng, Jing; Jiang, Jiuhui; Li, Cuiying

2015-08-01

To investigate the effect of the fibronectin extra domain A on the aggressiveness of salivary adenoid cystic carcinoma (SACC) cells, via the clustered regularly interspaced short palindromic repeats (CRISPR)/ associated proteins (Cas) system. One sgRNA was designed to target the upstream of the genome sequences of extra domain A(EDA) exon and the downstream. Then the sgRNA was linked into plasmid PX-330 and transfected into SACC-83 cells. PCR and DNA sequence were used to testify the knockout cells, and the monoclones of EDA absent SACC cells were selected (A+C-2, A+C-6, B+C-10). CCK-8 cell proliferation and invasion was then tested in control group and the experimental group. The sgRNA was successfully linked into PX-330 plasmid. Part of adenoid cystic carcinoma cells' SACC-83 genomic EDA exon was knocked out, and the knockdown efficiency was above 70%, but the total amount of fibronectin did not change significantly. Three monoclones of EDA absent SACC- 83 cells were successfully selected with diminished migration and proliferation. The CRISPR/Cas9 system was a simplified system with relatively high knockout efficiency and EDA knockout could inhibiting SACC cell's mobility and invasiveness.

Benchmarking CRISPR on-target sgRNA design.

Science.gov (United States)

Yan, Jifang; Chuai, Guohui; Zhou, Chi; Zhu, Chenyu; Yang, Jing; Zhang, Chao; Gu, Feng; Xu, Han; Wei, Jia; Liu, Qi

2017-02-15

CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)-based gene editing has been widely implemented in various cell types and organisms. A major challenge in the effective application of the CRISPR system is the need to design highly efficient single-guide RNA (sgRNA) with minimal off-target cleavage. Several tools are available for sgRNA design, while limited tools were compared. In our opinion, benchmarking the performance of the available tools and indicating their applicable scenarios are important issues. Moreover, whether the reported sgRNA design rules are reproducible across different sgRNA libraries, cell types and organisms remains unclear. In our study, a systematic and unbiased benchmark of the sgRNA predicting efficacy was performed on nine representative on-target design tools, based on six benchmark data sets covering five different cell types. The benchmark study presented here provides novel quantitative insights into the available CRISPR tools. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Structural genomics: keeping up with expanding knowledge of the protein universe

Science.gov (United States)

Grabowski, Marek; Joachimiak, Andrzej; Otwinowski, Zbyszek; Minor, Wladek

2010-01-01

Structural characterization of the protein universe is the main mission of Structural Genomics (SG) programs. However, progress in gene sequencing technology, set in motion in the 1990s, has resulted in rapid expansion of protein sequence space — a twelvefold increase in the past seven years. For the SG field, this creates new challenges and necessitates a reassessment of its strategies. Nevertheless, despite the growth of sequence space, at present nearly half of the content of the Swiss-Prot database and over 40% of Pfam protein families can be structurally modeled based on structures determined so far, with SG projects making an increasingly significant contribution. The SG contribution of new Pfam structures nearly doubled from 27.2% in 2003 to 51.6% in 2006. PMID:17587562

Structural genomics: keeping up with expanding knowledge of the protein universe.

Science.gov (United States)

Grabowski, Marek; Joachimiak, Andrzej; Otwinowski, Zbyszek; Minor, Wladek

2007-06-01

Structural characterization of the protein universe is the main mission of Structural Genomics (SG) programs. However, progress in gene sequencing technology, set in motion in the 1990s, has resulted in rapid expansion of protein sequence space--a twelvefold increase in the past seven years. For the SG field, this creates new challenges and necessitates a re-assessment of its strategies. Nevertheless, despite the growth of sequence space, at present nearly half of the content of the Swiss-Prot database and over 40% of Pfam protein families can be structurally modeled based on structures determined so far, with SG projects making an increasingly significant contribution. The SG contribution of new Pfam structures nearly doubled from 27.2% in 2003 to 51.6% in 2006.

Experimental and numerical study of deposit formation in secondary side SG TSP by electrokinetic approach

International Nuclear Information System (INIS)

Guillodo, Michael; Foucault, Marc; Ryckelynck, Natacha; Chahma, Farah; Guingo, Mathieu; Mansour, Carine; Alos-Ramos, Olga; Corredera, Geraldine

2012-09-01

Corrosion products deposit formation observed in PWR steam generators (SGs) - related to SG free span fouling and SG clogging - is now reported since several years. SG clogging is a localized phenomenon observed between the leading edge of the Tube Support Plate (TSP) and SG tubing materials. Based on visual inspections, it was found that the gaps between SG tubing material and TSP at the lower part of the broached holes were getting progressively blocked. Therefore, for safe operation, most affected PWRs had to be operated at reduced power. TSP blockage was mainly observed for low-pH water chemistry conditioning, which directly depends on the operating water chemistry. The TSP blockage mechanism is complex due to the localized conditions in which flow pattern change, chemistry and electrochemical conditions are not well understood. Electrokinetic considerations could be pointed out to explain the coupling of chemistry, materials and thermohydraulic (T/H) conditions. In this frame AREVA and EDF have launched a long-term R and D program in order to understand the mechanisms driving the formation of SG clogging. This study based on parametric laboratory tests aims to assess the role of secondary water chemistry, material and T/H conditions on deposit formation. The experimental approach focused on electrokinetic measurements of metallic substrates and on the assessment of oxidation properties of materials in secondary side chemistry. An overall analysis of recent results is presented to address SG deposit formation in secondary water chemistry for various conditioning amines - morpholine, ethanolamine and dimethylamine. To complete the study, the experimental results have been correlated to CFD simulations of particle deposition, by means of stochastic Lagrangian models. These calculations have in particular reproduced correctly the location of the most important particle deposit (the leading edge of the test tube), and have stressed the influence of the

Roles of endoplasmic reticulum stress and unfolded protein response associated genes in seed stratification and bud endodormancy during chilling accumulation in Prunus persica.

Directory of Open Access Journals (Sweden)

Xi Ling Fu

Full Text Available Dormancy mechanisms in seeds and buds arrest growth until environmental conditions are optimal for development. A genotype-specific period of chilling is usually required to release dormancy, but the underlying molecular mechanisms are still not fully understood. To discover transcriptional pathways associated with dormancy release common to seed stratification and bud endodormancy, we explored the chilling-dependent expression of 11 genes involved in endoplasmic reticulum stress and the unfolded protein response signal pathways. We propose that endoplasmic reticulum stress and the unfolded protein response impact on seed as well as bud germination and development by chilling-dependent mechanisms. The emerging discovery of similarities between seed stratification and bud endodormancy status indicate that these two processes are probably regulated by common endoplasmic reticulum stress and unfolded protein response signalling pathways. Clarification of regulatory pathways common to both seed and bud dormancy may enhance understanding of the mechanisms underlying dormancy and breeding programs may benefit from earlier prediction of chilling requirements for uniform blooming of novel genotypes of deciduous fruit tree species.

Characterization of proteins in soybean roots under flooding and drought stresses.

Science.gov (United States)

Oh, MyeongWon; Komatsu, Setsuko

2015-01-30

Flooding and drought affect soybean growth because soybean is a stress-sensitive crop. In 2-day-old plants exposed to 2-day flooding or drought, the fresh weight of roots was markedly suppressed, although the root morphology clearly differed between two conditions. To understand the response mechanisms of soybean to flooding and drought stresses, a gel-free proteomic technique was used. A total of 97 and 48 proteins were significantly changed in response to flooding and drought stresses, respectively. Proteins involved in protein synthesis were decreased by flooding stress and increased by drought. Glycolysis-related proteins were increased in roots by both flooding and drought stresses. Fermentation, stress, and cell wall-related proteins were increased in response to flooding stress, whereas cell organization and redox-related proteins were increased under drought stress. Among the identified proteins, three S-adenosylmethionine synthetases were commonly decreased and increased in response to flooding and drought stresses, respectively. The mRNA expression levels of S-adenosylmethionine synthetase genes displayed a similar tendency to the changes in protein abundance. These results suggest that S-adenosylmethionine synthetase is involved in the regulation of stress response because it was changed in response to flooding and drought stresses. This study reported on the response mechanisms of soybean to flooding and drought stresses using the gel-free proteomic technique. Proteins involved in protein synthesis were decreased by flooding stress and increased by drought. Glycolysis-related proteins were increased in roots by both flooding and drought stresses. Fermentation, stress, and cell wall-related proteins were increased in response to flooding stress, whereas cell organization and redox-related proteins were increased under drought stress. Among the identified proteins, three S-adenosylmethionine synthetases were commonly decreased and increased in response to

Selection of Reference Genes for qRT-PCR Analysis of Gene Expression in Stipa grandis during Environmental Stresses.

Directory of Open Access Journals (Sweden)

Dongli Wan

Full Text Available Stipa grandis P. Smirn. is a dominant plant species in the typical steppe of the Xilingole Plateau of Inner Mongolia. Selection of suitable reference genes for the quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR is important for gene expression analysis and research into the molecular mechanisms underlying the stress responses of S. grandis. In the present study, 15 candidate reference genes (EF1 beta, ACT, GAPDH, SamDC, CUL4, CAP, SNF2, SKIP1, SKIP5, SKIP11, UBC2, UBC15, UBC17, UCH, and HERC2 were evaluated for their stability as potential reference genes for qRT-PCR under different stresses. Four algorithms were used: GeNorm, NormFinder, BestKeeper, and RefFinder. The results showed that the most stable reference genes were different under different stress conditions: EF1beta and UBC15 during drought and salt stresses; ACT and GAPDH under heat stress; SKIP5 and UBC17 under cold stress; UBC15 and HERC2 under high pH stress; UBC2 and UBC15 under wounding stress; EF1beta and UBC17 under jasmonic acid treatment; UBC15 and CUL4 under abscisic acid treatment; and HERC2 and UBC17 under salicylic acid treatment. EF1beta and HERC2 were the most suitable genes for the global analysis of all samples. Furthermore, six target genes, SgPOD, SgPAL, SgLEA, SgLOX, SgHSP90 and SgPR1, were selected to validate the most and least stable reference genes under different treatments. Our results provide guidelines for reference gene selection for more accurate qRT-PCR quantification and will promote studies of gene expression in S. grandis subjected to environmental stress.

Model for Stress-induced Protein Degradation in Lemna minor1

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Cooke, Robert J.; Roberts, Keith; Davies, David D.

1980-01-01

Transfer of Lemna minor fronds to adverse or stress conditions produces a large increase in the rate of protein degradation. Cycloheximide partially inhibits stress-induced protein degradation and also partially inhibits the protein degradation which occurs in the absence of stress. The increased protein degradation does not appear to be due to an increase in activity of soluble proteolytic enzymes. Biochemical evidence indicates that stress, perhaps acting via hormones, affects the permeability of certain membranes, particularly the tonoplast. A general model for stress-induced protein degradation is presented in which changes in membrane properties allow vacuolar proteolytic enzymes increased access to cytoplasmic proteins. PMID:16661588

Development and application of an antibody-based protein microarray to assess physiological stress in grizzly bears (Ursus arctos).

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Carlson, Ruth I; Cattet, Marc R L; Sarauer, Bryan L; Nielsen, Scott E; Boulanger, John; Stenhouse, Gordon B; Janz, David M

2016-01-01

A novel antibody-based protein microarray was developed that simultaneously determines expression of 31 stress-associated proteins in skin samples collected from free-ranging grizzly bears (Ursus arctos) in Alberta, Canada. The microarray determines proteins belonging to four broad functional categories associated with stress physiology: hypothalamic-pituitary-adrenal axis proteins, apoptosis/cell cycle proteins, cellular stress/proteotoxicity proteins and oxidative stress/inflammation proteins. Small skin samples (50-100 mg) were collected from captured bears using biopsy punches. Proteins were isolated and labelled with fluorescent dyes, with labelled protein homogenates loaded onto microarrays to hybridize with antibodies. Relative protein expression was determined by comparison with a pooled standard skin sample. The assay was sensitive, requiring 80 µg of protein per sample to be run in triplicate on the microarray. Intra-array and inter-array coefficients of variation for individual proteins were generally bears. This suggests that remotely delivered biopsy darts could be used in future sampling. Using generalized linear mixed models, certain proteins within each functional category demonstrated altered expression with respect to differences in year, season, geographical sampling location within Alberta and bear biological parameters, suggesting that these general variables may influence expression of specific proteins in the microarray. Our goal is to apply the protein microarray as a conservation physiology tool that can detect, evaluate and monitor physiological stress in grizzly bears and other species at risk over time in response to environmental change.

The interactive association between heat shock factor 1 and heat shock proteins in primary myocardial cells subjected to heat stress.

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Tang, Shu; Chen, Hongbo; Cheng, Yanfen; Nasir, Mohammad Abdel; Kemper, Nicole; Bao, Endong

2016-01-01

Heat shock factor 1 (HSF1) is a heat shock transcription factor that rapidly induces heat shock gene transcription following thermal stress. In this study, we subjected primary neonatal rat myocardial cells to heat stress in vitro to create a model system for investigating the trends in expression and association between various heat shock proteins (HSPs) and HSF1 under adverse environmental conditions. After the cells were subjected to heat stress at 42˚C for different periods of time, HSP and HSF1 mRNA and protein levels were detected by qPCR and western blot analysis in the heat-stressed cells. The HSF1 expression levels significantly increased in the cells following 120 min of exposure to heat stess compared to the levels observed at the beginning of heat stress exposure. HSP90 followed a similar trend in expression to HSF1, whereas HSP70 followed an opposite trend. However, no significant changes were observed in the crystallin, alpha B (CRYAB, also known as HSP beta-5) expression levels during the 480‑min period of exposure to heat stress. The interaction between the HSPs and HSF1 was analyzed by STRING 9.1, and it was found that HSF1 interacted with HSP90 and HSP70, and that it did not play a role in regulating CRYAB expression. Based on our findings, HSP70 may suppress HSF1 in rat myocardial cells under conditions of heat stress. Furthermore, our data demonstrate that HSF1 is not the key factor for all HSPs, and this was particularly the case for CRYAB.

Gene Expression Responses to FUS, EWS, and TAF15 Reduction and Stress Granule Sequestration Analyses Identifies FET-Protein Non-Redundant Functions

DEFF Research Database (Denmark)

Blechingberg, Jenny; Luo, Yonglun; Bolund, Lars

2012-01-01

The FET family of proteins is composed of FUS/TLS, EWS/EWSR1, and TAF15 and possesses RNA- and DNA-binding capacities. The FET-proteins are involved in transcriptional regulation and RNA processing, and FET-gene deregulation is associated with development of cancer and protein granule formations...... in amyotrophic lateral sclerosis, frontotemporal lobar degeneration, and trinucleotide repeat expansion diseases. We here describe a comparative characterization of FET-protein localization and gene regulatory functions. We show that FUS and TAF15 locate to cellular stress granules to a larger extend than EWS....... FET-proteins have no major importance for stress granule formation and cellular stress responses, indicating that FET-protein stress granule association most likely is a downstream response to cellular stress. Gene expression analyses showed that the cellular response towards FUS and TAF15 reduction...

Periodic usage of low-protein methionine-fortified diets in broiler chickens under high ambient temperature conditions: effects on performance, slaughter traits, leukocyte profiles and antibody response

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Ghasemi, Hossein Ali; Ghasemi, Rohollah; Torki, Mehran

2014-09-01

This study was performed to evaluate the effects of adding methionine supplements to low-protein diets and subsequent re-feeding with a normal diet on the productive performance, slaughter parameters, leukocyte profiles and antibody response in broiler chickens reared under heat stress conditions. During the whole experimental period (6-49 days), the birds were raised in battery cages located in high ambient temperature in an open-sided housing system. A total of 360 6-day-old male chickens were divided into six treatments in six replicates with ten chicks each. Six isoenergetic diets, with similar total sulfur amino acids levels, were formulated to provide 100 and 100 (control), 85 and 100 (85S), 70 and 100 (70S), 85 and 85 (85SG), 70 and 85 (70S85G), and 70 and 70 % (70SG) of National Research Council recommended levels for crude protein during the starter (6-21 day) and grower (22-42 day) periods, respectively. Subsequently, all groups received a diet containing the same nutrients during the finisher period (43-49 day). The results showed that, under heat stress conditions, average daily gain and feed conversion ratio and performance index from day 6 to 49, breast and thigh yields and antibody titer against Newcastle disease in the birds fed diets 85S, 70S and 85SG were similar to those of birds fed control diet, whereas feeding diets 70S85G and 70SG significantly decreased the values of above-mentioned parameters. Additionally, diets 85S, 70S and 85SG significantly decreased mortality rate and heterophil:lymphocyte ratio compared with the control diet. In conclusion, the results indicate that supplementation of methionine to diets 85S, 70S and 85SG, and then re-feeding with a conventional diet is an effective tool to maintain productive performance and to improve health indices and heat resistance in broilers under high ambient temperature conditions.

Designing sgRNAs with CRISPy web

DEFF Research Database (Denmark)

Blin, Kai; Lee, Sang Yup; Weber, Tilmann

2017-01-01

Tilmann Weber’s group at the Novo Nordisk Foundation Center for Biosustainability developed a user-friendly, web server implementation of the sgRNA prediction software, CRISPy, for non-computer scientists.......Tilmann Weber’s group at the Novo Nordisk Foundation Center for Biosustainability developed a user-friendly, web server implementation of the sgRNA prediction software, CRISPy, for non-computer scientists....

Patterns of gene expression associated with recovery and injury in heat-stressed rats.

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Stallings, Jonathan D; Ippolito, Danielle L; Rakesh, Vineet; Baer, Christine E; Dennis, William E; Helwig, Bryan G; Jackson, David A; Leon, Lisa R; Lewis, John A; Reifman, Jaques

2014-12-03

The in vivo gene response associated with hyperthermia is poorly understood. Here, we perform a global, multiorgan characterization of the gene response to heat stress using an in vivo conscious rat model. We heated rats until implanted thermal probes indicated a maximal core temperature of 41.8°C (Tc,Max). We then compared transcriptomic profiles of liver, lung, kidney, and heart tissues harvested from groups of experimental animals at Tc,Max, 24 hours, and 48 hours after heat stress to time-matched controls kept at an ambient temperature. Cardiac histopathology at 48 hours supported persistent cardiac injury in three out of six animals. Microarray analysis identified 78 differentially expressed genes common to all four organs at Tc,Max. Self-organizing maps identified gene-specific signatures corresponding to protein-folding disorders in heat-stressed rats with histopathological evidence of cardiac injury at 48 hours. Quantitative proteomics analysis by iTRAQ (isobaric tag for relative and absolute quantitation) demonstrated that differential protein expression most closely matched the transcriptomic profile in heat-injured animals at 48 hours. Calculation of protein supersaturation scores supported an increased propensity of proteins to aggregate for proteins that were found to be changing in abundance at 24 hours and in animals with cardiac injury at 48 hours, suggesting a mechanistic association between protein misfolding and the heat-stress response. Pathway analyses at both the transcript and protein levels supported catastrophic deficits in energetics and cellular metabolism and activation of the unfolded protein response in heat-stressed rats with histopathological evidence of persistent heat injury, providing the basis for a systems-level physiological model of heat illness and recovery.

Root cause analysis of SG tube leakage at Fessenheim unit 2 in 2008

International Nuclear Information System (INIS)

Berger, J.; Deotto, G.; Mathon, C.; Madurel, A.; Pitner, P.; Gay, N.; Guivarch, M.

2015-01-01

In February 2008, a primary-to-secondary leak caused an unscheduled shutdown at Fessenheim Unit 2 NPP. A circumferential crack was observed just above the top support plate of Row 12 Column 62 U-bend tube on Steam Generator (SG) number 3, which has been attributed to high cycle fatigue. This tube was pulled out in 2011, just before the SG replacement at the third decenal outage, in order to perform exhaustive metallurgical investigations. The destructive examinations revealed that the circumferential crack (70 degrees of extension) was due to high cycle fatigue, with several external initiation areas associated with the presence of small piles of Intergranular Attack (IGA) (600 MA tube) and with very low stress intensity factors ΔK (close to the non-propagating threshold region). This paper complements the metallurgical investigations by carrying out numerical analyses (thermal-hydraulic computation, fluid-elastic instability evaluation, tube vibratory response analysis and fatigue evaluation). The first objective of the study is to attempt to clarify the effect of IGA and the role of several competing factors that could be involved in the tube vibration induced fatigue failure. From these results, a root cause analysis of the R12C62 tube fatigue failure is then provided. It appears that a combination of various factors led to the failure of the tube

Excessive endoplasmic reticulum stress and decreased neuroplasticity-associated proteins in prefrontal cortex of obese rats and the regulatory effects of aerobic exercise.

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Li, Feng; Liu, Bei Bei; Cai, Ming; Li, Jing Jing; Lou, Shu-Jie

2018-04-06

Studies have shown high fat diet induced obesity may cause cognition impairment and down-regulation of neuroplasticity-associated proteins, while aerobic exercise could improve that damage. Endoplasmic reticulum stress (ERS) has been reported to play a key role in regulating neuroplasticity-associated proteins expression, folding and post-translational modification in hippocampus of obese rodent models, however, the effects of ERS on neuroplasticity-associated proteins and possible underlying mechanisms in prefrontal cortex are not fully clear. In order to clarify changes of neuroplasticity-associated proteins and ERS in the prefrontal cortex of obese rats, male SD rats were fed on high fat diet for 8 weeks to establish the obese model. Then, 8 weeks of aerobic exercise treadmill intervention was arranged for the obese rats. Results showed that high fat diet induced obesity caused hyperlipidemia, and significantly promoted FATP1 expression in the prefrontal cortex, meanwhile, we found up-regulation of GRP78, p-PERK, p-eIF2α, caspase-12, CHOP, and Bax/Bcl-2, reflecting the activation of ERS and ERS-mediated apoptosis. Moreover, reduced BDNF and SYN was found in obese rats. However, FATP1, GRP78, p-PERK, p-eIF2α, caspase-12, CHOP, and Bax/Bcl-2 expressions were obviously reversed by aerobic exercise intervention. These results suggested that dietary obesity could induce Prefrontal ERS in SD rats and excessive ERS may play a critical role in decreasing the levels of neuroplasticity-associated proteins. Moreover, aerobic exercise could relieve ERS, thus promoted the expression of neuroplasticity-associated proteins. Copyright © 2018. Published by Elsevier Inc.

A Nucleocytoplasmic Shuttling Protein in Oxidative Stress Tolerance

Energy Technology Data Exchange (ETDEWEB)

Ow, David W.; Song, Wen

2003-03-26

Plants for effective extraction of toxic metals and radionuclides must tolerate oxidative stress. To identify genes that enhance oxidative stress tolerance, an S. pombe cDNA expression plasmid library was screened for the ability to yield hypertolerant colonies. Here, we report on the properties of one gene that confers hypertolerance to cadmium and oxidizing chemicals. This gene appears to be conserved in other organisms as homologous genes are found in human, mouse, fruitfly and Arabidopsis. The fruitfly and Arabidopsis genes likewise enhance oxidative stress tolerance in fission yeast. During oxidative stress, the amount of mRNA does not change, but protein fusions to GFP relocate from the cytoplasm to the nucleus. The same pattern is observed with the Arabidopsis homologue-GFP fusion protein. This behavior suggests a signaling role in oxidative stress tolerance and these conserved proteins may be targets for engineering stress tolerant plants for phytoremediation.

Schizosaccharomyces pombe Homologs of Human DJ-1 Are Stationary Phase-Associated Proteins That Are Involved in Autophagy and Oxidative Stress Resistance.

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Yang Su

Full Text Available The Parkinson's disease protein DJ-1 is involved in various cellular functions including detoxification of dicarbonyl compounds, autophagy and oxidative stress response. DJ-1 homologs are widely found in both prokaryotes and eukaryotes, constituting a superfamily of proteins that appear to be involved in stress response. Schizosaccharomyces pombe contains six DJ-1 homologs, designated Hsp3101-Hsp3105 and Sdj1 (previously named SpDJ-1. Here we show that deletion of any one of these six genes somehow affects autophagy during prolonged stationary phase. Furthermore, deletions of each of these DJ-1 homologs result in reduced stationary phase survival. Deletion of sdj1 also increases the sensitivity of stationary-phase cells to oxidative stress induced by hydrogen peroxide (H2O2 whereas overexpression of sdj1 has the opposite effect. Consistent with their role in stationary phase, expression of hsp3101, hsp3102, hsp3105 and sdj1, and to a lesser extent hsp3103 and hsp3104, is increased in stationary phase. The induction of hsp3101, hsp3102, hsp3105 and sdj1 involves the Sty1-regulated transcription factor Atf1 but not the transcription factor Pap1. Our results firmly establish that S. pombe homologs of DJ-1 are stationary-phase associated proteins and are likely involved in autophagy and antioxidant defense in stationary phase of S. pombe cells.

Development of a graphical user interface for sgRNAcas9 and its application.

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Zhao, Chang-zhi; Zhang, Yi; Li, Guang-lei; Chen, Ji-liang; Li, Jing-Jin; Ren, Rui-min; Ni, Pan; Zhao, Shu-hong; Xie, Sheng-song

2015-10-01

The CRISPR/Cas9 genome editing technique is a powerful tool for researchers. However, off-target effects of the Cas9 nuclease activity is a recurrent concern of the CRISPR system. Thus, designing sgRNA (single guide RNA) with minimal off-target effects is very important. sgRNAcas9 is a software package, which can be used to design sgRNA and to evaluate potential off-target cleavage sites. In this study, a graphical user interface for sgRNAcas9 was developed using the Java programming language. In addition, off-target effect for sgRNAs was evaluated according to mismatched number and "seed sequence" specification. Moreover, sgRNAcas9 software was used to design 34 124 sgRNAs, which can target 4691 microRNA (miRNA) precursors from human, mouse, rat, pig, and chicken. In particular, the off-target effect of a sgRNA targeting to human miR-206 precursor was analyzed, and the on/off-target activity of this sgRNA was validated by T7E1 assay in vitro. Taken together, these data showed that the interface can simplify the usage of the sgRNAcas9 program, which can be used to design sgRNAs for the majority of miRNA precursors. We also found that the GC% of those sgRNAs ranged from 40% to 60%. In summary, the sgRNAcas9 software can be easily used to design sgRNA with minimal off-target effects for any species. The software can be downloaded from BiooTools website (http://www.biootools.com/).

Inhibition of Ribosome Recruitment Induces Stress Granule Formation Independently of Eukaryotic Initiation Factor 2α Phosphorylation

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Mazroui, Rachid; Sukarieh, Rami; Bordeleau, Marie-Eve; Kaufman, Randal J.; Northcote, Peter; Tanaka, Junichi; Gallouzi, Imed

2006-01-01

Cytoplasmic aggregates known as stress granules (SGs) arise as a consequence of cellular stress and contain stalled translation preinitiation complexes. These foci are thought to serve as sites of mRNA storage or triage during the cell stress response. SG formation has been shown to require induction of eukaryotic initiation factor (eIF)2α phosphorylation. Herein, we investigate the potential role of other initiation factors in this process and demonstrate that interfering with eIF4A activity, an RNA helicase required for the ribosome recruitment phase of translation initiation, induces SG formation and that this event is not dependent on eIF2α phosphorylation. We also show that inhibition of eIF4A activity does not impair the ability of eIF2α to be phosphorylated under stress conditions. Furthermore, we observed SG assembly upon inhibition of cap-dependent translation after poliovirus infection. We propose that SG modeling can occur via both eIF2α phosphorylation-dependent and -independent pathways that target translation initiation. PMID:16870703

Inhibition of ribosome recruitment induces stress granule formation independently of eukaryotic initiation factor 2alpha phosphorylation.

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Mazroui, Rachid; Sukarieh, Rami; Bordeleau, Marie-Eve; Kaufman, Randal J; Northcote, Peter; Tanaka, Junichi; Gallouzi, Imed; Pelletier, Jerry

2006-10-01

Cytoplasmic aggregates known as stress granules (SGs) arise as a consequence of cellular stress and contain stalled translation preinitiation complexes. These foci are thought to serve as sites of mRNA storage or triage during the cell stress response. SG formation has been shown to require induction of eukaryotic initiation factor (eIF)2alpha phosphorylation. Herein, we investigate the potential role of other initiation factors in this process and demonstrate that interfering with eIF4A activity, an RNA helicase required for the ribosome recruitment phase of translation initiation, induces SG formation and that this event is not dependent on eIF2alpha phosphorylation. We also show that inhibition of eIF4A activity does not impair the ability of eIF2alpha to be phosphorylated under stress conditions. Furthermore, we observed SG assembly upon inhibition of cap-dependent translation after poliovirus infection. We propose that SG modeling can occur via both eIF2alpha phosphorylation-dependent and -independent pathways that target translation initiation.

Genome-wide analysis of uncapped mRNAs under heat stress in Arabidopsis

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Emilio Gutierrez-Beltran

2015-09-01

Full Text Available Recently, we have showed that Tudor Staphylococcal Nuclease (TSN or Tudor-SN proteins (TSN1 and TSN2 are localized in cytoplasmic messenger ribonucleoprotein (mRNP complexes called stress granules (SG and processing bodies (PB under heat stress in Arabidopsis. One of the primary functions of these mRNP complexes is mRNA decay, which generates uncapped mRNAs by the action of endonucleases and decapping enzymes (Thomas et al., 2011 [1]. In order to figure out whether TSN proteins could be implicated in mRNA decay, we isolated uncapped and total mRNAs of Wild type (WT; Col and Ler and TSN double knock-out (tsn1tsn2 seedlings grown under heat stress (39 °C for 40 min and control (23 °C conditions. Here, we provide the experimental procedure to reproduce the results (NCBI GEO accession number GSE63522 published by Gutierrez-Beltran et al. (2015 in The Plant Cell [2].

Hypoxic-induced stress protein expression in rat cardiac myocytes

International Nuclear Information System (INIS)

Howard, G.; Geoghegan, T.E.

1986-01-01

Mammalian stress proteins can be induced in cells and tissues exposed to a variety of conditions including hyperthermia and diminished O 2 supply. The authors have previously shown that the expression of three stress proteins (71, 85, and 95 kDa) was induced in cardiac tissue from mice exposed to hypoxic conditions. The expression of mRNAs coding for the 85 and 95 kDa proteins increase with time of exposure to hypoxia, while the mRNA coding for the 71 kDa protein is transiently induced. The authors extended these studies to investigate the expression of stress proteins in isolated rat cardiac myocytes. Freshly prepared myocytes were exposed to control, hypoxic, anoxic, or heat-shock environments for up to 16 h. The proteins were then labeled for 6 hours with [ 35 S]methionine. Analysis of the solubilized proteins by SDS-PAGE and autoradiography showed that there was a 6-fold increase in synthesis of the 85 kDa protein upon exposure to hypoxia but not heat-shock conditions. The 71 kDa protein was present at high levels in both control and treated myocyte protein preparations, and presumably had been induced during the isolation procedure. Total RNA isolated from intact rat heart and isolated myocytes was compared by cell-free translation analysis and showed induction of RNAs coding for several stress proteins in the myocyte preparation. The induced proteins at 85 and 95 kDa have molecular weights similar to reported cell stress and/or glucose-regulated proteins

Dispersant Application during SG Wet Layup at SK Unit 1

Energy Technology Data Exchange (ETDEWEB)

Kwon, Hyukchul; Lee, Dooho; Sung, Kibang [KHNP Central Institute, Daejeon (Korea, Republic of)

2014-05-15

The corrosion products in the feedwater are deposited onto the steam generators (SGs) despite the effort to control them within limit of impurity. This deposit is one of causes for occurrence of SCC (Stress Corrosion Cracking), water level fluctuation and further corrosion of SGs. To minimize corrosion and remove deposit, the nuclear power plants apply high pH to the secondary system and SG chemical cleaning, respectively. But these methods can be costly and carry risks of extended outages or incomplete cleaning. Another method is an on-line dispersant application. The role of dispersant is to make deposit suspended in the SGs. Then, the suspended deposit is discharged to the blowdown system. The iron removal is increased in the blowdown system during the dispersant application. Additional significant benefit in the form of reduced corrosion product transport may be obtained through applying dispersant in the SGs wet lay operational mode. This method helps to reduce the total SGs loading without affecting critical outage activities and with minimal additional effort on the part of the utilities. This study provides the results of the dispersant application trial during the SG wet layup at SK Unit 1. As the PAA concentrations were increased, the corrosion rates of Alloy 690 and SA 106 Gr.B were increased. The corrosion rate of Alloy 690 was 2 times less than that of SA 106 Gr.B at 100 ppm of PAA based on the electrochemical experimental. There were no significant feasibility problems with application of PAA during the SG wet layup. The reasonable estimation of the additional mass removed by the presence of PAA during SGs wet layup is 460 g. The iron removal depended on PAA concentration injected based on the comparative results of the SK Unit 1 and TMI-1. It is expected that injection of PAA into the SG result in a significant decrease in the amount of iron transported to the SGs during the startup.

Recent progress in SG level control in French PWR plants

International Nuclear Information System (INIS)

Parry, A.; Petetrot, J.F.; Vivier, M.J.

1985-10-01

Controlling the steam generator (SG) level is of major importance in a large PWR plant. This has led to extensive work on SG computer models. This paper presents results of the comparison between calculations and tests on the first four-loop plant in France. Four-loop plants started up after 1985 will be equipped with digital instead of analog controllers. A new SG level control has been designed and then optimised using the validated SG model. A prototype of this new system has been successfully tested on a three-loop plant. 4 refs

Network Mechanisms of Clinical Response to Transcranial Magnetic Stimulation in Posttraumatic Stress Disorder and Major Depressive Disorder.

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Philip, Noah S; Barredo, Jennifer; van 't Wout-Frank, Mascha; Tyrka, Audrey R; Price, Lawrence H; Carpenter, Linda L

2018-02-01

Repetitive transcranial magnetic stimulation (TMS) therapy can modulate pathological neural network functional connectivity in major depressive disorder (MDD). Posttraumatic stress disorder is often comorbid with MDD, and symptoms of both disorders can be alleviated with TMS therapy. This is the first study to evaluate TMS-associated changes in connectivity in patients with comorbid posttraumatic stress disorder and MDD. Resting-state functional connectivity magnetic resonance imaging was acquired before and after TMS therapy in 33 adult outpatients in a prospective open trial. TMS at 5 Hz was delivered, in up to 40 daily sessions, to the left dorsolateral prefrontal cortex. Analyses used a priori seeds relevant to TMS, posttraumatic stress disorder, or MDD (subgenual anterior cingulate cortex [sgACC], left dorsolateral prefrontal cortex, hippocampus, and basolateral amygdala) to identify imaging predictors of response and to evaluate clinically relevant changes in connectivity after TMS, followed by leave-one-out cross-validation. Imaging results were explored using data-driven multivoxel pattern activation. More negative pretreatment connectivity between the sgACC and the default mode network predicted clinical improvement, as did more positive amygdala-to-ventromedial prefrontal cortex connectivity. After TMS, symptom reduction was associated with reduced connectivity between the sgACC and the default mode network, left dorsolateral prefrontal cortex, and insula, and reduced connectivity between the hippocampus and the salience network. Multivoxel pattern activation confirmed seed-based predictors and correlates of treatment outcomes. These results highlight the central role of the sgACC, default mode network, and salience network as predictors of TMS response and suggest their involvement in mechanisms of action. Furthermore, this work indicates that there may be network-based biomarkers of clinical response relevant to these commonly comorbid disorders

Imbalance of heterologous protein folding and disulfide bond formation rates yields runaway oxidative stress

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Tyo Keith EJ

2012-03-01

Full Text Available Abstract Background The protein secretory pathway must process a wide assortment of native proteins for eukaryotic cells to function. As well, recombinant protein secretion is used extensively to produce many biologics and industrial enzymes. Therefore, secretory pathway dysfunction can be highly detrimental to the cell and can drastically inhibit product titers in biochemical production. Because the secretory pathway is a highly-integrated, multi-organelle system, dysfunction can happen at many levels and dissecting the root cause can be challenging. In this study, we apply a systems biology approach to analyze secretory pathway dysfunctions resulting from heterologous production of a small protein (insulin precursor or a larger protein (α-amylase. Results HAC1-dependent and independent dysfunctions and cellular responses were apparent across multiple datasets. In particular, processes involving (a degradation of protein/recycling amino acids, (b overall transcription/translation repression, and (c oxidative stress were broadly associated with secretory stress. Conclusions Apparent runaway oxidative stress due to radical production observed here and elsewhere can be explained by a futile cycle of disulfide formation and breaking that consumes reduced glutathione and produces reactive oxygen species. The futile cycle is dominating when protein folding rates are low relative to disulfide bond formation rates. While not strictly conclusive with the present data, this insight does provide a molecular interpretation to an, until now, largely empirical understanding of optimizing heterologous protein secretion. This molecular insight has direct implications on engineering a broad range of recombinant proteins for secretion and provides potential hypotheses for the root causes of several secretory-associated diseases.

Effect of temperature stress on protein methyl esters

International Nuclear Information System (INIS)

Welch, W.; Kracaw, K.

1986-01-01

Protein methyl esters have been implicated in a number of physiological processes. They have measured the effect of temperature stress on the levels of protein methyl esters in the mesophilic fungus Penicillium chrysogenum (PCPS) and the thermophilic fungus P. duponti (PD). PD and PCPS were incubated with [methyl- 3 H]methionine. The mycelia were collected by filtration, frozen in liquid nitrogen and ground to a fine powder. The nitrogen powder was extracted with either phosphate buffer or with SDS, glycerol, phosphate, 2-mercaptoethanol. Insoluble material was removed by centrifugation. The supernatants were assayed for protein methyl esters. The released [ 3 H]methanol was extracted into toluene:isoamyl alcohol (3:2) and quantitated by liquid scintillation. The production of volatile methanol was confirmed by use of Conway diffusion cells. Soluble proteins accounted for about one-fourth of the total protein methyl ester extracted by SDS. In PCPS, the SDS extracted proteins have about three times the level of esterification of the soluble proteins whereas in PD there is little difference between soluble and SDS extracted protein. The level of protein esterification in PD is about one-tenth that observed in PCPS. Temperature stress caused large changes in the level of protein esterification. The data suggest protein methyl esters may contribute to the adaptation to environmental stress

The stress response system of proteins: Implications for bioreactor scaleup

Science.gov (United States)

Goochee, Charles F.

1988-01-01

Animal cells face a variety of environmental stresses in large scale bioreactors, including periodic variations in shear stress and dissolved oxygen concentration. Diagnostic techniques were developed for identifying the particular sources of environmental stresses for animal cells in a given bioreactor configuration. The mechanisms by which cells cope with such stresses was examined. The individual concentrations and synthesis rates of hundreds of intracellular proteins are affected by the extracellular environment (medium composition, dissolved oxygen concentration, ph, and level of surface shear stress). Techniques are currently being developed for quantifying the synthesis rates and concentrations of the intracellular proteins which are most sensitive to environmental stress. Previous research has demonstrated that a particular set of stress response proteins are synthesized by mammalian cells in response to temperature fluctuations, dissolved oxygen deprivation, and glucose deprivation. Recently, it was demonstrated that exposure of human kidney cells to high shear stress results in expression of a completely distinct set of intracellular proteins.

Molecular elucidation of a new allelic variation at the Sg-5 gene associated with the absence of group A saponins in wild soybean.

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Sundaramoorthy, Jagadeesh; Park, Gyu Tae; Mukaiyama, Kyosuke; Tsukamoto, Chigen; Chang, Jeong Ho; Lee, Jeong-Dong; Kim, Jeong Hoe; Seo, Hak Soo; Song, Jong Tae

2018-01-01

In soybean, triterpenoid saponin is one of the major secondary metabolites and is further classified into group A and DDMP saponins. Although they have known health benefits for humans and animals, acetylation of group A saponins causes bitterness and gives an astringent taste to soy products. Therefore, several studies are being conducted to eliminate acetylated group A saponins. Previous studies have isolated and characterized the Sg-5 (Glyma.15g243300) gene, which encodes the cytochrome P450 72A69 enzyme and is responsible for soyasapogenol A biosynthesis. In this study, we elucidated the molecular identity of a novel mutant of Glycine soja, 'CWS5095'. Phenotypic analysis using TLC and LC-PDA/MS/MS showed that the mutant 'CWS5095' did not produce any group A saponins. Segregation analysis showed that the absence of group A saponins is controlled by a single recessive allele. The locus was mapped on chromosome 15 (4.3 Mb) between Affx-89193969 and Affx-89134397 where the previously identified Glyma.15g243300 gene is positioned. Sequence analysis of the coding region for the Glyma.15g243300 gene revealed the presence of four SNPs in 'CWS5095' compared to the control lines. One of these four SNPs (G1127A) leads to the amino acid change Arg376Lys in the EXXR motif, which is invariably conserved among the CYP450 superfamily proteins. Co-segregation analysis showed that the missense mutation (Arg376Lys) was tightly linked with the absence of group A saponins in 'CWS5095'. Even though Arg and Lys have similar chemical features, the 3D modelled protein structure indicates that the replacement of Arg with Lys may cause a loss-of-function of the Sg-5 protein by inhibiting the stable binding of a heme cofactor to the CYP72A69 apoenzyme.

Laser performance upgrade for precise ICF experiment in SG-Ⅲ laser facility

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Wanguo Zheng

2017-09-01

Full Text Available The SG-Ⅲ laser facility (SG-Ⅲ is the largest laser driver for inertial confinement fusion (ICF researches in China, which has 48 beamlines and can deliver 180 kJ ultraviolet laser energy in 3 ns. In order to meet the requirements of precise physics experiments, some new functionalities need to be added to SG-Ⅲ and some intrinsic laser performances need upgrade. So at the end of SG-Ⅲ's engineering construction, the 2-year laser performance upgrade project started. This paper will introduce the newly added functionalities and the latest laser performance of SG-Ⅲ. With these function extensions and performance upgrade, SG-Ⅲ is now fully prepared for precise ICF experiments and solidly paves the way towards fusion ignition.

The responses of antioxidant system in bitter melon, sponge gourd, and winter squash under flooding and chilling stresses

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Do, Tuong Ha; Nguyen, Hoang Chinh; Lin, Kuan-Hung

2018-04-01

The objective of this paper was to review the responses of antioxidant system and physiological parameters of bitter melon (BM), sponge gourd (SG), and winter squash (WS) under waterlogged and low temperature conditions. The BM and SG plants were subjected to 0-72 h flooding treatments, and BM and WS plants were exposed to chilling at 12/7 °C (day/night) for 0-72 h. Different genotypes responded differently to environmental stress according to their various antioxidant system and physiological parameters. Increased ascorbate peroxidase (APX) and superoxide dismutase (SOD) activities provided SG and WS plants with increased waterlogging and chilling stress tolerance, respectively, compared to BM plants. The APX gene from SG and the SOD gene from WS were then cloned, and the regulation of APX and SOD gene expressions under flooding and chilling stress, respectively, were also measured. Increased expression of APX and SOD genes was accompanied by the increased activity of the enzyme involved in detoxifying reactive oxygen species (ROS) in response to those stresses. Both APX and SOD activities can be used for selecting BM lines with the best tolerances to water logging and chilling stresses.

Stress responses during ageing: molecular pathways regulating protein homeostasis.

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Kyriakakis, Emmanouil; Princz, Andrea; Tavernarakis, Nektarios

2015-01-01

The ageing process is characterized by deterioration of physiological function accompanied by frailty and ageing-associated diseases. The most broadly and well-studied pathways influencing ageing are the insulin/insulin-like growth factor 1 signaling pathway and the dietary restriction pathway. Recent studies in diverse organisms have also delineated emerging pathways, which collectively or independently contribute to ageing. Among them the proteostatic-stress-response networks, inextricably affect normal ageing by maintaining or restoring protein homeostasis to preserve proper cellular and organismal function. In this chapter, we survey the involvement of heat stress and endoplasmic reticulum stress responses in the regulation of longevity, placing emphasis on the cross talk between different response mechanisms and their systemic effects. We further discuss novel insights relevant to the molecular pathways mediating these stress responses that may facilitate the development of innovative interventions targeting age-related pathologies such as diabetes, cancer, cardiovascular and neurodegenerative diseases.

Field experience with KWU SG chemical cleaning process

International Nuclear Information System (INIS)

Odar, S.

1989-01-01

The ingress of corrosion products into PWR steam generators (SG's) their deposition and the subsequent concentration of salt impurities can induce a variety of mechanisms for corrosion attack on SG tubing. Already, some plants have had to replace their steam generators due to severe corrosion damage and others are seriously considering the same costly action in the near future. One of the most effective ways to counteract corrosion mechanisms and thus to reduce the likelihood of SG replacement becoming necessary is to clean the SG's and to keep them clean. For many years, the industry has been involved in developing different types of cleaning techniques. Among these, chemical cleaning has been shown to be especially effective. In this article, the KWU chemical cleaning process, for which there is considerable application experience, is described. The results of field applications will be presented together with material compatibility data and information on cleaning effectiveness. (author)

Fluoride induces endoplasmic reticulum stress and inhibits protein synthesis and secretion.

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Sharma, Ramaswamy; Tsuchiya, Masahiro; Bartlett, John D

2008-09-01

Exposure to excessive amounts of fluoride (F(-)) causes dental fluorosis in susceptible individuals; however, the mechanism of F(-)-induced toxicity is unclear. Previously, we have shown that high-dose F(-) activates the unfolded protein response (UPR) in ameloblasts that are responsible for dental enamel formation. The UPR is a signaling pathway responsible for either alleviating endoplasmic reticulum (ER) stress or for inducing apoptosis of the stressed cells. In this study we determined if low-dose F(-) causes ER stress and activates the UPR, and we also determined whether F(-) interferes with the secretion of proteins from the ER. We stably transfected the ameloblast-derived LS8 cell line with secreted alkaline phosphatase (SEAP) and determined activity and localization of SEAP and F(-)-mediated induction of UPR proteins. Also, incisors from mice given drinking water containing various concentrations of F(-) were examined for eucaryotic initiation factor-2, subunit alpha (eIF2alpha) phosphorylation. We found that F(-) decreases the extracellular secretion of SEAP in a linear, dose-dependent manner. We also found a corresponding increase in the intracellular accumulation of SEAP after exposure to F(-). These changes are associated with the induction of UPR proteins such as the molecular chaperone BiP and phosphorylation of the UPR sensor PKR-like ER kinase, and its substrate, eIF2alpha. Importantly, F(-)-induced phosphorylation of eIF2alphawas confirmed in vivo. These data suggest that F(-) initiates an ER stress response in ameloblasts that interferes with protein synthesis and secretion. Consequently, ameloblast function during enamel development may be impaired, and this may culminate in dental fluorosis.

Does low-protein diet improve broiler performance under heat stress conditions?

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RL Furlan

2004-06-01

Full Text Available Nutrition for broilers under high temperatures is extremely important for brazilian broiler chicken industry because the amounts of consumed nutrients and environmental temperature have great effects on bird performance and carcass quality. Among diet nutrients, protein has the highest heat increment; thus, during many years, diets with low protein level were recommended in order to reduce heat production in broiler chickens under heat stress. However, reports have shown that low-protein diets have negative effects on broiler performance when environmental temperature is high, because during heat stress, low food intake associated to a low diet protein induce amino acid deficiencies. Other studies have shown that broilers fed low-protein diets increase their energy requirement for maintenance with higher heat production. Thus, with the growth of broiler industry in tropical areas more challenges need to be faced by the farmers. So, both the ambient and nutritional conditions ought to be well managed to avoid negative effects on poultry production once they can affect the metabolism (body heat production under low temperature and body heat dissipation under high temperature with consequence on poultry performance (meat and eggs.

Stress dependence of microstructures in experimentally deformed calcite

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Platt, John P.; De Bresser, J. H. P.

2017-12-01

Optical measurements of microstructural features in experimentally deformed Carrara marble help define their dependence on stress. These features include dynamically recrystallized grain size (Dr), subgrain size (Sg), minimum bulge size (Lρ), and the maximum scale length for surface-energy driven grain-boundary migration (Lγ). Taken together with previously published data Dr defines a paleopiezometer over the range 15-291 MPa and temperature over the range 500-1000 °C, with a stress exponent of -1.09 (CI -1.27 to -0.95), showing no detectable dependence on temperature. Sg and Dr measured in the same samples are closely similar in size, suggesting that the new grains did not grow significantly after nucleation. Lρ and Lγ measured on each sample define a relationship to stress with an exponent of approximately -1.6, which helps define the boundary between a region of dominant strain-energy-driven grain-boundary migration at high stress, from a region of dominant surface-energy-driven grain-boundary migration at low stress.

The Parkinson’s disease-associated protein α-synuclein disrupts stress signaling – a possible implication for methamphetamine use?

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Shaoxiao Wang

2014-03-01

Full Text Available The human neuronal protein α-synuclein (α-syn has been linked by a plethora of studies as a causative factor in sporadic Parkinson’s disease (PD. To speed the pace of discovery about the biology and pathobiology of α-syn, organisms such as yeast, worms, and flies have been used to investigate the mechanisms by which elevated levels of α-syn are toxic to cells and to screen for drugs and genes that suppress this toxicity. We recently reported [Wang et al. Proc. Natl. Acad. Sci.(2012 109: 16119–16124] that human α-syn, at high expression levels, disrupts stress-activated signal transduction pathways in both yeast and human neuroblastoma cells. Disruption of these signaling pathways ultimately leads to vulnerability to stress and to cell death. Here we discuss how the disruption of cell signaling by α-syn may have relevance to the parkinsonism that is associated with the abuse of the drug methamphetamine (meth.

In Silico Meets In Vivo: Towards Computational CRISPR-Based sgRNA Design.

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Chuai, Guo-Hui; Wang, Qi-Long; Liu, Qi

2017-01-01

CRISPR-based genome editing has been widely implemented in various cell types. In silico single guide RNA (sgRNA) design is a key step for successful gene editing using the CRISPR system, and continuing efforts are aimed at refining in silico sgRNA design with high on-target efficacy and reduced off-target effects. Many sgRNA design tools are available, but careful assessments of their application scenarios and performance benchmarks across different types of genome-editing data are needed. Efficient in silico models can be built that integrate current heterogeneous genome-editing data to derive unbiased sgRNA design rules and identify key features for improving sgRNA design. Comprehensive evaluation of on-target and off-target effects of sgRNA will allow more precise genome editing and gene therapies using the CRISPR system. Copyright © 2016 Elsevier Ltd. All rights reserved.

Pregnancy Associated Plasma Protein-A in Type 2 Diabetic Patient with Peripheral Neuropathy

International Nuclear Information System (INIS)

Nosseir, N.M.

2011-01-01

Metabolic changes induced by hyperglycemia lead to dysregulation of cytokines control, subclinical inflammation together with oxidative stress associated with diabetes. The aim of this study is to correlate the role of type 2 diabetic neuropathy on serum pregnancy associated plasma protein-A,interleukin-6 and c-reactive protein .The results denoted that both pregnancy associated plasma protein-A and interleukin-6 were significantly increased in those patients with diabetic neuropathy compared with those without neuropathy but while c-reactive proteins showed significant differences between the three groups, the results lead to the conclusion that PAPP-A,IL-6 are useful tests in monitoring the neuropathic complications associated with type 2 diabetes

Comparative Evaluation of Common Savannahgrass on a Range of Soils Subjected to Different Stresses I: Productivity and Quality

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Raymond Springer

2014-04-01

Full Text Available Turfgrass growth, performance and quality are affected by abiotic stress factors and are of primary concern for persons managing turfgrass areas under seasonal tropical climates. Under these conditions, common Savannahgrass (SG may have a performance advantage over imported hybrid turfgrasses. A greenhouse study was conducted to comparatively evaluate the performance of tropical turfgrasses exposed to water and compaction related stresses across a range of soils, with or without the addition of a surface sand layer. Turfgrass productivity and quality was monitored over a four-month growth period. Clipping yield (CY was lower at the higher compaction effort for all turfgrasses, but across all stresses, drought (D and waterlogging (WL resulted in lower CY. Values were significantly lower under D. SG had the highest clipping yield across all soils. The chlorophyll index (CI was lower for all turfgrasses under water-induced stress compared to compaction stresses. SG had a significantly higher CI across all stress treatments. Correlation analysis showed a positive (r² = 0.420 and significant (p < 0.05 relationship between CY and CI. Similar to CI, stress type influenced turfgrass visual quality (VQ, with D stress, resulting in the lowest VQ rating among turfgrasses. Bermudagrass (BG had the lowest VQ across all stress treatments, whilst, comparatively, Zoysiagrass (ZG had significantly higher VQ under high compaction (HC, low compaction (LC and WL stress. Overall, SG showed a higher level of tolerance to applied stresses and warrants greater attention as a potential turfgrass under tropical conditions.

CRISPRscan: designing highly efficient sgRNAs for CRISPR/Cas9 targeting in vivo

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Moreno-Mateos, Miguel A.; Vejnar, Charles E.; Beaudoin, Jean-Denis; Fernandez, Juan P.; Mis, Emily K.; Khokha, Mustafa K.; Giraldez, Antonio J.

2015-01-01

CRISPR/Cas9 technology provides a powerful system for genome engineering. However, variable activity across different single guide RNAs (sgRNAs) remains a significant limitation. We have analyzed the molecular features that influence sgRNA stability, activity and loading into Cas9 in vivo. We observe that guanine enrichment and adenine depletion increase sgRNA stability and activity, while loading, nucleosome positioning and Cas9 off-target binding are not major determinants. We additionally identified truncated and 5′ mismatch-containing sgRNAs as efficient alternatives to canonical sgRNAs. Based on these results, we created a predictive sgRNA-scoring algorithm (CRISPRscan.org) that effectively captures the sequence features affecting Cas9/sgRNA activity in vivo. Finally, we show that targeting Cas9 to the germ line using a Cas9-nanos-3′-UTR fusion can generate maternal-zygotic mutants, increase viability and reduce somatic mutations. Together, these results provide novel insights into the determinants that influence Cas9 activity and a framework to identify highly efficient sgRNAs for genome targeting in vivo. PMID:26322839

Identification of indicator proteins associated with flooding injury in soybean seedlings using label-free quantitative proteomics.

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Nanjo, Yohei; Nakamura, Takuji; Komatsu, Setsuko

2013-11-01

Flooding injury is one of the abiotic constraints on soybean growth. An experimental system established for evaluating flooding injury in soybean seedlings indicated that the degree of injury is dependent on seedling density in floodwater. Dissolved oxygen levels in the floodwater were decreased by the seedlings and correlated with the degree of injury. To understand the molecular mechanism responsible for the injury, proteomic alterations in soybean seedlings that correlated with severity of stress were analyzed using label-free quantitative proteomics. The analysis showed that the abundance of proteins involved in cell wall modification, such as polygalacturonase inhibitor-like and expansin-like B1-like proteins, which may be associated with the defense system, increased dependence on stress at both the protein and mRNA levels in all organs during flooding. The manner of alteration in abundance of these proteins was distinct from those of other responsive proteins. Furthermore, proteins also showing specific changes in abundance in the root tip included protein phosphatase 2A subunit-like proteins, which are possibly involved in flooding-induced root tip cell death. Additionally, decreases in abundance of cell wall synthesis-related proteins, such as cinnamyl-alcohol dehydrogenase and cellulose synthase-interactive protein-like proteins, were identified in hypocotyls of seedlings grown for 3 days after flooding, and these proteins may be associated with suppression of growth after flooding. These flooding injury-associated proteins can be defined as indicator proteins for severity of flooding stress in soybean.

Financial strain is associated with increased oxidative stress levels: the Women's Health and Aging Studies.

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Palta, Priya; Szanton, Sarah L; Semba, Richard D; Thorpe, Roland J; Varadhan, Ravi; Fried, Linda P

2015-01-01

Elevated oxidative stress levels may be one mechanism contributing to poor health outcomes. Financial strain and oxidative stress are each predictors of morbidity and mortality, but little research has investigated their relationship. Community-dwelling older adults (n = 728) from the Women's Health and Aging Studies I and II were included in this cross-sectional analysis. Financial strain was ascertained as an ordinal response to: "At the end of the month, do you have more than enough money left over, just enough, or not enough?" Oxidative stress was measured using serum protein carbonyl concentrations. Linear regression was used to quantify the relationship between financial strain and oxidative stress. Participants who reported high financial strain exhibited 13.4% higher protein carbonyl concentrations compared to individuals who reported low financial strain (p = 0.002). High financial strain may be associated with increased oxidative stress, suggesting that oxidative stress could mediate associations between financial strain and poor health. Copyright © 2015 Elsevier Inc. All rights reserved.

Protein Network Signatures Associated with Exogenous Biofuels Treatments in Cyanobacterium Synechocystis sp. PCC 6803

International Nuclear Information System (INIS)

Pei, Guangsheng; Chen, Lei; Wang, Jiangxin; Qiao, Jianjun; Zhang, Weiwen

2014-01-01

Although recognized as a promising microbial cell factory for producing biofuels, current productivity in cyanobacterial systems is low. To make the processes economically feasible, one of the hurdles, which need to be overcome is the low tolerance of hosts to toxic biofuels. Meanwhile, little information is available regarding the cellular responses to biofuels stress in cyanobacteria, which makes it challenging for tolerance engineering. Using large proteomic datasets of Synechocystis under various biofuels stress and environmental perturbation, a protein co-expression network was first constructed and then combined with the experimentally determined protein–protein interaction network. Proteins with statistically higher topological overlap in the integrated network were identified as common responsive proteins to both biofuels stress and environmental perturbations. In addition, a weighted gene co-expression network analysis was performed to distinguish unique responses to biofuels from those to environmental perturbations and to uncover metabolic modules and proteins uniquely associated with biofuels stress. The results showed that biofuel-specific proteins and modules were enriched in several functional categories, including photosynthesis, carbon fixation, and amino acid metabolism, which may represent potential key signatures for biofuels stress responses in Synechocystis. Network-based analysis allowed determination of the responses specifically related to biofuels stress, and the results constituted an important knowledge foundation for tolerance engineering against biofuels in Synechocystis.

Protein Network Signatures Associated with Exogenous Biofuels Treatments in Cyanobacterium Synechocystis sp. PCC 6803

Energy Technology Data Exchange (ETDEWEB)

Pei, Guangsheng; Chen, Lei; Wang, Jiangxin; Qiao, Jianjun, E-mail: jianjunq@tju.edu.cn; Zhang, Weiwen, E-mail: jianjunq@tju.edu.cn [Laboratory of Synthetic Microbiology, School of Chemical Engineering and Technology, Tianjin University, Tianjin (China); Key Laboratory of Systems Bioengineering, Ministry of Education of China, Tianjin (China); SynBio Research Platform, Collaborative Innovation Center of Chemical Science and Engineering, Tianjin (China)

2014-11-03

Although recognized as a promising microbial cell factory for producing biofuels, current productivity in cyanobacterial systems is low. To make the processes economically feasible, one of the hurdles, which need to be overcome is the low tolerance of hosts to toxic biofuels. Meanwhile, little information is available regarding the cellular responses to biofuels stress in cyanobacteria, which makes it challenging for tolerance engineering. Using large proteomic datasets of Synechocystis under various biofuels stress and environmental perturbation, a protein co-expression network was first constructed and then combined with the experimentally determined protein–protein interaction network. Proteins with statistically higher topological overlap in the integrated network were identified as common responsive proteins to both biofuels stress and environmental perturbations. In addition, a weighted gene co-expression network analysis was performed to distinguish unique responses to biofuels from those to environmental perturbations and to uncover metabolic modules and proteins uniquely associated with biofuels stress. The results showed that biofuel-specific proteins and modules were enriched in several functional categories, including photosynthesis, carbon fixation, and amino acid metabolism, which may represent potential key signatures for biofuels stress responses in Synechocystis. Network-based analysis allowed determination of the responses specifically related to biofuels stress, and the results constituted an important knowledge foundation for tolerance engineering against biofuels in Synechocystis.

Genome-Wide Association Mapping of Loci Associated with Plant Growth and Forage Production under Salt Stress in Alfalfa (Medicago sativa L.

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Xiang-Ping Liu

2017-05-01

Full Text Available Salinity tolerance is highly desirable to sustain alfalfa production in marginal lands that have been rendered saline. In this study, we used a diverse panel of 198 alfalfa accessions for mapping loci associated with plant growth and forage production under salt stress using genome-wide association studies (GWAS. The plants were genotyped using genotyping-by-sequencing (GBS. A greenhouse procedure was used for phenotyping four agronomic and physiological traits affected by salt stress, including dry weight (DW, plant height (PH, leaf chlorophyll content (LCC, and stomatal conductance (SC. For each trait, a stress susceptibility index (SSI was used to evaluate plant performance under stressed and non-stressed conditions. Marker-trait association identified a total of 42 markers significantly associated with salt tolerance. They were located on all chromosomes except chromosome 2 based on the alignment of their flanking sequences to the reference genome (Medicago truncatula. Of those identified, 13 were associated with multiple traits. Several loci identified in the present study were also identified in previous reports. BLAST search revealed that 19 putative candidate genes linked to 24 significant markers. Among them, B3 DNA-binding protein, Thiaminepyrophosphokinase and IQ calmodulin-binding motif protein were identified among multiple traits in the present and previous studies. With further investigation, these markers and candidates would be useful for developing markers for marker-assisted selection in breeding programs to improve alfalfa cultivars with enhanced tolerance to salt stress.

Intracellular proteins produced by mammalian cells in response to environmental stress

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Goochee, Charles F.; Passini, Cheryl A.

1988-01-01

The nature of the response of mammalian cells to environmental stress is examined by reviewing results of studies where cultured mouse L cells and baby hamster kidney cells were exposed to heat shock and the synthesis of heat-shock proteins and stress-response proteins (including HSP70, HSC70, HSP90, ubiquitin, and GRP70) in stressed and unstressed cells was evaluated using 2D-PAGE. The intracellular roles of the individual stress response proteins are discussed together with the regulation of the stress response system.

Drought Stress and Its Impact on Protein in Three Species of Vitex

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John A. De Britto

2011-09-01

Full Text Available Drought is one of the most important natural phenomenon which affects on plant growth. When drought stress is imposed different molecular and biochemical responses took place in the plants. The protein profile of three species of Vitex (Vitex trifolia L., Vitex altissima L. and Vitex negundo L. under normally irrigated condition and severe drought plants was analyzed through SDS-PAGE. Drought stress significantly affects proteins in plants when compared the normal conditioned plants. Several new protein bands were identified in the stressed plants. It seems that Vitex species can be adapted to drought stress conditions. Hence it was concluded that number of new proteins were synthesized in stressed plants for their adaptation in the stressed conditions. These proteins could be used as markers in identifying the stressed plants.

WRKY proteins: signaling and regulation of expression during abiotic stress responses.

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Banerjee, Aditya; Roychoudhury, Aryadeep

2015-01-01

WRKY proteins are emerging players in plant signaling and have been thoroughly reported to play important roles in plants under biotic stress like pathogen attack. However, recent advances in this field do reveal the enormous significance of these proteins in eliciting responses induced by abiotic stresses. WRKY proteins act as major transcription factors, either as positive or negative regulators. Specific WRKY factors which help in the expression of a cluster of stress-responsive genes are being targeted and genetically modified to induce improved abiotic stress tolerance in plants. The knowledge regarding the signaling cascade leading to the activation of the WRKY proteins, their interaction with other proteins of the signaling pathway, and the downstream genes activated by them are altogether vital for justified targeting of the WRKY genes. WRKY proteins have also been considered to generate tolerance against multiple abiotic stresses with possible roles in mediating a cross talk between abiotic and biotic stress responses. In this review, we have reckoned the diverse signaling pattern and biological functions of WRKY proteins throughout the plant kingdom along with the growing prospects in this field of research.

ARI3SG: Aerosol retention in the secondary side of a steam generator. Part II: Model validation and uncertainty analysis

International Nuclear Information System (INIS)

Lopez, Claudia; Herranz, Luis E.

2012-01-01

Highlights: ► Validation of a model (ARI3SG) for the aerosol retention in the break stage of a steam generator under SGTR conditions. ► Interpretation of the experimental SGTR and CAAT data by using the ARI3SG model. ► Assessment of the epistemic and stochastic uncertainties effect on the ARI3SG results. - Abstract: A large body of data has been gathered in the last decade through the EU-SGTR, ARTIST and ARTIST 2 projects for aerosol retention in the steam generator during SGTR severe accident sequences. At the same time the attempt to extend the analytical capability has resulted in models that need to be validated. The ARI3SG is one of such developments and it has been built to estimate the aerosol retention in the break stage of a “dry” steam generator. This paper assesses the ARI3SG predictability by comparing its estimates to open data and by analyzing the effect of associated uncertainties. Datamodel comparison has been shown to be satisfactory and highlight the potential use of an ARI3SG-like formulation in system codes.

A role for SR proteins in plant stress responses.

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Duque, Paula

2011-01-01

Members of the SR (serine/arginine-rich) protein gene family are key players in the regulation of alternative splicing, an important means of generating proteome diversity and regulating gene expression. In plants, marked changes in alternative splicing are induced by a wide variety of abiotic stresses, suggesting a role for this highly versatile gene regulation mechanism in the response to environmental cues. In support of this notion, the expression of plant SR proteins is stress-regulated at multiple levels, with environmental signals controlling their own alternative splicing patterns, phosphorylation status and subcellular distribution. Most importantly, functional links between these RNA-binding proteins and plant stress tolerance are beginning to emerge, including a role in the regulation of abscisic acid (ABA) signaling. Future identification of the physiological mRNA targets of plant SR proteins holds much promise for the elucidation of the molecular mechanisms underlying their role in the response to abiotic stress.

Membrane Stored Curvature Elastic Stress Modulates Recruitment of Maintenance Proteins PspA and Vipp1.

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McDonald, Christopher; Jovanovic, Goran; Ces, Oscar; Buck, Martin

2015-09-01

Phage shock protein A (PspA), which is responsible for maintaining inner membrane integrity under stress in enterobacteria, and vesicle-inducting protein in plastids 1 (Vipp1), which functions for membrane maintenance and thylakoid biogenesis in cyanobacteria and plants, are similar peripheral membrane-binding proteins. Their homologous N-terminal amphipathic helices are required for membrane binding; however, the membrane features recognized and required for expressing their functionalities have remained largely uncharacterized. Rigorously controlled, in vitro methodologies with lipid vesicles and purified proteins were used in this study and provided the first biochemical and biophysical characterizations of membrane binding by PspA and Vipp1. Both proteins are found to sense stored curvature elastic (SCE) stress and anionic lipids within the membrane. PspA has an enhanced sensitivity for SCE stress and a higher affinity for the membrane than Vipp1. These variations in binding may be crucial for some of the proteins' differing roles in vivo. Assays probing the transcriptional regulatory function of PspA in the presence of vesicles showed that a relief of transcription inhibition occurs in an SCE stress-specific manner. This in vitro recapitulation of membrane stress-dependent transcription control suggests that the Psp response may be mounted in vivo when a cell's inner membrane experiences increased SCE stress. All cell types maintain the integrity of their membrane systems. One widely distributed membrane stress response system in bacteria is the phage shock protein (Psp) system. The central component, peripheral membrane protein PspA, which mitigates inner membrane stress in bacteria, has a counterpart, Vipp1, which functions for membrane maintenance and thylakoid biogenesis in plants and photosynthetic bacteria. Membrane association of both these proteins is accepted as playing a pivotal role in their functions. Here we show that direct membrane binding by

Frequent sgRNA-barcode recombination in single-cell perturbation assays.

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Shiqi Xie

Full Text Available Simultaneously detecting CRISPR-based perturbations and induced transcriptional changes in the same cell is a powerful approach to unraveling genome function. Several lentiviral approaches have been developed, some of which rely on the detection of distally located genetic barcodes as an indirect proxy of sgRNA identity. Since barcodes are often several kilobases from their corresponding sgRNAs, viral recombination-mediated swapping of barcodes and sgRNAs is feasible. Using a self-circularization-based sgRNA-barcode library preparation protocol, we estimate the recombination rate to be ~50% and we trace this phenomenon to the pooled viral packaging step. Recombination is random, and decreases the signal-to-noise ratio of the assay. Our results suggest that alternative approaches can increase the throughput and sensitivity of single-cell perturbation assays.

Seminal Plasma Proteins as Androgen Receptor Corregulators Promote Prostate Cancer Growth

Science.gov (United States)

2016-12-01

protease activity of PSA [14]. Semenogelins are expressed in other male genital organs, such as the vas def - erens, epididymis, and prostate, as well...expression and secretion in prostate cancer lines stably expressing SgI. Cell extracts (A) or acetone-precipitated proteins in con - ditioned media... con - firmed this by demonstrating the failure to detect SgI signals in the conditioned medium after culturing control LNCaP with endogenous SgI and

Overexpression of BAX INHIBITOR-1 Links Plasma Membrane Microdomain Proteins to Stress.

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Ishikawa, Toshiki; Aki, Toshihiko; Yanagisawa, Shuichi; Uchimiya, Hirofumi; Kawai-Yamada, Maki

2015-10-01

BAX INHIBITOR-1 (BI-1) is a cell death suppressor widely conserved in plants and animals. Overexpression of BI-1 enhances tolerance to stress-induced cell death in plant cells, although the molecular mechanism behind this enhancement is unclear. We recently found that Arabidopsis (Arabidopsis thaliana) BI-1 is involved in the metabolism of sphingolipids, such as the synthesis of 2-hydroxy fatty acids, suggesting the involvement of sphingolipids in the cell death regulatory mechanism downstream of BI-1. Here, we show that BI-1 affects cell death-associated components localized in sphingolipid-enriched microdomains of the plasma membrane in rice (Oryza sativa) cells. The amount of 2-hydroxy fatty acid-containing glucosylceramide increased in the detergent-resistant membrane (DRM; a biochemical counterpart of plasma membrane microdomains) fraction obtained from BI-1-overexpressing rice cells. Comparative proteomics analysis showed quantitative changes of DRM proteins in BI-1-overexpressing cells. In particular, the protein abundance of FLOTILLIN HOMOLOG (FLOT) and HYPERSENSITIVE-INDUCED REACTION PROTEIN3 (HIR3) markedly decreased in DRM of BI-1-overexpressing cells. Loss-of-function analysis demonstrated that FLOT and HIR3 are required for cell death by oxidative stress and salicylic acid, suggesting that the decreased levels of these proteins directly contribute to the stress-tolerant phenotypes in BI-1-overexpressing rice cells. These findings provide a novel biological implication of plant membrane microdomains in stress-induced cell death, which is negatively modulated by BI-1 overexpression via decreasing the abundance of a set of key proteins involved in cell death. © 2015 American Society of Plant Biologists. All Rights Reserved.

Endoplasmic Reticulum Stress and Associated ROS

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Hafiz Maher Ali Zeeshan

2016-03-01

Full Text Available The endoplasmic reticulum (ER is a fascinating network of tubules through which secretory and transmembrane proteins enter unfolded and exit as either folded or misfolded proteins, after which they are directed either toward other organelles or to degradation, respectively. The ER redox environment dictates the fate of entering proteins, and the level of redox signaling mediators modulates the level of reactive oxygen species (ROS. Accumulating evidence suggests the interrelation of ER stress and ROS with redox signaling mediators such as protein disulfide isomerase (PDI-endoplasmic reticulum oxidoreductin (ERO-1, glutathione (GSH/glutathione disuphide (GSSG, NADPH oxidase 4 (Nox4, NADPH-P450 reductase (NPR, and calcium. Here, we reviewed persistent ER stress and protein misfolding-initiated ROS cascades and their significant roles in the pathogenesis of multiple human disorders, including neurodegenerative diseases, diabetes mellitus, atherosclerosis, inflammation, ischemia, and kidney and liver diseases.

Aerobic exercise training rescues cardiac protein quality control and blunts endoplasmic reticulum stress in heart failure rats.

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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.

Multidrug-resistance-associated protein plays a protective role in menadione-induced oxidative stress in endothelial cells.

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Takahashi, Kyohei; Shibata, Tomohito; Oba, Tatsuya; Ishikawa, Tetsuya; Yoshikawa, Masahito; Tatsunami, Ryosuke; Takahashi, Kazuhiko; Tampo, Yoshiko

2009-02-13

Menadione, a redox-cycling quinone known to cause oxidative stress, binds to reduced glutathione (GSH) to form glutathione S-conjugate. Glutathione S-conjugates efflux is often mediated by multidrug-resistance-associated protein (MRP). We investigated the effect of a transporter inhibitor, MK571 (3-[[3-[2-(7-chloroquinolin-2-yl)vinyl]phenyl]-(2-dimethylcarbamoylethylsulfanyl)methylsulfanyl] propionic acid), on menadione-induced oxidative stress in bovine aortic endothelial cells (BAECs). BAECs were treated with menadione and MK571, and cell viability was measured. Modulation of intracellular GSH levels was performed with buthionine sulfoximine and GSH ethyl ester treatments. Intracellular superoxide was estimated by dihydroethidium oxidation using fluorescence microscopy or flow cytometry. Expression of MRP was determined by flow cytometry using phycoerythrin-conjugated anti-MRP monoclonal antibody. Intracellular GSH depletion by buthionine sulfoximine promoted the loss of viability of BAECs exposed to menadione. Exogenous GSH, which does not permeate the cell membrane, or GSH ethyl ester protected BAECs against the loss of viability induced by menadione. The results suggest that GSH binds to menadione outside the cells as well as inside. Pretreatment of BAECs with MK571 dramatically increased intracellular levels of superoxide generated from menadione, indicating that menadione may accumulate in the intracellular milieu. Finally, we found that MK571 aggravated menadione-induced toxicity in BAECs and that MRP levels were increased in menadione-treated cells. We conclude that MRP plays a vital role in protecting BAECs against menadione-induced oxidative stress, presumably due to its ability to transport glutathione S-conjugate.

Salt stress-induced protein pattern associated with photosynthetic parameters and andrographolide content in Andrographis paniculata Nees.

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Talei, Daryush; Valdiani, Alireza; Maziah, Mahmood; Sagineedu, Sreenivasa Rao; Abiri, Rambod

2015-01-01

Andrographis paniculata is a multifunctional medicinal plant and a potent source of bioactive compounds. Impact of environmental stresses such as salinity on protein diversification, as well as the consequent changes in the photosynthetic parameters and andrographolide content (AG) of the herb, has not yet been thoroughly investigated. The present study showed that the salinity affects the protein pattern, and subsequently, it decreased the photosynthetic parameters, protein content, total dry weight, and total crude extract. Exceptionally, the AG content was increased (p ≤ 0.01). Moreover, it was noticed that the salinity at 12 dS m(-1) led to the maximum increase in AG content in all accessions. Interestingly, the leaf protein analysis revealed that the two polymorphic protein bands as low- and medium-sized of 17 and 45 kDa acted as the activator agents for the photosynthetic parameters and AG content. Protein sequencing and proteomic analysis can be conducted based on the present findings in the future.

Overexpression of a cytosolic abiotic stress responsive universal stress protein (SbUSP mitigates salt and osmotic stress in transgenic tobacco plants

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Pushpika eUdawat

2016-04-01

Full Text Available The Universal Stress Protein (USP is a ubiquitous protein and plays an indispensable role in plant abiotic stress tolerance. The genome of Salicornia brachiata contains two homologues of intron less SbUSP gene which encodes for salt and osmotic responsive universal stress protein. In vivo localization reveals that SbUSP is a membrane bound cytosolic protein. The role of the gene was functionally validated by developing transgenic tobacco and compared with control (wild type and vector control plants under different abiotic stress condition. Transgenic lines (T1 exhibited higher chlorophyll, relative water, proline, total sugar, reducing sugar, free amino acids, polyphenol contents, osmotic potential, membrane stability and lower electrolyte leakage and lipid peroxidation (malondialdehyde content under stress treatments than control (WT and VC plants. Lower accumulation of H2O2 and O2- radicals was also detected in transgenic lines compared to control plants under stress conditions. Present study confers that overexpression of the SbUSP gene enhances plant growth, alleviates ROS buildup, maintains ion homeostasis and improves the physiological status of the plant under salt and osmotic stresses. Principal component analysis (PCA exhibited a statistical distinction of plant response to salinity stress, and a significant response was observed for transgenic lines under stress, which provides stress endurance to the plant. A possible signaling role is proposed that some downstream genes may get activated by abiotic stress responsive cytosolic SbUSP, which leads to the protection of cell from oxidative damages. The study unveils that ectopic expression of the gene mitigates salt or osmotic stress by scavenging ROS and modulating the physiological process of the plant.

Trash to Supply Gas (TtSG) Project Overview

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Hintze, Paul; Santiago-Maldonado, Edgardo; Kulis, Michael J.; Lytle, John K.; Fisher, John W.; Vaccaro, Helen; Ewert, Michael K.; Broyan, James L.

2012-01-01

Technologies that reduce logistical needs are a key to long term space missions. Currently, trash and waste generated during a mission is carried during the entire roundtrip mission or stored inside a logistic module which is de-orbited into Earth's atmosphere for destruction. The goal of the Trash to Supply Gas (TtSG) project is to develop space technology alternatives for converting trash and other waste materials from human spaceflight into high-value products that might include propellants or power system fuels in addition to life support oxygen and water. In addition to producing a useful product from waste, TtSG will decrease the volume needed to store waste on long term space missions. This paper presents an overview of the TtSG technologies and future plans for the project.

CIKS, a connection to Ikappa B kinase and stress-activated protein kinase.

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Leonardi, A; Chariot, A; Claudio, E; Cunningham, K; Siebenlist, U

2000-09-12

Pathogens, inflammatory signals, and stress cause acute transcriptional responses in cells. The induced expression of genes in response to these signals invariably involves transcription factors of the NF-kappaB and AP-1/ATF families. Activation of NF-kappaB factors is thought to be mediated primarily via IkappaB kinases (IKK), whereas that of AP-1/ATF can be mediated by stress-activated protein kinases (SAPKs; also named Jun kinases or JNKs). IKKalpha and IKKbeta are two catalytic subunits of a core IKK complex that also contains the regulatory subunit NEMO (NF-kappaB essential modulator)/IKKgamma. The latter protein is essential for activation of the IKKs, but its mechanism of action is not known. Here we describe the molecular cloning of CIKS (connection to IKK and SAPK/JNK), a previously unknown protein that directly interacts with NEMO/IKKgamma in cells. When ectopically expressed, CIKS stimulates IKK and SAPK/JNK kinases and it transactivates an NF-kappaB-dependent reporter. Activation of NF-kappaB is prevented in the presence of kinase-deficient, interfering mutants of the IKKs. CIKS may help to connect upstream signaling events to IKK and SAPK/JNK modules. CIKS could coordinate the activation of two stress-induced signaling pathways, functions reminiscent of those noted for tumor necrosis factor receptor-associated factor adaptor proteins.

CIKS, a connection to IκB kinase and stress-activated protein kinase

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Leonardi, Antonio; Chariot, Alain; Claudio, Estefania; Cunningham, Kirk; Siebenlist, Ulrich

2000-01-01

Pathogens, inflammatory signals, and stress cause acute transcriptional responses in cells. The induced expression of genes in response to these signals invariably involves transcription factors of the NF-κB and AP-1/ATF families. Activation of NF-κB factors is thought to be mediated primarily via IκB kinases (IKK), whereas that of AP-1/ATF can be mediated by stress-activated protein kinases (SAPKs; also named Jun kinases or JNKs). IKKα and IKKβ are two catalytic subunits of a core IKK complex that also contains the regulatory subunit NEMO (NF-κB essential modulator)/IKKγ. The latter protein is essential for activation of the IKKs, but its mechanism of action is not known. Here we describe the molecular cloning of CIKS (connection to IKK and SAPK/JNK), a previously unknown protein that directly interacts with NEMO/IKKγ in cells. When ectopically expressed, CIKS stimulates IKK and SAPK/JNK kinases and it transactivates an NF-κB-dependent reporter. Activation of NF-κB is prevented in the presence of kinase-deficient, interfering mutants of the IKKs. CIKS may help to connect upstream signaling events to IKK and SAPK/JNK modules. CIKS could coordinate the activation of two stress-induced signaling pathways, functions reminiscent of those noted for tumor necrosis factor receptor-associated factor adaptor proteins. PMID:10962033

Impact of Post-Translational Modifications of Crop Proteins under Abiotic Stress

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Akiko Hashiguchi

2016-12-01

Full Text Available The efficiency of stress-induced adaptive responses of plants depends on intricate coordination of multiple signal transduction pathways that act coordinately or, in some cases, antagonistically. Protein post-translational modifications (PTMs can regulate protein activity and localization as well as protein–protein interactions in numerous cellular processes, thus leading to elaborate regulation of plant responses to various external stimuli. Understanding responses of crop plants under field conditions is crucial to design novel stress-tolerant cultivars that maintain robust homeostasis even under extreme conditions. In this review, proteomic studies of PTMs in crops are summarized. Although the research on the roles of crop PTMs in regulating stress response mechanisms is still in its early stage, several novel insights have been retrieved so far. This review covers techniques for detection of PTMs in plants, representative PTMs in plants under abiotic stress, and how PTMs control functions of representative proteins. In addition, because PTMs under abiotic stresses are well described in soybeans under submergence, recent findings in PTMs of soybean proteins under flooding stress are introduced. This review provides information on advances in PTM study in relation to plant adaptations to abiotic stresses, underlining the importance of PTM study to ensure adequate agricultural production in the future.

Inclusive search for b → sg

International Nuclear Information System (INIS)

1998-07-01

The authors describe an inclusive search for flavor changing neutral current decays of the type b → s gluon in the SLD experiment. Models of b → sg indicate that the production of high momentum kaons is enhanced over background from standard B decays. If the branching ratio for b → sg is ∼ 10%, then such an enhancement should have a good signal to background ratio. The analysis makes use of the particle identification and high precision vertexing capabilities of SLD to search for such an enhancement. The data sample consists of 300K hadronic Z 0 decays collected between 1993 and 1997

Analysis of soybean root proteins affected by gibberellic acid treatment under flooding stress.

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Oh, Myeong Won; Nanjo, Yohei; Komatsu, Setsuko

2014-01-01

Flooding is a serious abiotic stress for soybean because it restricts growth and reduces grain yields. To investigate the effect of gibberellic acid (GA) on soybean under flooding stress, root proteins were analyzed using a gel-free proteomic technique. Proteins were extracted from the roots of 4-days-old soybean seedlings exposed to flooding stress in the presence and absence of exogenous GA3 for 2 days. A total of 307, 324, and 250 proteins were identified from untreated, and flooding-treated soybean seedlings without or with GA3, respectively. Secondary metabolism- and cell-related proteins, and proteins involved in protein degradation/synthesis were decreased by flooding stress; however, the levels of these proteins were restored by GA3 supplementation under flooding. Fermentation- and cell wall-related proteins were not affected by GA3 supplementation. Furthermore, putative GA-responsive proteins, which were identified by the presence of a GA-responsive element in the promoter region, were less abundant by flooding stress; however, these proteins were more abundant by GA3 supplementation under flooding. Taken together, these results suggest that GA3 affects the abundance of proteins involved in secondary metabolism, cell cycle, and protein degradation/synthesis in soybeans under flooding stress.

Class I and II Small Heat Shock Proteins Together with HSP101 Protect Protein Translation Factors during Heat Stress.

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McLoughlin, Fionn; Basha, Eman; Fowler, Mary E; Kim, Minsoo; Bordowitz, Juliana; Katiyar-Agarwal, Surekha; Vierling, Elizabeth

2016-10-01

The ubiquitous small heat shock proteins (sHSPs) are well documented to act in vitro as molecular chaperones to prevent the irreversible aggregation of heat-sensitive proteins. However, the in vivo activities of sHSPs remain unclear. To investigate the two most abundant classes of plant cytosolic sHSPs (class I [CI] and class II [CII]), RNA interference (RNAi) and overexpression lines were created in Arabidopsis (Arabidopsis thaliana) and shown to have reduced and enhanced tolerance, respectively, to extreme heat stress. Affinity purification of CI and CII sHSPs from heat-stressed seedlings recovered eukaryotic translation elongation factor (eEF) 1B (α-, β-, and γ-subunits) and eukaryotic translation initiation factor 4A (three isoforms), although the association with CI sHSPs was stronger and additional proteins involved in translation were recovered with CI sHSPs. eEF1B subunits became partially insoluble during heat stress and, in the CI and CII RNAi lines, showed reduced recovery to the soluble cell fraction after heat stress, which was also dependent on HSP101. Furthermore, after heat stress, CI sHSPs showed increased retention in the insoluble fraction in the CII RNAi line and vice versa. Immunolocalization revealed that both CI and CII sHSPs were present in cytosolic foci, some of which colocalized with HSP101 and with eEF1Bγ and eEF1Bβ. Thus, CI and CII sHSPs have both unique and overlapping functions and act either directly or indirectly to protect specific translation factors in cytosolic stress granules. © 2016 American Society of Plant Biologists. All Rights Reserved.

Identification of genes and pathways associated with aluminum stress and tolerance using transcriptome profiling of wheat near-isogenic lines.

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Houde, Mario; Diallo, Amadou Oury

2008-08-27

Aluminum is considered the most limiting factor for plant productivity in acidic soils, which cover large areas of the world's potential arable lands. The inhibition of root growth is recognized as the primary effect of Al toxicity. To identify genes associated with Al stress and tolerance, transcriptome analyses of four different wheat lines (2 Al-tolerant and 2 Al sensitive) that differ in their response to Al were performed. Microarray expression profiling revealed that 83 candidate genes are associated with Al stress and 25 are associated with tolerance. The stress-associated genes include important enzymes such as pyruvate dehydrogenase, alternative oxidase, and galactonolactone oxidase, ABC transporter and ascorbate oxido-reducatase. The Al tolerance-associated genes include the ALMT-1 malate transporter, glutathione S-transferase, germin/oxalate oxidase, fructose 1,6-bisphosphatase, cysteine-rich proteins, cytochrome P450 monooxygenase, cellulose synthase, zinc finger transcription factor, disease resistance response protein and F-box containing domain protein. In this survey, we identified stress- and tolerance-associated genes that may be involved in the detoxification of Al and reactive oxygen species. Alternative pathways could help maintain the supply of important metabolites (H2O2, ascorbate, NADH, and phosphate) needed for Al tolerance and root growth. The Al tolerance-associated genes may be key factors that regulate these pathways.

Microtubule-dependent association of AKAP350A and CCAR1 with RNA stress granules

International Nuclear Information System (INIS)

Kolobova, Elena; Efimov, Andrey; Kaverina, Irina; Rishi, Arun K.; Schrader, John W.; Ham, Amy-Joan; Larocca, M. Cecilia; Goldenring, James R.

2009-01-01

Recent investigations have highlighted the importance of subcellular localization of mRNAs to cell function. While AKAP350A, a multifunctional scaffolding protein, localizes to the Golgi apparatus and centrosomes, we have now identified a cytosolic pool of AKAP350A. Analysis of AKAP350A scaffolded complexes revealed two novel interacting proteins, CCAR1 and caprin-1. CCAR1, caprin-1 and AKAP350A along with G3BP, a stress granule marker, relocate to RNA stress granules after arsenite treatment. Stress also caused loss of AKAP350 from the Golgi and fragmentation of the Golgi apparatus. Disruption of microtubules with nocodazole altered stress granule formation and changed their morphology by preventing fusion of stress granules. In the presence of nocodazole, arsenite induced smaller granules with the vast majority of AKAP350A and CCAR1 separated from G3BP-containing granules. Similar to nocodazole treatment, reduction of AKAP350A or CCAR1 expression also altered the size and number of G3BP-containing stress granules induced by arsenite treatment. A limited set of 69 mRNA transcripts was immunoisolated with AKAP350A even in the absence of stress, suggesting the association of AKAP350A with mRNA transcripts. These results provide the first evidence for the microtubule dependent association of AKAP350A and CCAR1 with RNA stress granules

Apoptosis in response to heat stress is positively associated with heat-shock protein 90 expression in chicken myocardial cells in vitro.

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Zhang, Xiao-Hui; Wu, Hong; Tang, Shu; Li, Qiao-Ning; Xu, Jiao; Zhang, Miao; Su, Ya-Nan; Yin, Bin; Zhao, Qi-Ling; Kemper, Nicole; Hartung, Joerg; Bao, En-Dong

2017-06-30

To determine heat-shock protein (Hsp)90 expression is connected with cellular apoptotic response to heat stress and its mechanism, chicken ( Gallus gallus ) primary myocardial cells were treated with the Hsp90 promoter, aspirin, and its inhibitor, geldanamycin (GA), before heat stress. Cellular viability, heat-stressed apoptosis and reactive oxygen species level under different treatments were measured, and the expression of key proteins of the signaling pathway related to Hsp90 and their colocalization with Hsp90 were detected. The results showed that aspirin treatment increased the expression of protein kinase B (Akt), the signal transducer and activator of transcription (STAT)-3 and p-IKKα/β and the colocalization of Akt and STAT-3 with Hsp90 during heat stress, which was accompanied by improved viability and low apoptosis. GA significantly inhibited Akt expression and p-IKKα/β level, but not STAT-3 quantity, while the colocalization of Akt and STAT-3 with Hsp90 was weakened, followed by lower cell viability and higher apoptosis. Aspirin after GA treatment partially improved the stress response and apoptosis rate of tested cells caused by the recovery of Akt expression and colocalization, rather than the level of STAT-3 (including its co-localization with Hsp90) and p-IKKα/β. Therefore, Hsp90 expression has a positive effect on cellular capacity to resist heat-stressed injury and apoptosis. Moreover, inhibition of Hsp90 before stress partially attenuated its positive effects.

Fluorescent proteins such as eGFP lead to catalytic oxidative stress in cells.

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Ganini, Douglas; Leinisch, Fabian; Kumar, Ashutosh; Jiang, JinJie; Tokar, Erik J; Malone, Christine C; Petrovich, Robert M; Mason, Ronald P

2017-08-01

Fluorescent proteins are an important tool that has become omnipresent in life sciences research. They are frequently used for localization of proteins and monitoring of cells [1,2]. Green fluorescent protein (GFP) was the first and has been the most used fluorescent protein. Enhanced GFP (eGFP) was optimized from wild-type GFP for increased fluorescence yield and improved expression in mammalian systems [3]. Many GFP-like fluorescent proteins have been discovered, optimized or created, such as the red fluorescent protein TagRFP [4]. Fluorescent proteins are expressed colorless and immature and, for eGFP, the conversion to the fluorescent form, mature, is known to produce one equivalent of hydrogen peroxide (H 2 O 2 ) per molecule of chromophore [5,6]. Even though it has been proposed that this process is non-catalytic and generates nontoxic levels of H 2 O 2 [6], this study investigates the role of fluorescent proteins in generating free radicals and inducing oxidative stress in biological systems. Immature eGFP and TagRFP catalytically generate the free radical superoxide anion (O 2 •- ) and H 2 O 2 in the presence of NADH. Generation of the free radical O 2 •- and H 2 O 2 by eGFP in the presence of NADH affects the gene expression of cells. Many biological pathways are altered, such as a decrease in HIF1α stabilization and activity. The biological pathways altered by eGFP are known to be implicated in the pathophysiology of many diseases associated with oxidative stress; therefore, it is critical that such experiments using fluorescent proteins are validated with alternative methodologies and the results are carefully interpreted. Since cells inevitably experience oxidative stress when fluorescent proteins are expressed, the use of this tool for cell labeling and in vivo cell tracing also requires validation using alternative methodologies. Published by Elsevier B.V.

A Heavy Metal-Associated Protein (AcHMA1 from the Halophyte, Atriplex canescens (Pursh Nutt., Confers Tolerance to Iron and Other Abiotic Stresses When Expressed in Saccharomyces cerevisiae

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Xin-Hua Sun

2014-08-01

Full Text Available Many heavy metals are essential for metabolic processes, but are toxic at elevated levels. Metal tolerance proteins provide resistance to this toxicity. In this study, we identified and characterized a heavy metal-associated protein, AcHMA1, from the halophyte, Atriplex canescens. Sequence analysis has revealed that AcHMA1 contains two heavy metal binding domains. Treatments with metals (Fe, Cu, Ni, Cd or Pb, PEG6000 and NaHCO3 highly induced AcHMA1 expression in A. canescens, whereas NaCl and low temperature decreased its expression. The role of AcHMA1 in metal stress tolerance was examined using a yeast expression system. Expression of the AcHMA1 gene significantly increased the ability of yeast cells to adapt to and recover from exposure to excess iron. AcHMA1 expression also provided salt, alkaline, osmotic and oxidant stress tolerance in yeast cells. Finally, subcellular localization of an AcHMA1/GFP fusion protein expressed in tobacco cells showed that AcHMA1 was localized in the plasma membrane. Thus, our results suggest that AcHMA1 encodes a membrane-localized metal tolerance protein that mediates the detoxification of iron in eukaryotes. Furthermore, AcHMA1 also participates in the response to abiotic stress.

Protein intake and stress levels in nurses and housewives of Pakistan

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Wattoo, Feroza Hamid; Memon, Muhammad Saleh; Memon, Allah Nawaz; Wattoo, Muhammad Hamid Sarwar; Asad, Muhammad Javaid; Siddique, Farzana

2011-01-01

Stress has many biological effects on human daily life. In the present study, dietary protein intake was correlated with the investigated stress levels of nurses and housewives of the targeted urban population. Age group ranged from 30 to 45 years and both the groups belonged to middle socioeconomic status. After calculations of environmental, psychological and physiological stresses, it was observed that the levels of stress in housewives were significantly higher than those of nurses. Recommended dietary allowances, RDA and actual protein intakes, API were also compared in both the groups. The found protein intake was less in housewives as compared to that of nurses. PMID:23961140

Cluster Differentiating 36 (CD36) Deficiency Attenuates Obesity-Associated Oxidative Stress in the Heart.

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Gharib, Mohamed; Tao, Huan; Fungwe, Thomas V; Hajri, Tahar

2016-01-01

Obesity is often associated with a state of oxidative stress and increased lipid deposition in the heart. More importantly, obesity increases lipid influx into the heart and induces excessive production of reactive oxygen species (ROS) leading to cell toxicity and metabolic dysfunction. Cluster differentiating 36 (CD36) protein is highly expressed in the heart and regulates lipid utilization but its role in obesity-associated oxidative stress is still not clear. The aim of this study was to determine the impact of CD36 deficiency on cardiac steatosis, oxidative stress and lipotoxicity associated with obesity. Studies were conducted in control (Lean), obese leptin-deficient (Lepob/ob) and leptin-CD36 double null (Lepob/obCD36-/-) mice. Compared to lean mice, cardiac steatosis, and fatty acid (FA) uptake and oxidation were increased in Lepob/ob mice, while glucose uptake and oxidation was reduced. Moreover, insulin resistance, oxidative stress markers and NADPH oxidase-dependent ROS production were markedly enhanced. This was associated with the induction of NADPH oxidase expression, and increased membrane-associated p47phox, p67phox and protein kinase C. Silencing CD36 in Lepob/ob mice prevented cardiac steatosis, increased insulin sensitivity and glucose utilization, but reduced FA uptake and oxidation. Moreover, CD36 deficiency reduced NADPH oxidase activity and decreased NADPH oxidase-dependent ROS production. In isolated cardiomyocytes, CD36 deficiency reduced palmitate-induced ROS production and normalized NADPH oxidase activity. CD36 deficiency prevented obesity-associated cardiac steatosis and insulin resistance, and reduced NADPH oxidase-dependent ROS production. The study demonstrates that CD36 regulates NADPH oxidase activity and mediates FA-induced oxidative stress.

Association between prenatal psychological stress and oxidative stress during pregnancy.

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Eick, Stephanie M; Barrett, Emily S; van 't Erve, Thomas J; Nguyen, Ruby H N; Bush, Nicole R; Milne, Ginger; Swan, Shanna H; Ferguson, Kelly K

2018-03-30

Prenatal psychological stress during pregnancy has been associated with adverse reproductive outcomes. A growing animal literature supports an association between psychological stress and oxidative stress. We assessed this relationship in pregnant women, hypothesising that psychological stress is associated with higher concentrations of oxidative stress biomarkers during pregnancy. Psychosocial status and stressful life events (SLE) were self-reported. 8-iso-prostaglandin F 2α (8-iso-PGF 2α ) was measured as a biomarker of oxidative stress in urine samples at median 32 weeks' gestation. We examined SLEs individually (ever vs never) and in summary (any vs none) and psychosocial status as measured by individual subscales and in summary (poor vs good). Linear models estimated associations between these parameters and urinary 8-iso-PGF 2α concentrations after adjusting for covariates. The geometric mean of 8-iso-PGF 2α was significantly higher among pregnant women who were non-White, smokers, had less than a college education, higher pre-pregnancy BMI and were unmarried. Having ever had a death in the family (n = 39) during pregnancy was associated with a 22.9% increase in 8-iso-PGF 2α in unadjusted models (95% confidence interval [CI] 1.50, 48.8). Poor psychosocial status was associated with a 13.1% (95% CI 2.43, 25.0) greater mean 8-iso-PGF 2α in unadjusted analyses. Associations were attenuated, but remained suggestive, after covariate adjustment. These data suggest that 8-iso-PGF 2α is elevated in pregnant women with who are at a sociodemographic disadvantage and who have higher psychological stress in pregnancy. Previous studies have observed that 8-iso-PGF 2α levels are associated with adverse birth outcomes, oxidative stress could be a mediator in these relationships. © 2018 John Wiley & Sons Ltd.

Growth inhibition of head and neck squamous cell carcinoma cells by sgRNA targeting the cyclin D1 mRNA based on TRUE gene silencing.

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Satoshi Iizuka

Full Text Available Head and neck squamous cell carcinoma (HNSCC exhibits increased expression of cyclin D1 (CCND1. Previous studies have shown a correlation between poor prognosis of HNSCC and cyclin D1 overexpression. tRNase ZL-utilizing efficacious gene silencing (TRUE gene silencing is one of the RNA-mediated gene expression control technologies that have therapeutic potential. This technology is based on a unique enzymatic property of mammalian tRNase ZL, which is that it can cleave any target RNA at any desired site by recognizing a pre-tRNA-like complex formed between the target RNA and an artificial small guide RNA (sgRNA. In this study, we designed several sgRNAs targeting human cyclin D1 mRNA to examine growth inhibition of HNSCC cells. Transfection of certain sgRNAs decreased levels of cyclin D1 mRNA and protein in HSC-2 and HSC-3 cells, and also inhibited their proliferation. The combination of these sgRNAs and cisplatin showed more than additive inhibition of cancer cell growth. These findings demonstrate that TRUE gene silencing of cyclin D1 leads to inhibition of the growth of HNSCC cells and suggest that these sgRNAs alone or combined with cisplatin may be a useful new therapy for HNSCCs.

Measurement of Hepatic Protein Fractional Synthetic Rate with Stable Isotope Labeling Technique in Thapsigargin Stressed HepG2 Cells

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Song, Juquan; Zhang, Xiao-jun; Boehning, Darren; Brooks, Natasha C.; Herndon, David N.; Jeschke, Marc G.

2012-01-01

Severe burn-induced liver damage and dysfunction is associated with endoplasmic reticulum (ER) stress. ER stress has been shown to regulate global protein synthesis. In the current study, we induced ER stress in vitro and estimated the effect of ER stress on hepatic protein synthesis. The aim was two-fold: (1) to establish an in vitro model to isotopically measure hepatic protein synthesis and (2) to evaluate protein fractional synthetic rate (FSR) in response to ER stress. Human hepatocellular carcinoma cells (HepG2) were cultured in medium supplemented with stable isotopes 1,2-13C2-glycine and L-[ring-13C6]phenylalanine. ER stress was induced by exposing the cells to 100 nM of thapsigargin (TG). Cell content was collected from day 0 to 14. Alterations in cytosolic calcium were measured by calcium imaging and ER stress markers were confirmed by Western blotting. The precursor and product enrichments were detected by GC-MS analysis for FSR calculation. We found that the hepatic protein FSR were 0.97±0.02 and 0.99±0.05%/hr calculated from 1,2-13C2-glycine and L-[ring-13C6]phenylalanine, respectively. TG depleted ER calcium stores and induced ER stress by upregulating p-IRE-1 and Bip. FSR dramatically decreased to 0.68±0.03 and 0.60±0.06%/hr in the TG treatment group (pisotope tracer incorporation technique is a useful method for studying the effects of ER stress on hepatic protein synthesis. PMID:22298954

Advanced evaluation method of SG TSP BEC hole blockage rate

International Nuclear Information System (INIS)

Izumida, Hiroyuki; Nagata, Yasuyuki; Harada, Yutaka; Murakami, Ryuji

2003-01-01

In spite of the control of the water chemistry of SG secondary feed-water in PWR-SG, SG TSP BEC holes, which are the flow path of secondary water, are often clogged. In the past, the trending of BEC hole blockage rate has conducted by evaluating ECT original signals and visual inspections. However, the ECT original signals of deposits are diversified, it becomes difficult to analyze them with the existing evaluation method using the ECT original signals. In this regard, we have developed the secondary side visual inspection system, which enables the high-accuracy evaluation of BEC hole blockage rate, and new ECT signal evaluation method. (author)

dFOXO Activates Large and Small Heat Shock Protein Genes in Response to Oxidative Stress to Maintain Proteostasis in Drosophila.

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Donovan, Marissa R; Marr, Michael T

2016-09-02

Maintaining protein homeostasis is critical for survival at the cellular and organismal level (Morimoto, R. I. (2011) Cold Spring Harb. Symp. Quant. Biol. 76, 91-99). Cells express a family of molecular chaperones, the heat shock proteins, during times of oxidative stress to protect against proteotoxicity. We have identified a second stress responsive transcription factor, dFOXO, that works alongside the heat shock transcription factor to activate transcription of both the small heat shock protein and the large heat shock protein genes. This expression likely protects cells from protein misfolding associated with oxidative stress. Here we identify the regions of the Hsp70 promoter essential for FOXO-dependent transcription using in vitro methods and find a physiological role for FOXO-dependent expression of heat shock proteins in vivo. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Transfer-messenger RNA controls the translation of cell-cycle and stress proteins in Streptomyces

DEFF Research Database (Denmark)

Barends, Sharief; Zehl, Martin; Bialek, Sylwia

2010-01-01

coelicolor, trans-translation has a specialized role in stress management. Analysis of proteins that were carboxy-terminally His(8)-tagged by a recombinant tmRNA identified only 10 targets, including the stress proteins: DnaK heat-shock protein 70, thiostrepton-induced protein A, universal stress protein A...... functionality for tmRNA, promoting the translation of the same mRNA it targets, at the expense of sacrificing the first nascent protein. In streptomycetes, tmRNA has evolved into a dedicated task force that ensures the instantaneous response to the exposure to stress....

PROTEIN ENRICHMENT OF SPENT SORGHUM RESIDUE USING ...

African Journals Online (AJOL)

BSN

The optimum concentration of spent sorghum for protein enrichment with S. cerevisiae was 7.Sg/100 ml. Th.: protein ... production of single sell protein using Candida utilis and cassava starch effluem as substrate. ... wastes as substrates, Kluyveromyces fragilis and milk whey coconut water as substrate (Rahmat et al.,. 1995 ...

CRISPRpred: A flexible and efficient tool for sgRNAs on-target activity prediction in CRISPR/Cas9 systems.

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Md Khaledur Rahman

Full Text Available The CRISPR/Cas9-sgRNA system has recently become a popular tool for genome editing and a very hot topic in the field of medical research. In this system, Cas9 protein is directed to a desired location for gene engineering and cleaves target DNA sequence which is complementary to a 20-nucleotide guide sequence found within the sgRNA. A lot of experimental efforts, ranging from in vivo selection to in silico modeling, have been made for efficient designing of sgRNAs in CRISPR/Cas9 system. In this article, we present a novel tool, called CRISPRpred, for efficient in silico prediction of sgRNAs on-target activity which is based on the applications of Support Vector Machine (SVM model. To conduct experiments, we have used a benchmark dataset of 17 genes and 5310 guide sequences where there are only 20% true values. CRISPRpred achieves Area Under Receiver Operating Characteristics Curve (AUROC-Curve, Area Under Precision Recall Curve (AUPR-Curve and maximum Matthews Correlation Coefficient (MCC as 0.85, 0.56 and 0.48, respectively. Our tool shows approximately 5% improvement in AUPR-Curve and after analyzing all evaluation metrics, we find that CRISPRpred is better than the current state-of-the-art. CRISPRpred is enough flexible to extract relevant features and use them in a learning algorithm. The source code of our entire software with relevant dataset can be found in the following link: https://github.com/khaled-buet/CRISPRpred.

Endoplasmic reticulum stress and N-glycosylation modulate expression of WFS1 protein

International Nuclear Information System (INIS)

Yamaguchi, Suguru; Ishihara, Hisamitsu; Tamura, Akira; Yamada, Takahiro; Takahashi, Rui; Takei, Daisuke; Katagiri, Hideki; Oka, Yoshitomo

2004-01-01

Mutations of the WFS1 gene are responsible for two hereditary diseases, Wolfram syndrome and low frequency sensorineural hearing loss. The WFS1 protein is a glycoprotein located in the endoplasmic reticulum (ER) membrane but its function is poorly understood. Herein we show WFS1 mRNA and protein levels in pancreatic islets to be increased with ER-stress inducers, thapsigargin and dithiothreitol. Another ER-stress inducer, the N-glycosylation inhibitor tunicamycin, also raised WFS1 mRNA but not protein levels. Site-directed mutagenesis showed both Asn-663 and Asn-748 to be N-glycosylated in mouse WFS1 protein. The glycosylation-defective WFS1 protein, in which Asn-663 and Asn-748 had been substituted with aspartate, exhibited an increased protein turnover rate. Consistent with this, the WFS1 protein was more rapidly degraded in the presence of tunicamycin. These data indicate that ER-stress and N-glycosylation play important roles in WFS1 expression and stability, and also suggest regulatory roles for this protein in ER-stress induced cell death

Identification of genes and pathways associated with aluminum stress and tolerance using transcriptome profiling of wheat near-isogenic lines

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Diallo Amadou

2008-08-01

Full Text Available Abstract Background Aluminum is considered the most limiting factor for plant productivity in acidic soils, which cover large areas of the world's potential arable lands. The inhibition of root growth is recognized as the primary effect of Al toxicity. To identify genes associated with Al stress and tolerance, transcriptome analyses of four different wheat lines (2 Al-tolerant and 2 Al sensitive that differ in their response to Al were performed. Results Microarray expression profiling revealed that 83 candidate genes are associated with Al stress and 25 are associated with tolerance. The stress-associated genes include important enzymes such as pyruvate dehydrogenase, alternative oxidase, and galactonolactone oxidase, ABC transporter and ascorbate oxido-reducatase. The Al tolerance-associated genes include the ALMT-1 malate transporter, glutathione S-transferase, germin/oxalate oxidase, fructose 1,6-bisphosphatase, cysteine-rich proteins, cytochrome P450 monooxygenase, cellulose synthase, zinc finger transcription factor, disease resistance response protein and F-box containing domain protein. Conclusion In this survey, we identified stress- and tolerance-associated genes that may be involved in the detoxification of Al and reactive oxygen species. Alternative pathways could help maintain the supply of important metabolites (H2O2, ascorbate, NADH, and phosphate needed for Al tolerance and root growth. The Al tolerance-associated genes may be key factors that regulate these pathways.

Stress tolerances of nullmutants of function-unknown genes encoding menadione stress-responsive proteins in Aspergillus nidulans.

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Leiter, Éva; Bálint, Mihály; Miskei, Márton; Orosz, Erzsébet; Szabó, Zsuzsa; Pócsi, István

2016-07-01

A group of menadione stress-responsive function-unkown genes of Aspergillus nidulans (Locus IDs ANID_03987.1, ANID_06058.1, ANID_10219.1, and ANID_10260.1) was deleted and phenotypically characterized. Importantly, comparative and phylogenetic analyses of the tested A. nidulans genes and their orthologs shed light only on the presence of a TANGO2 domain with NRDE protein motif in the translated ANID_06058.1 gene but did not reveal any recognizable protein-encoding domains in other protein sequences. The gene deletion strains were subjected to oxidative, osmotic, and metal ion stress and, surprisingly, only the ΔANID_10219.1 mutant showed an increased sensitivity to 0.12 mmol l(-1) menadione sodium bisulfite. The gene deletions affected the stress sensitivities (tolerances) irregularly, for example, some strains grew more slowly when exposed to various oxidants and/or osmotic stress generating agents, meanwhile the ΔANID_10260.1 mutant possessed a wild-type tolerance to all stressors tested. Our results are in line with earlier studies demonstrating that the deletions of stress-responsive genes do not confer necessarily any stress-sensitivity phenotypes, which can be attributed to compensatory mechanisms based on other elements of the stress response system with overlapping functions. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Cold and Heat Stress Diversely Alter Both Cauliflower Respiration and Distinct Mitochondrial Proteins Including OXPHOS Components and Matrix Enzymes

Science.gov (United States)

Rurek, Michał; Czołpińska, Magdalena; Pawłowski, Tomasz Andrzej; Krzesiński, Włodzimierz; Spiżewski, Tomasz

2018-01-01

Complex proteomic and physiological approaches for studying cold and heat stress responses in plant mitochondria are still limited. Variations in the mitochondrial proteome of cauliflower (Brassica oleracea var. botrytis) curds after cold and heat and after stress recovery were assayed by two-dimensional polyacrylamide gel electrophoresis (2D PAGE) in relation to mRNA abundance and respiratory parameters. Quantitative analysis of the mitochondrial proteome revealed numerous stress-affected protein spots. In cold, major downregulations in the level of photorespiratory enzymes, porine isoforms, oxidative phosphorylation (OXPHOS) and some low-abundant proteins were observed. In contrast, carbohydrate metabolism enzymes, heat-shock proteins, translation, protein import, and OXPHOS components were involved in heat response and recovery. Several transcriptomic and metabolic regulation mechanisms are also suggested. Cauliflower plants appeared less susceptible to heat; closed stomata in heat stress resulted in moderate photosynthetic, but only minor respiratory impairments, however, photosystem II performance was unaffected. Decreased photorespiration corresponded with proteomic alterations in cold. Our results show that cold and heat stress not only operate in diverse modes (exemplified by cold-specific accumulation of some heat shock proteins), but exert some associations at molecular and physiological levels. This implies a more complex model of action of investigated stresses on plant mitochondria. PMID:29547512

Nitrosative stress and nitrated proteins in trichloroethene-mediated autoimmunity.

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Gangduo Wang

Full Text Available Exposure to trichloroethene (TCE, a ubiquitous environmental contaminant, has been linked to a variety of autoimmune diseases (ADs including SLE, scleroderma and hepatitis. Mechanisms involved in the pathogenesis of ADs are largely unknown. Earlier studies from our laboratory in MRL+/+ mice suggested the contribution of oxidative/nitrosative stress in TCE-induced autoimmunity, and N-acetylcysteine (NAC supplementation provided protection by attenuating oxidative stress. This study was undertaken to further evaluate the contribution of nitrosative stress in TCE-mediated autoimmunity and to identify proteins susceptible to nitrosative stress. Groups of female MRL +/+ mice were given TCE, NAC or TCE + NAC for 6 weeks (TCE, 10 mmol/kg, i.p., every 4th day; NAC, ∼ 250 mg/kg/day via drinking water. TCE exposure led to significant increases in serum anti-nuclear and anti-histone antibodies together with significant induction of iNOS and increased formation of nitrotyrosine (NT in sera and livers. Proteomic analysis identified 14 additional nitrated proteins in the livers of TCE-treated mice. Furthermore, TCE exposure led to decreased GSH levels and increased activation of NF-κB. Remarkably, NAC supplementation not only ameliorated TCE-induced nitrosative stress as evident from decreased iNOS, NT, nitrated proteins, NF-κB p65 activation and increased GSH levels, but also the markers of autoimmunity, as evident from decreased levels of autoantibodies in the sera. These findings provide support to the role of nitrosative stress in TCE-mediated autoimmune response and identify specific nitrated proteins which could have autoimmune potential. Attenuation of TCE-induced autoimmunity in mice by NAC provides an approach for designing therapeutic strategies.

Rotavirus replication is correlated with S/G2 interphase arrest of the host cell cycle.

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Selene Glück

Full Text Available In infected cells rotavirus (RV replicates in viroplasms, cytosolic structures that require a stabilized microtubule (MT network for their assembly, maintenance of the structure and perinuclear localization. Therefore, we hypothesized that RV could interfere with the MT-breakdown that takes place in mitosis during cell division. Using synchronized RV-permissive cells, we show that RV infection arrests the cell cycle in S/G2 phase, thus favoring replication by improving viroplasms formation, viral protein translation, and viral assembly. The arrest in S/G2 phase is independent of the host or viral strain and relies on active RV replication. RV infection causes cyclin B1 down-regulation, consistent with blocking entry into mitosis. With the aid of chemical inhibitors, the cytoskeleton network was linked to specific signaling pathways of the RV-induced cell cycle arrest. We found that upon RV infection Eg5 kinesin was delocalized from the pericentriolar region to the viroplasms. We used a MA104-Fucci system to identify three RV proteins (NSP3, NSP5, and VP2 involved in cell cycle arrest in the S-phase. Our data indicate that there is a strong correlation between the cell cycle arrest and RV replication.

Deficiency of heat shock transcription factor 1 suppresses heat stress-associated increase in slow soleus muscle mass of mice.

Science.gov (United States)

Ohno, Y; Egawa, T; Yokoyama, S; Nakai, A; Sugiura, T; Ohira, Y; Yoshioka, T; Goto, K

2015-12-01

Effects of heat shock transcription factor 1 (HSF1) deficiency on heat stress-associated increase in slow soleus muscle mass of mice were investigated. Both HSF1-null and wild-type mice were randomly assigned to control and heat-stressed groups. Mice in heat-stressed group were exposed to heat stress (41 °C for 60 min) in an incubator without anaesthesia. Significant increase in wet and dry weights, and protein content of soleus muscle in wild-type mice was observed seven days after the application of the heat stress. However, heat stress had no impact on soleus muscle mass in HSF1-null mice. Neither type of mice exhibited much effect of heat stress on HSF mRNA expression (HSF1, HSF2 and HSF4). On the other hand, heat stress upregulated heat shock proteins (HSPs) at the mRNA (HSP72) and protein (HSP72 and HSP110) levels in wild-type mice, but not in HSF1-null mice. The population of Pax7-positive nuclei relative to total myonuclei of soleus muscle in wild-type mice was significantly increased by heat stress, but not in HSF1-null mice. Furthermore, the absence of HSF1 gene suppressed heat stress-associated phosphorylation of Akt and p70 S6 kinase (p-p70S6K) in soleus muscle. Heat stress-associated increase in skeletal muscle mass may be induced by HSF1 and/or HSF1-mediated stress response that activates muscle satellite cells and Akt/p70S6K signalling pathway. © 2015 Scandinavian Physiological Society. Published by John Wiley & Sons Ltd.

Experimental residual stress evaluation of hydraulic expansion transitions in Alloy 690 steam generator tubing

International Nuclear Information System (INIS)

McGregor, R.; Doherty, P.; Hornbach, D.; Abdelsalam, U.

1995-01-01

Nuclear Steam Generator (SG) service reliability and longevity have been seriously affected worldwide by corrosion at the tube-to-tubesheet joint expansion. Current SG designs for new facilities and replacement projects enhance corrosion resistance through the use of advanced tubing materials and improved joint design and fabrication techniques. Here, transition zones of hydraulic expansions have undergone detailed experimental evaluation to define residual stress and cold-work distribution on and below the secondary-side surface. Using X-ray diffraction techniques, with supporting finite element analysis, variations are compared in tubing metallurgical condition, tube/pitch geometry, expansion pressure, and tube-to-hole clearance. Initial measurements to characterize the unexpanded tube reveal compressive stresses associated with a thin work-hardened layer on the outer surface of the tube. The gradient of cold-work was measured as 3% to 0% within .001 inch of the surface. The levels and character of residual stresses following hydraulic expansion are primarily dependent on this work-hardened surface layer and initial stress state that is unique to each tube fabrication process. Tensile stresses following expansion are less than 25% of the local yield stress and are found on the transition in a narrow circumferential band at the immediate tube surface (< .0002 inch/0.005 mm depth). The measurements otherwise indicate a predominance of compressive stresses on and below the secondary-side surface of the transition zone. Excellent resistance to SWSCC initiation is offered by the low levels of tensile stress and cold-work. Propagation of any possible cracking would be deterred by the compressive stress field that surrounds this small volume of tensile material

The Stress Granule RNA-Binding Protein TIAR-1 Protects Female Germ Cells from Heat Shock in Caenorhabditis elegans

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Gabriela Huelgas-Morales

2016-04-01

Full Text Available In response to stressful conditions, eukaryotic cells launch an arsenal of regulatory programs to protect the proteome. One major protective response involves the arrest of protein translation and the formation of stress granules, cytoplasmic ribonucleoprotein complexes containing the conserved RNA-binding proteins TIA-1 and TIAR. The stress granule response is thought to preserve mRNA for translation when conditions improve. For cells of the germline—the immortal cell lineage required for sexual reproduction—protection from stress is critically important for perpetuation of the species, yet how stress granule regulatory mechanisms are deployed in animal reproduction is incompletely understood. Here, we show that the stress granule protein TIAR-1 protects the Caenorhabditis elegans germline from the adverse effects of heat shock. Animals containing strong loss-of-function mutations in tiar-1 exhibit significantly reduced fertility compared to the wild type following heat shock. Analysis of a heat-shock protein promoter indicates that tiar-1 mutants display an impaired heat-shock response. We observed that TIAR-1 was associated with granules in the gonad core and oocytes during several stressful conditions. Both gonad core and oocyte granules are dynamic structures that depend on translation; protein synthesis inhibitors altered their formation. Nonetheless, tiar-1 was required for the formation of gonad core granules only. Interestingly, the gonad core granules did not seem to be needed for the germ cells to develop viable embryos after heat shock. This suggests that TIAR-1 is able to protect the germline from heat stress independently of these structures.

The Stress Granule RNA-Binding Protein TIAR-1 Protects Female Germ Cells from Heat Shock in Caenorhabditis elegans.

Science.gov (United States)

Huelgas-Morales, Gabriela; Silva-García, Carlos Giovanni; Salinas, Laura S; Greenstein, David; Navarro, Rosa E

2016-04-07

In response to stressful conditions, eukaryotic cells launch an arsenal of regulatory programs to protect the proteome. One major protective response involves the arrest of protein translation and the formation of stress granules, cytoplasmic ribonucleoprotein complexes containing the conserved RNA-binding proteins TIA-1 and TIAR. The stress granule response is thought to preserve mRNA for translation when conditions improve. For cells of the germline-the immortal cell lineage required for sexual reproduction-protection from stress is critically important for perpetuation of the species, yet how stress granule regulatory mechanisms are deployed in animal reproduction is incompletely understood. Here, we show that the stress granule protein TIAR-1 protects the Caenorhabditis elegans germline from the adverse effects of heat shock. Animals containing strong loss-of-function mutations in tiar-1 exhibit significantly reduced fertility compared to the wild type following heat shock. Analysis of a heat-shock protein promoter indicates that tiar-1 mutants display an impaired heat-shock response. We observed that TIAR-1 was associated with granules in the gonad core and oocytes during several stressful conditions. Both gonad core and oocyte granules are dynamic structures that depend on translation; protein synthesis inhibitors altered their formation. Nonetheless, tiar-1 was required for the formation of gonad core granules only. Interestingly, the gonad core granules did not seem to be needed for the germ cells to develop viable embryos after heat shock. This suggests that TIAR-1 is able to protect the germline from heat stress independently of these structures. Copyright © 2016 Huelgas-Morales et al.

On the dynamical incompleteness of the Protein Data Bank.

Science.gov (United States)

Marino-Buslje, Cristina; Monzon, Alexander Miguel; Zea, Diego Javier; Fornasari, María Silvina; Parisi, Gustavo

2017-08-02

Major scientific challenges that are beyond the capability of individuals need to be addressed by multi-disciplinary and multi-institutional consortia. Examples of these endeavours include the Human Genome Project, and more recently, the Structural Genomics (SG) initiative. The SG initiative pursues the expansion of structural coverage to include at least one structural representative for each protein family to derive the remaining structures using homology modelling. However, biological function is inherently connected with protein dynamics that can be studied by knowing different structures of the same protein. This ensemble of structures provides snapshots of protein conformational diversity under native conditions. Thus, sequence redundancy in the Protein Data Bank (PDB) (i.e. crystallization of the same protein under different conditions) is therefore an essential input contributing to experimentally based studies of protein dynamics and providing insights into protein function. In this work, we show that sequence redundancy, a key concept for exploring protein dynamics, is highly biased and fundamentally incomplete in the PDB. Additionally, our results show that dynamical behaviour of proteins cannot be inferred using homologous proteins. Minor to moderate changes in sequence can produce great differences in dynamical behaviour. Nonetheless, the structural and dynamical incompleteness of the PDB is apparently unrelated concepts in SG. While the first could be reversed by promoting the extension of the structural coverage, we would like to emphasize that further focused efforts will be needed to amend the incompleteness of the PDB in terms of dynamical information content, essential to fully understand protein function. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Burst pressure and leak rate from fretted SG tubes

International Nuclear Information System (INIS)

Hwang, Seong Sik; Jung, Man Kyo; Kim, Hong Pyo; Kim, Joung Soo

2005-01-01

Steam generator(SG) tubes of a pressurized water reactor(PWR) have suffered from various types of corrosion, such as pitting, wastage and stress corrosion cracking (SCC) on both the primary and secondary side. Recently, fretting/wear degradation at the tube support region has been reported in some Korean nuclear power plants. In order to prevent the primary coolant from leaking to the secondary side, the tubes are repaired by a sleeving or plugging. It is important to establish the repair criteria to assure a reactor integrity and yet maintain the plugging ratio within the limits needed for an efficient operation. The objective of the burst test is to obtain a relationship between the burst/leak rate and the shape of the fretted flaws machined with an electro discharge machining (EDM)

Sol-Generating Chemical Vapor into Liquid (SG-CViL) deposition – a facile method for encapsulation of diverse cell types in silica matrices

Energy Technology Data Exchange (ETDEWEB)

Johnston, Robert [New Mexico Inst. of Mining and Technology, Socorro, NM (United States). Materials Engineering Dept.; Rogelj, Snezna [New Mexico Inst. of Mining and Technology, Socorro, NM (United States). Biology Dept.; Harper, Jason C. [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States). Bioenergy and Biodefense Technologies Dept.; Tartis, Michaelann [New Mexico Inst. of Mining and Technology, Socorro, NM (United States). Materials and Chemical Engineering Dept.

2014-12-12

In nature, cells perform a variety of complex functions such as sensing, catalysis, and energy conversion which hold great potential for biotechnological device construction. However, cellular sensitivity to ex vivo environments necessitates development of bio–nano interfaces which allow integration of cells into devices and maintain their desired functionality. In order to develop such an interface, the use of a novel Sol-Generating Chemical Vapor into Liquid (SG-CViL) deposition process for whole cell encapsulation in silica was explored. In SG-CViL, the high vapor pressure of tetramethyl orthosilicate (TMOS) is utilized to deliver silica into an aqueous medium, creating a silica sol. Cells are then mixed with the resulting silica sol, facilitating encapsulation of cells in silica while minimizing cell contact with the cytotoxic products of silica generating reactions (i.e. methanol), and reduce exposure of cells to compressive stresses induced from silica condensation reactions. Using SG-CVIL, Saccharomyces cerevisiae (S. cerevisiae) engineered with an inducible beta galactosidase system were encapsulated in silica solids and remained both viable and responsive 29 days post encapsulation. By tuning SG-CViL parameters, thin layer silica deposition on mammalian HeLa and U87 human cancer cells was also achieved. Thus, the ability to encapsulate various cell types in either a multi cell (S. cerevisiae) or a thin layer (HeLa and U87 cells) fashion shows the promise of SG-CViL as an encapsulation strategy for generating cell–silica constructs with diverse functions for incorporation into devices for sensing, bioelectronics, biocatalysis, and biofuel applications.

Absence of Wdr13 Gene Predisposes Mice to Mild Social Isolation – Chronic Stress, Leading to Depression-Like Phenotype Associated With Differential Expression of Synaptic Proteins

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Mitra, Shiladitya; Sameer Kumar, Ghantasala S.; Jyothi Lakshmi, B.; Thakur, Suman; Kumar, Satish

2018-01-01

We earlier reported that the male mice lacking the Wdr13 gene (Wdr13-/0) showed mild anxiety, better memory retention, and up-regulation of synaptic proteins in the hippocampus. With increasing evidences from parallel studies in our laboratory about the possible role of Wdr13 in stress response, we investigated its role in brain. We observed that Wdr13 transcript gets up-regulated in the hippocampus of the wild-type mice exposed to stress. To further dissect its function, we analyzed the behavioral and molecular phenotypes of Wdr13-/0 mice when subjected to mild chronic psychological stress, namely; mild (attenuated) social isolation. We employed iTRAQ based quantitative proteomics, real time PCR and western blotting to investigate molecular changes. Three weeks of social isolation predisposed Wdr13-/0 mice to anhedonia, heightened anxiety-measured by Open field test (OFT), increased behavior despair- measured by Forced swim test (FST) and reduced dendritic branching along with decreased spine density of hippocampal CA1 neurons as compared to wild-type counterparts. This depression-like-phenotype was however ameliorated when treated with anti-depressant imipramine. Molecular analysis revealed that out of 1002 quantified proteins [1% False discovery rate (FDR), at-least two unique peptides], strikingly, a significant proportion of synaptic proteins including, SYN1, CAMK2A, and RAB3A were down-regulated in the socially isolated Wdr13-/0 mice as compared to its wild-type counterparts. This was in contrast to the elevated levels of these proteins in non-stressed mutants as compared to the controls. We hypothesized that a de-regulated transcription factor upstream of the synaptic genes might be responsible for the observed phenotype. Indeed, in the socially isolated Wdr13-/0 mice, there was an up-regulation of GATA1 – a transcription factor that negatively regulates synaptic genes and has been associated with Major Depression (MD) in humans. The present study

Preliminary analysis of cold stress responsive proteins in Mesocestoides corti larvae.

Science.gov (United States)

Canclini, Lucía; Esteves, Adriana

2007-07-01

Many parasites undergo sudden changes in environmental conditions at some stage during their life cycle. The molecular response to this variation is characterised by a rapid transcriptional activation of a specific set of genes coding for proteins generically known as stress proteins. They appear to be also involved in various biological processes including cell proliferation and differentiation. The platyhelminth parasite, Mesocestoides corti (Cestoda) presents important properties as a model organism. Under stress conditions, key molecules involved in metabolic pathways as well as in the growth and differentiation of the parasite can be identified. 2D protein expression profile of tetrathyridia of M. corti, submitted to nutritional starvation and cold stress is described, as well as the recovery pattern. A set of specifically expressed proteins was observed in each experimental condition. Quantitative and qualitative differences and stress recovery pattern are also reported. This work makes evident the high plasticity and resistance to extreme environmental conditions of these parasites at the molecular level.

Optimum Frequency for Eddy Current Testing Method of SMART SG tubes

International Nuclear Information System (INIS)

Lee, Yoon Sang; Jung, Hyun Kyu; Choung, Yun Hang

2009-01-01

The SMART SG tubes will be made of Alloy 690. The outside diameter will be 17 mm and the thickness will be 2.5 mm. They will be assembled helically around, and their innermost diameter will be about 600 mm and the total length will be about 32 meters. For safety, SMART SG tubes are designed for use with thick tubes such as 2.5 mm thickness compared to about 1 mm thickness of normal Korean standard pressurized water reactor tubes. Due to using thick tubes such as 2.5 mm varieties, it was doubted that the Eddy Current Testing Method (ECT) would be a feasible method. Therefore we are trying to simulate the bobbin probe signal for SMART SG tubes and comparing it to PWR SG ECT probe signal using VIM software, checking for the applicability of ECT. Also we are trying to compare the ECT signal of 2.5 mm thick stainless tubes to check if they are possible substitute material

Effect of the unfolded protein response on ER protein export: a potential new mechanism to relieve ER stress.

Science.gov (United States)

Shaheen, Alaa

2018-05-05

The unfolded protein response (UPR) is an adaptive cellular response that aims to relieve endoplasmic reticulum (ER) stress via several mechanisms, including inhibition of protein synthesis and enhancement of protein folding and degradation. There is a controversy over the effect of the UPR on ER protein export. While some investigators suggested that ER export is inhibited during ER stress, others suggested the opposite. In this article, their conflicting studies are analyzed and compared in attempt to solve this controversy. The UPR appears indeed to enhance ER export, possibly via multiple mechanisms. However, another factor, which is the integrity of the folding machinery/environment inside ER, determines whether ER export will appear increased or decreased during experimentation. Also, different methods of stress induction appear to have different effects on ER export. Thus, improvement of ER export may represent a new mechanism by which the UPR alleviates ER stress. This may help researchers to understand how the UPR works inside cells and how to manipulate it to alter cell fate during stress, either to promote cell survival or death. This may open up new approaches for the treatment of ER stress-related diseases.

Anti-Apoptotic Protein Bcl-xL Expression in the Midbrain Raphe Region Is Sensitive to Stress and Glucocorticoids.

Science.gov (United States)

Shishkina, Galina T; Kalinina, Tatyana S; Bulygina, Veta V; Lanshakov, Dmitry A; Babluk, Ekaterina V; Dygalo, Nikolay N

2015-01-01

Anti-apoptotic proteins are suggested to be important for the normal health of neurons and synapses as well as for resilience to stress. In order to determine whether stressful events may influence the expression of anti-apoptotic protein Bcl-xL in the midbrain and specifically in the midbrain serotonergic (5-HT) neurons involved in neurobehavioral responses to adverse stimuli, adult male rats were subjected to short-term or chronic forced swim stress. A short-term stress rapidly increased the midbrain bcl-xl mRNA levels and significantly elevated Bcl-xL immunoreactivity in the midbrain 5-HT cells. Stress-induced increase in glucocorticoid secretion was implicated in the observed effect. The levels of bcl-xl mRNA were decreased after stress when glucocorticoid elevation was inhibited by metyrapone (MET, 150 mg/kg), and this decrease was attenuated by glucocorticoid replacement with dexamethasone (DEX; 0.2 mg/kg). Both short-term stress and acute DEX administration, in parallel with Bcl-xL, caused a significant increase in tph2 mRNA levels and slightly enhanced tryptophan hydroxylase immunoreactivity in the midbrain. The increasing effect on the bcl-xl expression was specific to the short-term stress. Forced swim repeated daily for 2 weeks led to a decrease in bcl-xl mRNA in the midbrain without any effects on the Bcl-xL protein expression in the 5-HT neurons. In chronically stressed animals, an increase in tph2 gene expression was not associated with any changes in tryptophan hydroxylase protein levels. Our findings are the first to demonstrate that both short-term stress and acute glucocorticoid exposures induce Bcl-xL protein expression in the midbrain 5-HT neurons concomitantly with the activation of the 5-HT synthesis pathway in these neurons.

Anti-Apoptotic Protein Bcl-xL Expression in the Midbrain Raphe Region Is Sensitive to Stress and Glucocorticoids.

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Galina T Shishkina

Full Text Available Anti-apoptotic proteins are suggested to be important for the normal health of neurons and synapses as well as for resilience to stress. In order to determine whether stressful events may influence the expression of anti-apoptotic protein Bcl-xL in the midbrain and specifically in the midbrain serotonergic (5-HT neurons involved in neurobehavioral responses to adverse stimuli, adult male rats were subjected to short-term or chronic forced swim stress. A short-term stress rapidly increased the midbrain bcl-xl mRNA levels and significantly elevated Bcl-xL immunoreactivity in the midbrain 5-HT cells. Stress-induced increase in glucocorticoid secretion was implicated in the observed effect. The levels of bcl-xl mRNA were decreased after stress when glucocorticoid elevation was inhibited by metyrapone (MET, 150 mg/kg, and this decrease was attenuated by glucocorticoid replacement with dexamethasone (DEX; 0.2 mg/kg. Both short-term stress and acute DEX administration, in parallel with Bcl-xL, caused a significant increase in tph2 mRNA levels and slightly enhanced tryptophan hydroxylase immunoreactivity in the midbrain. The increasing effect on the bcl-xl expression was specific to the short-term stress. Forced swim repeated daily for 2 weeks led to a decrease in bcl-xl mRNA in the midbrain without any effects on the Bcl-xL protein expression in the 5-HT neurons. In chronically stressed animals, an increase in tph2 gene expression was not associated with any changes in tryptophan hydroxylase protein levels. Our findings are the first to demonstrate that both short-term stress and acute glucocorticoid exposures induce Bcl-xL protein expression in the midbrain 5-HT neurons concomitantly with the activation of the 5-HT synthesis pathway in these neurons.

Overexpression of a Cytosolic Abiotic Stress Responsive Universal Stress Protein (SbUSP) Mitigates Salt and Osmotic Stress in Transgenic Tobacco Plants

Science.gov (United States)

Udawat, Pushpika; Jha, Rajesh K.; Sinha, Dinkar; Mishra, Avinash; Jha, Bhavanath

2016-01-01

The universal stress protein (USP) is a ubiquitous protein and plays an indispensable role in plant abiotic stress tolerance. The genome of Salicornia brachiata contains two homologs of intron less SbUSP gene which encodes for salt and osmotic responsive USP. In vivo localization reveals that SbUSP is a membrane bound cytosolic protein. The role of the gene was functionally validated by developing transgenic tobacco and compared with control [wild-type (WT) and vector control (VC)] plants under different abiotic stress condition. Transgenic lines (T1) exhibited higher chlorophyll, relative water, proline, total sugar, reducing sugar, free amino acids, polyphenol contents, osmotic potential, membrane stability, and lower electrolyte leakage and lipid peroxidation (malondialdehyde content) under stress treatments than control (WT and VC) plants. Lower accumulation of H2O2 and O2− radicals was also detected in transgenic lines compared to control plants under stress conditions. Present study confers that overexpression of the SbUSP gene enhances plant growth, alleviates ROS buildup, maintains ion homeostasis and improves the physiological status of the plant under salt and osmotic stresses. Principal component analysis exhibited a statistical distinction of plant response to salinity stress, and a significant response was observed for transgenic lines under stress, which provides stress endurance to the plant. A possible signaling role is proposed that some downstream genes may get activated by abiotic stress responsive cytosolic SbUSP, which leads to the protection of cell from oxidative damages. The study unveils that ectopic expression of the gene mitigates salt or osmotic stress by scavenging ROS and modulating the physiological process of the plant. PMID:27148338

Relationship of oxidative stress in skeletal muscle with obesity and obesity-associated hyperinsulinemia in horses.

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Banse, Heidi E; Frank, Nicholas; Kwong, Grace P S; McFarlane, Dianne

2015-10-01

In horses, hyperinsulinemia and insulin resistance (insulin dysregulation) are associated with the development of laminitis. Although obesity is associated with insulin dysregulation, the mechanism of obesity-associated insulin dysregulation remains to be established. We hypothesized that oxidative stress in skeletal muscle is associated with obesity-associated hyperinsulinemia in horses. Thirty-five light breed horses with body condition scores (BCS) of 3/9 to 9/9 were studied, including 7 obese, normoinsulinemic (BCS ≥ 7, resting serum insulin obese, hyperinsulinemic (resting serum insulin ≥ 30 μIU/mL) horses. Markers of oxidative stress (oxidative damage, mitochondrial function, and antioxidant capacity) were evaluated in skeletal muscle biopsies. A Spearman's rank correlation coefficient was used to determine relationships between markers of oxidative stress and BCS. Furthermore, to assess the role of oxidative stress in obesity-related hyperinsulinemia, markers of antioxidant capacity and oxidative damage were compared among lean, normoinsulinemic (L-NI); obese, normoinsulinemic (O-NI); and obese, hyperinsulinemic (O-HI) horses. Increasing BCS was associated with an increase in gene expression of a mitochondrial protein responsible for mitochondrial biogenesis (estrogen-related receptor alpha, ERRα) and with increased antioxidant enzyme total superoxide dismutase (TotSOD) activity. When groups (L-NI, O-NI, and O-HI) were compared, TotSOD activity was increased and protein carbonyls, a marker of oxidative damage, decreased in the O-HI compared to the L-NI horses. These findings suggest that a protective antioxidant response occurred in the muscle of obese animals and that obesity-associated oxidative damage in skeletal muscle is not central to the pathogenesis of equine hyperinsulinemia.

Calcium affecting protein expression in longan under simulated acid rain stress.

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Pan, Tengfei; Li, Yongyu; Ma, Cuilan; Qiu, Dongliang

2015-08-01

Longan (Dimocarpus longana Lour. cv. Wulongling) of uniform one-aged seedlings grown in pots were selected to study specific proteins expressed in leaves under simulated acid rain (SiAR) stress and exogenous Ca(2+) regulation. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) results showed that there was a protein band specifically expressed under SiAR of pH 2.5 stress for 15 days with its molecular weight of about 23 kD. A 17 kD protein band specifically expressed after SiAR stress 5 days. Compared with pH 2.5, the pH 3.5 of SiAR made a less influence to protein expression. Two-dimensional electrophoresis (2-DE) results showed that six new specific proteins including C4 (20.2 kD pI 6.0), F (24 kD pI 6.35), B3 (22.3 kD pI 6.35), B4 (23.5 kD pI 6.5), C5 (21.8 kD pI 5.6), and C6 (20.2 kD pI 5.6) specifically expressed. C4 always expressed during SiAR stress. F expressed under the stress of pH 2.5 for 15 days and expressed in all pH SiAR stress for 20 days. The expression of proteins including B3, C5, and C6 was related to pH value and stress intensity of SiAR. The expression of B4 resulted from synergistic effects of SiAR and Ca. The expression of G1 (Mr 19.3 kD, pI 4.5), G2 (Mr 17.8 kD, pI 4.65), G3 (Mr 16.6 kD, pI 4.6), and G4 (Mr 14.7 kD, pI 4.4) enhanced under the treatment of 5 mM ethylene glycol tetraacetic acid (EGTA) and 2 mM chlorpromazine (CPZ). These proteins showed antagonistic effects and might be relative to the Ca-calmodulin (Ca-CaM) system of longan in response to SiAR stress.

Multidrug resistance-associated protein-1 (MRP1 genetic variants, MRP1 protein levels and severity of COPD

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Rutgers Bea

2010-05-01

Full Text Available Abstract Background Multidrug resistance-associated protein-1 (MRP1 protects against oxidative stress and toxic compounds generated by cigarette smoking, which is the main risk factor for chronic obstructive pulmonary disease (COPD. We have previously shown that single nucleotide polymorphisms (SNPs in MRP1 significantly associate with level of FEV1 in two independent population based cohorts. The aim of our study was to assess the associations of MRP1 SNPs with FEV1 level, MRP1 protein levels and inflammatory markers in bronchial biopsies and sputum of COPD patients. Methods Five SNPs (rs212093, rs4148382, rs504348, rs4781699, rs35621 in MRP1 were genotyped in 110 COPD patients. The effects of MRP1 SNPs were analyzed using linear regression models. Results One SNP, rs212093 was significantly associated with a higher FEV1 level and less airway wall inflammation. Another SNP, rs4148382 was significantly associated with a lower FEV1 level, higher number of inflammatory cells in induced sputum and with a higher MRP1 protein level in bronchial biopsies. Conclusions This is the first study linking MRP1 SNPs with lung function and inflammatory markers in COPD patients, suggesting a role of MRP1 SNPs in the severity of COPD in addition to their association with MRP1 protein level in bronchial biopsies.

Recovery from heat, salt and osmotic stress in Physcomitrella patens requires a functional small heat shock protein PpHsp16.4.

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Ruibal, Cecilia; Castro, Alexandra; Carballo, Valentina; Szabados, László; Vidal, Sabina

2013-11-05

Plant small heat shock proteins (sHsps) accumulate in response to various environmental stresses, including heat, drought, salt and oxidative stress. Numerous studies suggest a role for these proteins in stress tolerance by preventing stress-induced protein aggregation as well as by facilitating protein refolding by other chaperones. However, in vivo evidence for the involvement of sHsps in tolerance to different stress factors is still missing, mainly due to the lack of appropriate mutants in specific sHsp genes. In this study we characterized the function of a sHsp in abiotic stress tolerance in the moss Physcomitrella patens, a model for primitive land plants. Using suppression subtractive hybridization, we isolated an abscisic acid-upregulated gene from P. patens encoding a 16.4 kDa cytosolic class II sHsp. PpHsp16.4 was also induced by salicylic acid, dithiothreitol (DTT) and by exposure to various stimuli, including osmotic and salt stress, but not by oxidative stress-inducing compounds. Expression of the gene was maintained upon stress relief, suggesting a role for this protein in the recovery stage. PpHsp16.4 is encoded by two identical genes arranged in tandem in the genome. Targeted disruption of both genes resulted in the inability of plants to recover from heat, salt and osmotic stress. In vivo localization studies revealed that PpHsp16.4 localized in cytosolic granules in the vicinity of chloroplasts under non stress conditions, suggesting possible distinct roles for this protein under stress and optimal growth. We identified a member of the class II sHsp family that showed hormonal and abiotic stress gene regulation. Induction of the gene by DTT treatment suggests that damaged proteins may act as signals for the stress-induction of PpHsp16.4. The product of this gene was shown to localize in cytosolic granules near the chloroplasts, suggesting a role for the protein in association with these organelles. Our study provides the first direct genetic

Multiple nutrient stresses at intersecting Pacific Ocean biomes detected by protein biomarkers.

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Saito, Mak A; McIlvin, Matthew R; Moran, Dawn M; Goepfert, Tyler J; DiTullio, Giacomo R; Post, Anton F; Lamborg, Carl H

2014-09-05

Marine primary productivity is strongly influenced by the scarcity of required nutrients, yet our understanding of these nutrient limitations is informed by experimental observations with sparse geographical coverage and methodological limitations. We developed a quantitative proteomic method to directly assess nutrient stress in high-light ecotypes of the abundant cyanobacterium Prochlorococcus across a meridional transect in the central Pacific Ocean. Multiple peptide biomarkers detected widespread and overlapping regions of nutritional stress for nitrogen and phosphorus in the North Pacific Subtropical Gyre and iron in the equatorial Pacific. Quantitative protein analyses demonstrated simultaneous stress for these nutrients at biome interfaces. This application of proteomic biomarkers to diagnose ocean metabolism demonstrated Prochlorococcus actively and simultaneously deploying multiple biochemical strategies for low-nutrient conditions in the oceans. Copyright © 2014, American Association for the Advancement of Science.

Occupational role stress is associated with higher cortisol reactivity to acute stress.

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Wirtz, Petra H; Ehlert, Ulrike; Kottwitz, Maria U; La Marca, Roberto; Semmer, Norbert K

2013-04-01

We investigated whether occupational role stress is associated with differential levels of the stress hormone cortisol in response to acute psychosocial stress. Forty-three medication-free nonsmoking men aged between 22 and 65 years (mean ± SEM: 44.5 ± 2) underwent an acute standardized psychosocial stress task combining public speaking and mental arithmetic in front of an audience. We assessed occupational role stress in terms of role conflict and role ambiguity (combined into a measure of role uncertainty) as well as further work characteristics and psychological control variables including time pressure, overcommitment, perfectionism, and stress appraisal. Moreover, we repeatedly measured salivary cortisol and blood pressure levels before and after stress exposure, and several times up to 60 min thereafter. Higher role uncertainty was associated with a more pronounced cortisol stress reactivity (p = .016), even when controlling for the full set of potential confounders (p stress reactivity was not associated with role uncertainty. Our findings suggest that occupational role stress in terms of role uncertainty acts as a background stressor that is associated with increased HPA-axis reactivity to acute stress. This finding may represent a potential mechanism regarding how occupational role stress may precipitate adverse health outcomes.

ER stress-induced protein, VIGG, disturbs plant cation homeostasis, which is correlated with growth retardation and robustness to ER stress

International Nuclear Information System (INIS)

Katoh, Hironori; Fujita, Keiko; Takuhara, Yuki; Ogawa, Atsushi; Suzuki, Shunji

2011-01-01

Highlights: → VIGG is an ER stress-induced protein in plant. → We examine the characteristics of VIGG-overexpressing Arabidopsis plants. → VIGG-overexpressing plants reveal growth retardation and robustness to ER stress. → VIGG disturbs cation homeostasis in plant. -- Abstract: VIGG is a putative endoplasmic reticulum (ER) resident protein induced by virus infection and ER stress, and is correlated with fruit quality in grapevine. The present study was undertaken to determine the biological function of VIGG in grapevine. Experiments using fluorescent protein-VIGG fusion protein demonstrated that VIGG is localized in ER and the ER targeting sequence is in the N-terminus. The overexpression of VIGG in Arabidopsis plant led to growth retardation. The rosette leaves of VIGG-overexpressing plants were smaller than those of the control plants and rolled at 42 days after seeding. VIGG-overexpressing plants revealed robustness to ER stress as well as the low expression of ER stress marker proteins, such as the luminal binding proteins. These characteristics of VIGG-overexpressing plants were supported by a microarray experiment that demonstrated the disruption of genes related to ER stress response and flowering, as well as cation mobility, in the plants. Finally, cation homeostasis in the plants was disturbed by the overexpression of VIGG. Taken together, these results suggest that VIGG may disturb cation homeostasis in plant, which is correlated with the robustness to ER stress and growth retardation.

ER stress-induced protein, VIGG, disturbs plant cation homeostasis, which is correlated with growth retardation and robustness to ER stress

Energy Technology Data Exchange (ETDEWEB)

Katoh, Hironori; Fujita, Keiko; Takuhara, Yuki [Laboratory of Fruit Genetic Engineering, The Institute of Enology and Viticulture, University of Yamanashi, Kofu, Yamanashi 400-0005 (Japan); Ogawa, Atsushi [Department of Biological Production, Akita Prefectural University, Shimosinjyou-nakano 241-438, Akita 010-0195 (Japan); Suzuki, Shunji, E-mail: suzukis@yamanashi.ac.jp [Laboratory of Fruit Genetic Engineering, The Institute of Enology and Viticulture, University of Yamanashi, Kofu, Yamanashi 400-0005 (Japan)

2011-02-18

Highlights: {yields} VIGG is an ER stress-induced protein in plant. {yields} We examine the characteristics of VIGG-overexpressing Arabidopsis plants. {yields} VIGG-overexpressing plants reveal growth retardation and robustness to ER stress. {yields} VIGG disturbs cation homeostasis in plant. -- Abstract: VIGG is a putative endoplasmic reticulum (ER) resident protein induced by virus infection and ER stress, and is correlated with fruit quality in grapevine. The present study was undertaken to determine the biological function of VIGG in grapevine. Experiments using fluorescent protein-VIGG fusion protein demonstrated that VIGG is localized in ER and the ER targeting sequence is in the N-terminus. The overexpression of VIGG in Arabidopsis plant led to growth retardation. The rosette leaves of VIGG-overexpressing plants were smaller than those of the control plants and rolled at 42 days after seeding. VIGG-overexpressing plants revealed robustness to ER stress as well as the low expression of ER stress marker proteins, such as the luminal binding proteins. These characteristics of VIGG-overexpressing plants were supported by a microarray experiment that demonstrated the disruption of genes related to ER stress response and flowering, as well as cation mobility, in the plants. Finally, cation homeostasis in the plants was disturbed by the overexpression of VIGG. Taken together, these results suggest that VIGG may disturb cation homeostasis in plant, which is correlated with the robustness to ER stress and growth retardation.

Increased intracellular proteolysis reduces disease severity in an ER stress-associated dwarfism.

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Mullan, Lorna A; Mularczyk, Ewa J; Kung, Louise H; Forouhan, Mitra; Wragg, Jordan Ma; Goodacre, Royston; Bateman, John F; Swanton, Eileithyia; Briggs, Michael D; Boot-Handford, Raymond P

2017-10-02

The short-limbed dwarfism metaphyseal chondrodysplasia type Schmid (MCDS) is linked to mutations in type X collagen, which increase ER stress by inducing misfolding of the mutant protein and subsequently disrupting hypertrophic chondrocyte differentiation. Here, we show that carbamazepine (CBZ), an autophagy-stimulating drug that is clinically approved for the treatment of seizures and bipolar disease, reduced the ER stress induced by 4 different MCDS-causing mutant forms of collagen X in human cell culture. Depending on the nature of the mutation, CBZ application stimulated proteolysis of misfolded collagen X by either autophagy or proteasomal degradation, thereby reducing intracellular accumulation of mutant collagen. In MCDS mice expressing the Col10a1.pN617K mutation, CBZ reduced the MCDS-associated expansion of the growth plate hypertrophic zone, attenuated enhanced expression of ER stress markers such as Bip and Atf4, increased bone growth, and reduced skeletal dysplasia. CBZ produced these beneficial effects by reducing the MCDS-associated abnormalities in hypertrophic chondrocyte differentiation. Stimulation of intracellular proteolysis using CBZ treatment may therefore be a clinically viable way of treating the ER stress-associated dwarfism MCDS.

Effects of stress and adrenalectomy on activity-regulated cytoskeleton protein (Arc) gene expression

DEFF Research Database (Denmark)

Mikkelsen, Jens D; Larsen, Marianne Hald

2006-01-01

Activity-regulated cytoskeletal-associated protein (Arc) is an effector immediate early gene induced by novelty and involved in consolidation of long-term memory. Since activation of glucocorticoid receptors is a prerequisite for memory consolidation, we therefore aimed to study the effect of acute...... restraint stress on Arc gene expression in adrenalectomized rats. Acute stress produced a significant increase in Arc gene expression in the medial prefrontal cortex, but not in the parietal cortex or in the pyramidal cell layer of the hippocampus. The basal level of Arc mRNA in adrenalectomized animals...... was high in the medial prefrontal cortex and unaffected by acute stress in these animals. These data are consistent with the role of Arc as an integrative modulator of synaptic plasticity by emphasizing the potential role of stress and glucocorticoids in the control of Arc gene expression....

mTORC1 Coordinates Protein Synthesis and Immunoproteasome Formation via PRAS40 to Prevent Accumulation of Protein Stress.

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Yun, Young Sung; Kim, Kwan Hyun; Tschida, Barbara; Sachs, Zohar; Noble-Orcutt, Klara E; Moriarity, Branden S; Ai, Teng; Ding, Rui; Williams, Jessica; Chen, Liqiang; Largaespada, David; Kim, Do-Hyung

2016-02-18

Reduction of translational fidelity often occurs in cells with high rates of protein synthesis, generating defective ribosomal products. If not removed, such aberrant proteins can be a major source of cellular stress causing human diseases. Here, we demonstrate that mTORC1 promotes the formation of immunoproteasomes for efficient turnover of defective proteins and cell survival. mTORC1 sequesters precursors of immunoproteasome β subunits via PRAS40. When activated, mTORC1 phosphorylates PRAS40 to enhance protein synthesis and simultaneously to facilitate the assembly of the β subunits for forming immunoproteasomes. Consequently, the PRAS40 phosphorylations play crucial roles in clearing aberrant proteins that accumulate due to mTORC1 activation. Mutations of RAS, PTEN, and TSC1, which cause mTORC1 hyperactivation, enhance immunoproteasome formation in cells and tissues. Those mutations increase cellular dependence on immunoproteasomes for stress response and survival. These results define a mechanism by which mTORC1 couples elevated protein synthesis with immunoproteasome biogenesis to protect cells against protein stress. Copyright © 2016 Elsevier Inc. All rights reserved.

Stress proteins and phytohormones: their role in formation of plant resistance

International Nuclear Information System (INIS)

Kosakivska, I.V.

2005-01-01

Full text: Using the disc-electrophoresis methods, we have studied protein biosynthesis of different plants, including 11 species of Orchidaceae, some other tropical and subtropical plants, 9 different fruit plants, and 4 cultivars of Triticum aestivum L. under stresses factors such as high and low temperature, clinostating, radioactive irradiation and osmotic shock. Specific and unspecific reactions of plants protein system on stresses were found. De novo synthesis of 35 and 45 kD polypeptides were observed in total and mitochondrial proteins fractions after heat-shock and radioactive irradiation. This suggests that mitochondries participate in formation of plant resistance. Intensive synthesis of ABA revealed as the universal reaction of all studied plants on action of different kinds of stresses. Specific changes in balance of phytohormones were found under different stresses. We observed the correlation between endogenous ABA, IAA and cytokinin level and plant resistance. We also found the interaction between the process of biosynthesis of proteins and phytohormone balance, as well as their direct participation in formation of plant resistance. (author)

Herp enhances ER-associated protein degradation by recruiting ubiquilins

International Nuclear Information System (INIS)

Kim, Tae-Yeon; Kim, Eunmin; Yoon, Sungjoo Kim; Yoon, Jong-Bok

2008-01-01

ER-associated protein degradation (ERAD) is a protein quality control system of ER, which eliminates misfolded proteins by proteasome-dependent degradation and ensures export of only properly folded proteins from ER. Herp, an ER membrane protein upregulated by ER stress, is implicated in regulation of ERAD. In the present study, we show that Herp interacts with members of the ubiquilin family, which function as a shuttle factor to deliver ubiquitinated substrates to the proteasome for degradation. Knockdown of ubiquilin expression by small interfering RNA stabilized the ERAD substrate CD3δ, whereas it did not alter or increased degradation of non-ERAD substrates tested. CD3δ was stabilized by overexpressed Herp mutants which were capable of binding to ubiquilins but were impaired in ER membrane targeting by deletion of the transmembrane domain. Our data suggest that Herp binding to ubiquilin proteins plays an important role in the ERAD pathway and that ubiquilins are specifically involved in degradation of only a subset of ubiquitinated targets, including Herp-dependent ERAD substrates

Stress analysis of HTR-10 steam generator heat exchanging tubes

International Nuclear Information System (INIS)

Dong Jianling; Zhang Xiaohang; Yin Dejian; Fu Jiyang

2001-01-01

Steam Generator (SG) heat exchanging tubes of 10 MW High Temperature Gas Cooled Reactor (HTR-10) are protective screens between the primary loop of helium with radioactivity and the secondary loop of feeding water and steam without radioactivity. Water and steam will enter into the primary loop when rupture of the heat exchanging tubes occurs, which lead to increase of the primary loop pressure and discharge of radioactive materials. Therefore it is important to guarantee the integrity of the tubes. The tube structure is spiral tube with small bending radius, which make it impossible to test with volumetric in-service detection. For such kind of spiral tube, using LBB concept to guarantee the integrity of the tubes is an important option. The author conducts stress analysis and calculation of HTR-10 SG heat exchanging tubes using the FEM code of piping stress analysis, PIPESTRESS. The maximum stress and the dangerous positions are obtained

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

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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.

Synthesis of stress proteins in winter wheat seedlings under gamma-radiation

International Nuclear Information System (INIS)

Gudkova, N.V.; Kosakovskaya, I.V.; Major, P.S.

2001-01-01

A universal cellular response to a number of diverse stresses is the synthesis of a set of stress proteins. Most of them are heat shock proteins (HSP). We show that both heat shock and gamma-radiation enhance the synthesis of HSP70 in the total protein fractions of winter wheat seedlings. It is found that a dose of 15 Gy induced the synthesis of 35 and 45 kD proteins after 5 h of irradiation in both total and mitochondrial protein fractions. On the second day after exposure, both 35 and 45 kD proteins were not observed, but new total proteins with a molecular weight of 90 and 92 kD appeared. The synthesis of 35 and 45 kD proteins after gamma-irradiation is revealed for the first time, their function being now unknown

Whole genome analysis of CRISPR Cas9 sgRNA off-target homologies via an efficient computational algorithm.

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Zhou, Hong; Zhou, Michael; Li, Daisy; Manthey, Joseph; Lioutikova, Ekaterina; Wang, Hong; Zeng, Xiao

2017-11-17

The beauty and power of the genome editing mechanism, CRISPR Cas9 endonuclease system, lies in the fact that it is RNA-programmable such that Cas9 can be guided to any genomic loci complementary to a 20-nt RNA, single guide RNA (sgRNA), to cleave double stranded DNA, allowing the introduction of wanted mutations. Unfortunately, it has been reported repeatedly that the sgRNA can also guide Cas9 to off-target sites where the DNA sequence is homologous to sgRNA. Using human genome and Streptococcus pyogenes Cas9 (SpCas9) as an example, this article mathematically analyzed the probabilities of off-target homologies of sgRNAs and discovered that for large genome size such as human genome, potential off-target homologies are inevitable for sgRNA selection. A highly efficient computationl algorithm was developed for whole genome sgRNA design and off-target homology searches. By means of a dynamically constructed sequence-indexed database and a simplified sequence alignment method, this algorithm achieves very high efficiency while guaranteeing the identification of all existing potential off-target homologies. Via this algorithm, 1,876,775 sgRNAs were designed for the 19,153 human mRNA genes and only two sgRNAs were found to be free of off-target homology. By means of the novel and efficient sgRNA homology search algorithm introduced in this article, genome wide sgRNA design and off-target analysis were conducted and the results confirmed the mathematical analysis that for a sgRNA sequence, it is almost impossible to escape potential off-target homologies. Future innovations on the CRISPR Cas9 gene editing technology need to focus on how to eliminate the Cas9 off-target activity.

Stress granules at the intersection of autophagy and ALS.

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Monahan, Zachary; Shewmaker, Frank; Pandey, Udai Bhan

2016-10-15

Amyotrophic lateral sclerosis (ALS) is a progressive, fatal disease caused by loss of upper and lower motor neurons. The majority of ALS cases are classified as sporadic (80-90%), with the remaining considered familial based on patient history. The last decade has seen a surge in the identification of ALS-causing genes - including TARDBP (TDP-43), FUS, MATR3 (Matrin-3), C9ORF72 and several others - providing important insights into the molecular pathways involved in pathogenesis. Most of the protein products of ALS-linked genes fall into two functional categories: RNA-binding/homeostasis and protein-quality control (i.e. autophagy and proteasome). The RNA-binding proteins tend to be aggregation-prone with low-complexity domains similar to the prion-forming domains of yeast. Many also incorporate into stress granules (SGs), which are cytoplasmic ribonucleoprotein complexes that form in response to cellular stress. Mutant forms of TDP-43 and FUS perturb SG dynamics, lengthening their cytoplasmic persistence. Recent evidence suggests that SGs are regulated by the autophagy pathway, suggesting a unifying connection between many of the ALS-linked genes. Persistent SGs may give rise to intractable aggregates that disrupt neuronal homeostasis, thus failure to clear SGs by autophagic processes may promote ALS pathogenesis. This article is part of a Special Issue entitled SI:Autophagy. Copyright © 2016 Elsevier B.V. All rights reserved.

The BEACH Domain Protein SPIRRIG Is Essential for Arabidopsis Salt Stress Tolerance and Functions as a Regulator of Transcript Stabilization and Localization.

Directory of Open Access Journals (Sweden)

Alexandra Steffens

2015-07-01

Full Text Available Members of the highly conserved class of BEACH domain containing proteins (BDCPs have been established as broad facilitators of protein-protein interactions and membrane dynamics in the context of human diseases like albinism, bleeding diathesis, impaired cellular immunity, cancer predisposition, and neurological dysfunctions. Also, the Arabidopsis thaliana BDCP SPIRRIG (SPI is important for membrane integrity, as spi mutants exhibit split vacuoles. In this work, we report a novel molecular function of the BDCP SPI in ribonucleoprotein particle formation. We show that SPI interacts with the P-body core component DECAPPING PROTEIN 1 (DCP1, associates to mRNA processing bodies (P-bodies, and regulates their assembly upon salt stress. The finding that spi mutants exhibit salt hypersensitivity suggests that the local function of SPI at P-bodies is of biological relevance. Transcriptome-wide analysis revealed qualitative differences in the salt stress-regulated transcriptional response of Col-0 and spi. We show that SPI regulates the salt stress-dependent post-transcriptional stabilization, cytoplasmic agglomeration, and localization to P-bodies of a subset of salt stress-regulated mRNAs. Finally, we show that the PH-BEACH domains of SPI and its human homolog FAN (Factor Associated with Neutral sphingomyelinase activation interact with DCP1 isoforms from plants, mammals, and yeast, suggesting the evolutionary conservation of an association of BDCPs and P-bodies.

Immediate and residual effects of heat stress and restricted intake on milk protein and casein composition and energy metabolism.

Science.gov (United States)

Cowley, F C; Barber, D G; Houlihan, A V; Poppi, D P

2015-04-01

The effects of heat stress on dairy production can be separated into 2 distinct causes: those effects that are mediated by the reduced voluntary feed intake associated with heat stress, and the direct physiological and metabolic effects of heat stress. To distinguish between these, and identify their effect on milk protein and casein concentration, mid-lactation Holstein-Friesian cows (n = 24) were housed in temperature-controlled chambers and either subjected to heat stress [HS; temperature-humidity index (THI) ~78] or kept in a THIheat-stressed cows (TN-R) for 7 d. A control group of cows was kept in a THIheat stress. Heat stress reduced the milk protein concentration, casein number, and casein concentration and increased the urea concentration in milk beyond the effects of restriction of intake. Under HS, the proportion in total casein of αS1-casein increased and the proportion of αS2-casein decreased. Because no effect of HS on milk fat or lactose concentration was found, these effects appeared to be the result of specific downregulation of mammary protein synthesis, and not a general reduction in mammary activity. No residual effects were found of HS or TN-R on milk production or composition after THIHeat-stressed cows had elevated blood concentrations of urea and Ca, compared with TN-R and TN-AL. Cows in TN-R had higher serum nonesterified fatty acid concentrations than cows in HS. It was proposed that HS and TN-R cows may mobilize different tissues as endogenous sources of energy. Copyright © 2015 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Subchronic nandrolone administration reduces cardiac oxidative markers during restraint stress by modulating protein expression patterns.

Science.gov (United States)

Pergolizzi, Barbara; Carriero, Vitina; Abbadessa, Giuliana; Penna, Claudia; Berchialla, Paola; De Francia, Silvia; Bracco, Enrico; Racca, Silvia

2017-10-01

Nandrolone decanoate (ND), an anabolic-androgenic steroid prohibited in collegiate and professional sports, is associated with detrimental cardiovascular effects through redox-dependent mechanisms. We previously observed that high-dose short-term ND administration (15 mg/kg for 2 weeks) did not induce left heart ventricular hypertrophy and, paradoxically, improved postischemic response, whereas chronic ND treatment (5 mg/kg twice a week for 10 weeks) significantly reduced the cardioprotective effect of postconditioning, with an increase in infarct size and a decrease in cardiac performance. We wanted to determine whether short-term ND administration could affect the oxidative redox status in animals exposed to acute restraint stress. Our hypothesis was that, depending on treatment schedule, ND may have a double-edged sword effect. Measurement of malondialdehyde and 4-hydroxynonenal, two oxidative stress markers, in rat plasma and left heart ventricular tissue, revealed that the levels of both markers were increased in animals exposed to restraint stress, whereas no increase in marker levels was noted in animals pretreated with ND, indicating a possible protective action of ND against stress-induced oxidative damage. Furthermore, isolation and identification of proteins extracted from the left heart ventricular tissue samples of rats pretreated or not with ND and exposed to acute stress showed a prevalent expression of enzymes involved in amino acid synthesis and energy metabolism. Among other proteins, peroxiredoxin 6 and alpha B-crystallin, both involved in the oxidative stress response, were predominantly expressed in the left heart ventricular tissues of the ND-pretreated rats. In conclusion, ND seems to reduce oxidative stress by inducing the expression of antioxidant proteins in the hearts of restraint-stressed animals, thus contributing to amelioration of postischemic heart performance.

Quantitative determination of a synthetic amide derivative of gallic acid, SG-HQ2, using liquid chromatography tandem mass spectrometry, and its pharmacokinetics in rats.

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Seo, Seung-Yong; Kang, Wonku

2016-11-30

An amide derivative of gallic acid (GA), 3,4,5-trihydroxy-N-(8-hydroxyquinolin-2-yl)benzamide) (SG-HQ2) was recently synthesized, and its inhibitory actions were previously shown on histamine release and pro-inflammatory cytokine expression. In this study, a simultaneous quantification method was developed for the determination of SG-HQ2 and its possible metabolite, GA, in rat plasma using liquid chromatography with a tandem mass spectrometry (LC-MS/MS). After simple protein precipitation with acetonitrile including diclofenac (internal standard, IS), the analytes were chromatographed on a reversed phased column with a mobile phase of acetonitrile and water (60:40, v/v, including 0.1% formic acid). The ion transitions of the precursor to the product ion were principally protonated ion [M+H] + at m/z 313.2→160.6 for SG-HQ2, and deprotonated ions [M-H] - at m/z 168.7→124.9 for GA and 296.0→251.6 for the IS. The accuracy and precision of the assay were in accordance with FDA regulations for the validation of bioanalytical methods. This method was successfully applied to a pharmacokinetic study of SG-HQ2 after intravenous administration in rats. Copyright © 2016 Elsevier B.V. All rights reserved.

Production of functional proteins: balance of shear stress and gravity

Science.gov (United States)

Goodwin, Thomas John (Inventor); Hammond, Timothy Grant (Inventor); Kaysen, James Howard (Inventor)

2011-01-01

A method for the production of functional proteins including hormones by renal cells in a three dimensional culturing process responsive to shear stress uses a rotating wall vessel. Natural mixture of renal cells expresses the enzyme 1-.alpha.-hydroxylase which can be used to generate the active form of vitamin D: 1,25-diOH vitamin D.sub.3. The fibroblast cultures and co-culture of renal cortical cells express the gene for erythropoietin and secrete erythropoietin into the culture supernatant. Other shear stress response genes are also modulated by shear stress, such as toxin receptors megalin and cubulin (gp280). Also provided is a method of treating an in-need individual with the functional proteins produced in a three dimensional co-culture process responsive to shear stress using a rotating wall vessel.

Comprehensive Protein Interactome Analysis of a Key RNA Helicase: Detection of Novel Stress Granule Proteins

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Rebecca Bish

2015-07-01

Full Text Available DDX6 (p54/RCK is a human RNA helicase with central roles in mRNA decay and translation repression. To help our understanding of how DDX6 performs these multiple functions, we conducted the first unbiased, large-scale study to map the DDX6-centric protein-protein interactome using immunoprecipitation and mass spectrometry. Using DDX6 as bait, we identify a high-confidence and high-quality set of protein interaction partners which are enriched for functions in RNA metabolism and ribosomal proteins. The screen is highly specific, maximizing the number of true positives, as demonstrated by the validation of 81% (47/58 of the RNA-independent interactors through known functions and interactions. Importantly, we minimize the number of indirect interaction partners through use of a nuclease-based digestion to eliminate RNA. We describe eleven new interactors, including proteins involved in splicing which is an as-yet unknown role for DDX6. We validated and characterized in more detail the interaction of DDX6 with Nuclear fragile X mental retardation-interacting protein 2 (NUFIP2 and with two previously uncharacterized proteins, FAM195A and FAM195B (here referred to as granulin-1 and granulin-2, or GRAN1 and GRAN2. We show that NUFIP2, GRAN1, and GRAN2 are not P-body components, but re-localize to stress granules upon exposure to stress, suggesting a function in translation repression in the cellular stress response. Using a complementary analysis that resolved DDX6’s multiple complex memberships, we further validated these interaction partners and the presence of splicing factors. As DDX6 also interacts with the E3 SUMO ligase TIF1β, we tested for and observed a significant enrichment of sumoylation amongst DDX6’s interaction partners. Our results represent the most comprehensive screen for direct interaction partners of a key regulator of RNA life cycle and localization, highlighting new stress granule components and possible DDX6 functions�

WPEC Subgroup Meetings. Joint SG39/SG40-CIELO meeting, 14 May 2014

International Nuclear Information System (INIS)

Chadwick, M.; Yokoyama, Kenji; Ishikawa, M.; Archier, P.; Palmiotti, G.; Salvatores, Massimo; Dupont, E.; ); Leal, L.

2014-05-01

The aim of WPEC subgroup 39 'Methods and approaches to provide feedback from nuclear and covariance data adjustment for improvement of nuclear data files' is to provide criteria and practical approaches to use effectively the results of sensitivity analyses and cross section adjustments for feedback to evaluators and differential measurement experimentalists in order to improve the knowledge of neutron cross sections, uncertainties, and correlations to be used in a wide range of applications. WPEC subgroup 40-CIELO (Collaborative International Evaluated Library Organization) provides a new working paradigm to facilitate evaluated nuclear reaction data advances. It brings together experts from across the international nuclear reaction data community to identify and document discrepancies among existing evaluated data libraries, measured data, and model calculation interpretations, and aims to make progress in reconciling these discrepancies to create more accurate ENDF-formatted files. SG40-CIELO focusses on 6 important isotopes: "1H, "1"6O, "5"6Fe, "2"3"5","2"3"8U, "2"3"9Pu. This document is the proceedings of the Joint SG39/SG40-CIELO meeting, held at the NEA, Issy-les-Moulineaux, France, on 19-20 May 2015. It comprises all the available presentations (slides) given by the participants: 1 - CIELO Progress (M. Chadwick); 2 - Revised Recommendations from ADJ2010 Adjustment (K. Yokoyama); 3 - Comparisons and Discussions on Adjustment trends from JEFF (CEA) (P. Archier); 4 - Feedback on CIELO Isotopes from ENDF/B-VII.0 Adjustment (G. Palmiotti); 5 - Demonstration - Plot comparisons of participants' results (E. Dupont); 6 - Some very preliminary indications from recent adjustment studies intercomparison (M. Salvatores); 7 - Resonance Region of "5"6Fe for the CIELO Project (L. Leal); 8 - A-priori and a-posteriori covariance data in nuclear cross section adjustments: issues and challenges (G. Palmiotti); 9 - Comments on Covariance Data of JENDL-4.0 and ENDF/B-VII.1 (K

Effect of oxidative stress on homer scaffolding proteins.

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Igor Nepliouev

Full Text Available Homer proteins are a family of multifaceted scaffolding proteins that participate in the organization of signaling complexes at the post-synaptic density and in a variety of tissues including striated muscle. Homer isoforms form multimers via their C-terminal coiled coil domains, which allows for the formation of a polymeric network in combination with other scaffolding proteins. We hypothesized that the ability of Homer isoforms to serve as scaffolds would be influenced by oxidative stress. We have found by standard SDS-PAGE of lysates from adult mouse skeletal muscle exposed to air oxidation that Homer migrates as both a dimer and monomer in the absence of reducing agents and solely as a monomer in the presence of a reducing agent, suggesting that Homer dimers exposed to oxidation could be modified by the presence of an inter-molecular disulfide bond. Analysis of the peptide sequence of Homer 1b revealed the presence of only two cysteine residues located adjacent to the C-terminal coiled-coil domain. HEK 293 cells were transfected with wild-type and cysteine mutant forms of Homer 1b and exposed to oxidative stress by addition of menadione, which resulted in the formation of disulfide bonds except in the double mutant (C246G, C365G. Exposure of myofibers from adult mice to oxidative stress resulted in decreased solubility of endogenous Homer isoforms. This change in solubility was dependent on disulfide bond formation. In vitro binding assays revealed that cross-linking of Homer dimers enhanced the ability of Homer 1b to bind Drebrin, a known interacting partner. Our results show that oxidative stress results in disulfide cross-linking of Homer isoforms and loss of solubility of Homer scaffolds. This suggests that disulfide cross-linking of a Homer polymeric network may contribute to the pathophysiology seen in neurodegenerative diseases and myopathies characterized by oxidative stress.

Is occupational stress associated with work engagement ?

Science.gov (United States)

Padula, Rosimeire Simprini; Chiavegato, Luciana Dias; Cabral, Cristina Maria Nunes; Almeid, Talita; Ortiz, Thais; Carregaro, Rodrigo Luiz

2012-01-01

The occupational stress is associated with dissatisfaction, excessive demand at work and personal factors. Those factors can reduce work performance and can predispose workers to various diseases. Workers' health may be protected if there is encouragement to face challenges, which may lessen the impact on psychological and somatic stress and thus have greater personal and professional satisfaction. The aim of this study was to evaluate the association between occupational stress and work engagement. Participated in this study 457 male and female workers of a metallurgical industry. Subjects answered personal data, and the Job Stress Scale and Utrecht Work Engagement Scale were applied. Results showed an association between occupational stress and work engagement (P=0,001). The way the individual deals with his frustrations, or rather the work engagement, is associated with the occupational stress.

The tryptophan-rich sensory protein (TSPO is involved in stress-related and light-dependent processes in the cyanobacterium Fremyella diplosiphon

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Andrea eBusch

2015-12-01

Full Text Available The tryptophan-rich sensory protein (TSPO is a membrane protein, which is a member of the 18 kilodalton translocator protein/peripheral-type benzodiazepine receptor (MBR family of proteins that is present in most organisms and is also referred to as Translocator protein 18 kDa. Although TSPO is associated with stress- and disease-related processes in organisms from bacteria to mammals, full elucidation of the functional role of the TSPO protein is lacking for most organisms in which it is found. In this study, we describe the regulation and function of a TSPO homolog in the cyanobacterium Fremyella diplosiphon, designated FdTSPO. Accumulation of the FdTSPO transcript is upregulated by green light and in response to nutrient deficiency and stress. A F. diplosiphon TSPO deletion mutant (i.e., ΔFdTSPO showed altered responses compared to the wild type strain under stress conditions, including salt treatment, osmotic stress and induced oxidative stress. Under salt stress, the FdTSPO transcript is upregulated and a ΔFdTSPO mutant accumulates lower levels of reactive oxygen species (ROS and displays increased growth compared to WT. In response to osmotic stress, FdTSPO transcript levels are upregulated and ΔFdTSPO mutant cells exhibit impaired growth compared to the wild type. By comparison, methyl viologen-induced oxidative stress results in higher ROS levels in the ΔFdTSPO mutant compared to the wild type strain. Taken together, our results provide support for the involvement of membrane-localized FdTSPO in mediating cellular responses to stress in F. diplosiphon and represent detailed functional analysis of a cyanobacterial TSPO. This study advances our understanding of the functional roles of TSPO homologs in vivo.

P-body proteins regulate transcriptional rewiring to promote DNA replication stress resistance.

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Loll-Krippleber, Raphael; Brown, Grant W

2017-09-15

mRNA-processing (P-) bodies are cytoplasmic granules that form in eukaryotic cells in response to numerous stresses to serve as sites of degradation and storage of mRNAs. Functional P-bodies are critical for the DNA replication stress response in yeast, yet the repertoire of P-body targets and the mechanisms by which P-bodies promote replication stress resistance are unknown. In this study we identify the complete complement of mRNA targets of P-bodies during replication stress induced by hydroxyurea treatment. The key P-body protein Lsm1 controls the abundance of HHT1, ACF4, ARL3, TMA16, RRS1 and YOX1 mRNAs to prevent their toxic accumulation during replication stress. Accumulation of YOX1 mRNA causes aberrant downregulation of a network of genes critical for DNA replication stress resistance and leads to toxic acetaldehyde accumulation. Our data reveal the scope and the targets of regulation by P-body proteins during the DNA replication stress response.P-bodies form in response to stress and act as sites of mRNA storage and degradation. Here the authors identify the mRNA targets of P-bodies during DNA replication stress, and show that P-body proteins act to prevent toxic accumulation of these target transcripts.

Chaperones, but not oxidized proteins, are ubiquitinated after oxidative stress

DEFF Research Database (Denmark)

Kästle, Marc; Reeg, Sandra; Rogowska-Wrzesinska, Adelina

2012-01-01

of these proteins by MALDI tandem mass spectrometry (MALDI MS/MS). As a result we obtained 24 different proteins which can be categorized into the following groups: chaperones, energy metabolism, cytoskeleton/intermediate filaments, and protein translation/ribosome biogenesis. The special set of identified......, ubiquitinated proteins confirm the thesis that ubiquitination upon oxidative stress is no random process to degrade the mass of oxidized proteins, but concerns a special group of functional proteins....

COMPARISON OF THE THERMAL SHOCK RESlST ANCE IN Alı03-SG AND Zr02-l2°/oSi+AI COATING SYSTEMS

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AHMET ÖZEL

1998-12-01

Full Text Available In this investigation� thennal and structure fınite eleınent analysis has been employed to aııalyse tlıe level of the thennal stresses developed in A1203-SG and Zr02- 12%Si+ Al coatings subjected to them1al loading. Systeıns \\Vith 0.4nıın coating thickness and 4nıın substrate ınaterial thickness were nıodelled. Alumina -Ductile Ca st Iran coatings \\Vith Ni Al, Ni Cr AIY, NiCoCrAIY inte rlayer \\vere also modelled. Noıninal and shear stresses at the cıitical interface regions ( filnı /inter laycr/substrate \\vere obtained and compared. The results showed that the Al203-SG coatings has higher tlıennal shock resistance than Zr02- 12o/oSi+Al coating systeıns. Furthern1ore� tlıe inte rlayer thickness and ınate rial coınbinations have a significant influence on the level of the devcloped tl1ennal stresses. It is also concluded that the finite eleınent technique can be used to optiınise the design and the processing of ceranı ic . coatıngs.

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

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Tzu-Yen Yang

2018-05-01

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

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

Science.gov (United States)

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

2018-01-01

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

Long-lasting hippocampal synaptic protein loss in a mouse model of posttraumatic stress disorder.

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Leonie Herrmann

Full Text Available Despite intensive research efforts, the molecular pathogenesis of posttraumatic stress disorder (PTSD and especially of the hippocampal volume loss found in the majority of patients suffering from this anxiety disease still remains elusive. We demonstrated before that trauma-induced hippocampal shrinkage can also be observed in mice exhibiting a PTSD-like syndrome. Aiming to decipher the molecular correlates of these trans-species posttraumatic hippocampal alterations, we compared the expression levels of a set of neurostructural marker proteins between traumatized and control mice at different time points after their subjection to either an electric footshock or mock treatment which was followed by stressful re-exposure in several experimental groups. To our knowledge, this is the first systematic in vivo study analyzing the long-term neuromolecular sequelae of acute traumatic stress combined with re-exposure. We show here that a PTSD-like syndrome in mice is accompanied by a long-lasting reduction of hippocampal synaptic proteins which interestingly correlates with the strength of the generalized and conditioned fear response but not with the intensity of hyperarousal symptoms. Furthermore, we demonstrate that treatment with the serotonin reuptake inhibitor (SSRI fluoxetine is able to counteract both the PTSD-like syndrome and the posttraumatic synaptic protein loss. Taken together, this study demonstrates for the first time that a loss of hippocampal synaptic proteins is associated with a PTSD-like syndrome in mice. Further studies will have to reveal whether these findings are transferable to PTSD patients.

Drosophila DJ-1 decreases neural sensitivity to stress by negatively regulating Daxx-like protein through dFOXO.

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Soojin Hwang

2013-04-01

Full Text Available DJ-1, a Parkinson's disease (PD-associated gene, has been shown to protect against oxidative stress in Drosophila. However, the molecular mechanism underlying oxidative stress-induced phenotypes, including apoptosis, locomotive defects, and lethality, in DJ-1-deficient flies is not fully understood. Here we showed that Daxx-like protein (DLP, a Drosophila homologue of the mammalian Death domain-associated protein (Daxx, was upregulated under oxidative stress conditions in the loss-of-function mutants of Drosophila DJ-1β, a Drosophila homologue of DJ-1. DLP overexpression induced apoptosis via the c-Jun N-terminal kinase (JNK/Drosophila forkhead box subgroup O (dFOXO pathway, whereas loss of DLP increased resistance to oxidative stress and UV irradiation. Moreover, the oxidative stress-induced phenotypes of DJ-1β mutants were dramatically rescued by DLP deficiency, suggesting that enhanced expression of DLP contributes to the DJ-1β mutant phenotypes. Interestingly, we found that dFOXO was required for the increase in DLP expression in DJ-1β mutants and that dFOXO activity was increased in the heads of DJ-1β mutants. In addition, subcellular localization of DLP appeared to be influenced by DJ-1 expression so that cytosolic DLP was increased in DJ-1β mutants. Similarly, in mammalian cells, Daxx translocation from the nucleus to the cytosol was suppressed by overexpressed DJ-1β under oxidative stress conditions; and, furthermore, targeted expression of DJ-1β to mitochondria efficiently inhibited the Daxx translocation. Taken together, our findings demonstrate that DJ-1β protects flies against oxidative stress- and UV-induced apoptosis by regulating the subcellular localization and gene expression of DLP, thus implying that Daxx-induced apoptosis is involved in the pathogenesis of DJ-1-associated PD.

Dynamics of the association of heat shock protein HSPA6 (Hsp70B') and HSPA1A (Hsp70-1) with stress-sensitive cytoplasmic and nuclear structures in differentiated human neuronal cells.

Science.gov (United States)

Shorbagi, Sadek; Brown, Ian R

2016-11-01

Heat shock proteins (Hsps) are cellular repair agents that counter the effects of protein misfolding that is a characteristic feature of neurodegenerative diseases. HSPA1A (Hsp70-1) is a widely studied member of the HSPA (Hsp70) family. The little-studied HSPA6 (Hsp70B') is present in the human genome and absent in mouse and rat; hence, it is missing in current animal models of neurodegenerative diseases. Differentiated human neuronal SH-SY5Y cells were employed to compare the dynamics of the association of YFP-tagged HSPA6 and HSPA1A with stress-sensitive cytoplasmic and nuclear structures. Following thermal stress, live-imaging confocal microscopy and Fluorescence Recovery After Photobleaching (FRAP) demonstrated that HSPA6 displayed a prolonged and more dynamic association, compared to HSPA1A, with centrioles that play critical roles in neuronal polarity and migration. HSPA6 and HSPA1A also targeted nuclear speckles, rich in RNA splicing factors, and the granular component of the nucleolus that is involved in rRNA processing and ribosomal subunit assembly. HSPA6 and HSPA1A displayed similar FRAP kinetics in their interaction with nuclear speckles and the nucleolus. Subsequently, during the recovery from neuronal stress, HSPA6, but not HSPA1A, localized with the periphery of nuclear speckles (perispeckles) that have been characterized as transcription sites. The stress-induced association of HSPA6 with perispeckles displayed the greatest dynamism compared to the interaction of HSPA6 or HSPA1A with other stress-sensitive cytoplasmic and nuclear structures. This suggests involvement of HSPA6 in transcriptional recovery of human neurons from cellular stress that is not apparent for HSPA1A.

Beyond the ENDF format: A modern nuclear database structure. SG38 meeting, OECD Conference Centre, 9-11 May 2016

International Nuclear Information System (INIS)

Herman, Michal Wladyslaw; Cabellos De Francisco, Oscar; Beck, Bret; Trkov, Andrej; Conlin, Jeremy Lloyd; Mcnabb, Dennis P.; Malvagi, Fausto; Grudzevich, Oleg T.; Mattoon, Caleb; Wiarda, Dorothea; Brown, David; Chadwick, Mark; Roubtsov, Danila; Iwamoto, Osamu; Yokoyama, Kenji; White, Morgan C.; Coste-Delclaux, Mireille; Fiorito, Luca; Haeck, Wim; Dunn, Michael; Jouanne, Cedric

2016-05-01

WPEC subgroup 38 (SG38) was formed to develop a new structure for storing nuclear reaction data, that is meant to eventually replace ENDF-6 as the standard way to store and share evaluations. The work of SG38 covers the following tasks: Designing flexible, general-purpose data containers; Determining a logical and easy-to-understand top-level hierarchy for storing evaluated nuclear reaction data; Creating a particle database for storing particles, masses and level schemes; Specifying the infrastructure (plotting, processing, etc.) that must accompany the new structure; Developing an Application Programming Interface or API to allow other codes to access data stored in the new structure; Specifying what tests need to be implemented for quality assurance of the new structure and associated infrastructure; Ensuring documentation and governance of the structure and associated infrastructure. This document is the proceedings of the SG38 meeting which took place at the OECD Conference Centre, on 9-11 May 2016. It comprises all the available presentations (slides) given by the participants: - 1) Beyond the ENDF format: Working toward the first specifications (Dennis P. McNabb); - 2) Summary of LLNL/LANL/ORNL/BNL discussions on SG38 (Caleb M. Mattoon); - 3) Status of Top Level Hierarchy Requirements Document (D.A. Brown); - 4) GND: Storing multiple representations of a quantity using forms, components and styles (Bret Beck); - 5) Particle Database update (Caleb M. Mattoon); - 6) General Purpose Data Containers (Jeremy Lloyd Conlin); - 7) Functional data containers (Bret Beck); - 8) Top-level hierarchy specifications (D.A. Brown)

Small heat shock proteins mediate cell-autonomous and -nonautonomous protection in a Drosophila model for environmental-stress-induced degeneration.

Science.gov (United States)

Kawasaki, Fumiko; Koonce, Noelle L; Guo, Linda; Fatima, Shahroz; Qiu, Catherine; Moon, Mackenzie T; Zheng, Yunzhen; Ordway, Richard W

2016-09-01

Cell and tissue degeneration, and the development of degenerative diseases, are influenced by genetic and environmental factors that affect protein misfolding and proteotoxicity. To better understand the role of the environment in degeneration, we developed a genetic model for heat shock (HS)-stress-induced degeneration in Drosophila This model exhibits a unique combination of features that enhance genetic analysis of degeneration and protection mechanisms involving environmental stress. These include cell-type-specific failure of proteostasis and degeneration in response to global stress, cell-nonautonomous interactions within a simple and accessible network of susceptible cell types, and precise temporal control over the induction of degeneration. In wild-type flies, HS stress causes selective loss of the flight ability and degeneration of three susceptible cell types comprising the flight motor: muscle, motor neurons and associated glia. Other motor behaviors persist and, accordingly, the corresponding cell types controlling leg motor function are resistant to degeneration. Flight motor degeneration was preceded by a failure of muscle proteostasis characterized by diffuse ubiquitinated protein aggregates. Moreover, muscle-specific overexpression of a small heat shock protein (HSP), HSP23, promoted proteostasis and protected muscle from HS stress. Notably, neurons and glia were protected as well, indicating that a small HSP can mediate cell-nonautonomous protection. Cell-autonomous protection of muscle was characterized by a distinct distribution of ubiquitinated proteins, including perinuclear localization and clearance of protein aggregates associated with the perinuclear microtubule network. This network was severely disrupted in wild-type preparations prior to degeneration, suggesting that it serves an important role in muscle proteostasis and protection. Finally, studies of resistant leg muscles revealed that they sustain proteostasis and the microtubule

Dehydrin-like proteins in the necrotrophic fungus Alternaria brassicicola have a role in plant pathogenesis and stress response.

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Stéphanie Pochon

Full Text Available In this study, the roles of fungal dehydrin-like proteins in pathogenicity and protection against environmental stresses were investigated in the necrotrophic seed-borne fungus Alternaria brassicicola. Three proteins (called AbDhn1, AbDhn2 and AbDhn3, harbouring the asparagine-proline-arginine (DPR signature pattern and sharing the characteristic features of fungal dehydrin-like proteins, were identified in the A. brassicicola genome. The expression of these genes was induced in response to various stresses and found to be regulated by the AbHog1 mitogen-activated protein kinase (MAPK pathway. A knock-out approach showed that dehydrin-like proteins have an impact mainly on oxidative stress tolerance and on conidial survival upon exposure to high and freezing temperatures. The subcellular localization revealed that AbDhn1 and AbDhn2 were associated with peroxisomes, which is consistent with a possible perturbation of protective mechanisms to counteract oxidative stress and maintain the redox balance in AbDhn mutants. Finally, we show that the double deletion mutant ΔΔabdhn1-abdhn2 was highly compromised in its pathogenicity. By comparison to the wild-type, this mutant exhibited lower aggressiveness on B. oleracea leaves and a reduced capacity to be transmitted to Arabidopsis seeds via siliques. The double mutant was also affected with respect to conidiation, another crucial step in the epidemiology of the disease.

Acute multi-sgRNA knockdown of KEOPS complex genes reproduces the microcephaly phenotype of the stable knockout zebrafish model.

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Tilman Jobst-Schwan

Full Text Available Until recently, morpholino oligonucleotides have been widely employed in zebrafish as an acute and efficient loss-of-function assay. However, off-target effects and reproducibility issues when compared to stable knockout lines have compromised their further use. Here we employed an acute CRISPR/Cas approach using multiple single guide RNAs targeting simultaneously different positions in two exemplar genes (osgep or tprkb to increase the likelihood of generating mutations on both alleles in the injected F0 generation and to achieve a similar effect as morpholinos but with the reproducibility of stable lines. This multi single guide RNA approach resulted in median likelihoods for at least one mutation on each allele of >99% and sgRNA specific insertion/deletion profiles as revealed by deep-sequencing. Immunoblot showed a significant reduction for Osgep and Tprkb proteins. For both genes, the acute multi-sgRNA knockout recapitulated the microcephaly phenotype and reduction in survival that we observed previously in stable knockout lines, though milder in the acute multi-sgRNA knockout. Finally, we quantify the degree of mutagenesis by deep sequencing, and provide a mathematical model to quantitate the chance for a biallelic loss-of-function mutation. Our findings can be generalized to acute and stable CRISPR/Cas targeting for any zebrafish gene of interest.

Plant Glycine-Rich Proteins in Stress Response: An Emerging, Still Prospective Story

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Magdalena Czolpinska

2018-03-01

Full Text Available Seed plants are sessile organisms that have developed a plethora of strategies for sensing, avoiding, and responding to stress. Several proteins, including the glycine-rich protein (GRP superfamily, are involved in cellular stress responses and signaling. GRPs are characterized by high glycine content and the presence of conserved segments including glycine-containing structural motifs composed of repetitive amino acid residues. The general structure of this superfamily facilitates division of GRPs into five main subclasses. Although the participation of GRPs in plant stress response has been indicated in numerous model and non-model plant species, relatively little is known about the key physiological processes and molecular mechanisms in which those proteins are engaged. Class I, II, and IV members are known to be involved in hormone signaling, stress acclimation, and floral development, and are crucial for regulation of plant cells growth. GRPs of class IV [RNA-binding proteins (RBPs] are involved in alternative splicing or regulation of transcription and stomatal movement, seed, pollen, and stamen development; their accumulation is regulated by the circadian clock. Owing to the fact that the overexpression of GRPs can confer tolerance to stress (e.g., some are involved in cold acclimation and may improve growth at low temperatures, these proteins could play a promising role in agriculture through plant genetic engineering. Consequently, isolation, cloning, characterization, and functional validation of novel GRPs expressed in response to the diverse stress conditions are expected to be growing areas of research in the coming years. According to our knowledge, this is the first comprehensive review on participation of plant GRPs in the response to diverse stress stimuli.

Alpha-mangostin attenuates diabetic nephropathy in association with suppression of acid sphingomyelianse and endoplasmic reticulum stress.

Science.gov (United States)

Liu, Tingting; Duan, Wang; Nizigiyimana, Paul; Gao, Lin; Liao, Zhouning; Xu, Boya; Liu, Lerong; Lei, Minxiang

2018-02-05

Diabetic nephropathy is a common complication of diabetes, but there are currently few treatment options. The aim of this study was to gain insight into the effect of alpha-mangostin on diabetic nephropathy and possible related mechanisms. Goto-Kakizaki rats were used as a diabetic model and received alpha-mangostin or desipramine treatment with normal saline as a control. Ten age-matched Sprague Dawley rats were used as normal controls and treated with normal saline. At week 12, blood glucose, albuminuria, apoptosis and renal pathologic changes were assessed. Protein levels for acid sphingomyelinase, glucose-regulated protein 78, phosphorylated PKR-like ER-resident kinase, activated transcription factor 4, CCAAT/enhancer-binding protein, homologous protein), and cleaved-caspase12 were measured. The level of acid sphingomyelinase was significantly increased, and ER stress was activated in diabetic rat kidneys when compared to the control animals. When acid sphingomyelinase was inhibited by alpha-mangostin, the expression of ER stress-related proteins was down-regulated in association with decreased levels of diabetic kidney injury. Alpha-mangostin, an acid sphingomyelinase inhibitor plays a protective role in diabetic neuropathy by relieving ER stress induced-renal cell apoptosis. Copyright © 2018 Elsevier Inc. All rights reserved.

Cadmium induces the expression of specific stress proteins in sea urchin embryos

International Nuclear Information System (INIS)

Roccheri, Maria Carmela; Agnello, Maria; Bonaventura, Rosa; Matranga, Valeria

2004-01-01

Marine organisms are highly sensitive to many environmental stresses, and consequently, the analysis of their bio-molecular responses to different stress agents is very important for the understanding of putative repair mechanisms. Sea urchin embryos represent a simple though significant model system to test how specific stress can simultaneously affect development and protein expression. Here, we used Paracentrotus lividus sea urchin embryos to study the effects of time-dependent continuous exposure to subacute/sublethal cadmium concentrations. We found that, between 15 and 24 h of exposure, the synthesis of a specific set of stress proteins (90, 72-70, 56, 28, and 25 kDa) was induced, with an increase in the rate of synthesis of 72-70 kDa (hsps), 56 kDa (hsp), and 25 kDa, which was dependent on the lengths of treatment. Recovery experiments in which cadmium was removed showed that while stress proteins continued to be synthesized, embryo development was resumed only after short lengths of exposure

Cross-talk between lipid and protein carbonylation in a dynamic cardiomyocyte model of mild nitroxidative stress

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Eva Griesser

2017-04-01

Full Text Available Reactive oxygen and nitrogen species (ROS/RNS play an important role in the regulation of cardiac function. Increase in ROS/RNS concentration results in lipid and protein oxidation and is often associated with onset and/or progression of many cardiovascular disorders. However, interplay between lipid and protein modifications has not been simultaneously studied in detail so far. Biomolecule carbonylation is one of the most common biomarkers of oxidative stress. Using a dynamic model of nitroxidative stress we demonstrated rapid changes in biomolecule carbonylation in rat cardiomyocytes. Levels of carbonylated species increased as early as 15 min upon treatment with the peroxynitrite donor, 3-morpholinosydnonimine (SIN-1, and decreased to values close to control after 16 h. Total (lipids+proteins vs. protein-specific carbonylation showed different dynamics, with a significant increase in protein-bound carbonyls at later time points. Treatment with SIN-1 in combination with inhibitors of proteasomal and autophagy/lysosomal degradation pathways allowed confirmation of a significant role of the proteasome in the degradation of carbonylated proteins, whereas lipid carbonylation increased in the presence of autophagy/lysosomal inhibitors. Electrophilic aldehydes and ketones formed by lipid peroxidation were identified and relatively quantified using LC-MS/MS. Molecular identity of reactive species was used for data-driven analysis of their protein targets. Combination of different enrichment strategies with LC-MS/MS analysis allowed identification of more than 167 unique proteins with 332 sites modified by electrophilic lipid peroxidation products. Gene ontology analysis of modified proteins demonstrated enrichment of several functional categories including proteins involved in cytoskeleton, extracellular matrix, ion channels and their regulation. Using calcium mobilization assays, the effect of nitroxidative stress on the activity of several ion

Proteotoxic stress induces phosphorylation of p62/SQSTM1 by ULK1 to regulate selective autophagic clearance of protein aggregates.

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Junghyun Lim

Full Text Available Disruption of proteostasis, or protein homeostasis, is often associated with aberrant accumulation of misfolded proteins or protein aggregates. Autophagy offers protection to cells by removing toxic protein aggregates and injured organelles in response to proteotoxic stress. However, the exact mechanism whereby autophagy recognizes and degrades misfolded or aggregated proteins has yet to be elucidated. Mounting evidence demonstrates the selectivity of autophagy, which is mediated through autophagy receptor proteins (e.g. p62/SQSTM1 linking autophagy cargos and autophagosomes. Here we report that proteotoxic stress imposed by the proteasome inhibition or expression of polyglutamine expanded huntingtin (polyQ-Htt induces p62 phosphorylation at its ubiquitin-association (UBA domain that regulates its binding to ubiquitinated proteins. We find that autophagy-related kinase ULK1 phosphorylates p62 at a novel phosphorylation site S409 in UBA domain. Interestingly, phosphorylation of p62 by ULK1 does not occur upon nutrient starvation, in spite of its role in canonical autophagy signaling. ULK1 also phosphorylates S405, while S409 phosphorylation critically regulates S405 phosphorylation. We find that S409 phosphorylation destabilizes the UBA dimer interface, and increases binding affinity of p62 to ubiquitin. Furthermore, lack of S409 phosphorylation causes accumulation of p62, aberrant localization of autophagy proteins and inhibition of the clearance of ubiquitinated proteins or polyQ-Htt. Therefore, our data provide mechanistic insights into the regulation of selective autophagy by ULK1 and p62 upon proteotoxic stress. Our study suggests a potential novel drug target in developing autophagy-based therapeutics for the treatment of proteinopathies including Huntington's disease.

Cdk1-Cyclin B1-mediated Phosphorylation of Tumor-associated Microtubule-associated Protein/Cytoskeleton-associated Protein 2 in Mitosis*

OpenAIRE

Uk Hong, Kyung; Kim, Hyun-Jun; Kim, Hyo-Sil; Seong, Yeon-Sun; Hong, Kyeong-Man; Bae, Chang-Dae; Park, Joobae

2009-01-01

During mitosis, establishment of structurally and functionally sound bipolar spindles is necessary for maintaining the fidelity of chromosome segregation. Tumor-associated microtubule-associated protein (TMAP), also known as cytoskeleton-associated protein 2 (CKAP2), is a mitotic spindle-associated protein whose level is frequently up-regulated in various malignancies. Previous reports have suggested that TMAP is a potential regulator of mitotic spindle assembly and dynamics and that it is re...

Molecular characterization of SG33 and Borghi vaccines used against myxomatosis.

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Cavadini, Patrizia; Botti, Giuliana; Barbieri, Ilaria; Lavazza, Antonio; Capucci, Lorenzo

2010-07-26

Myxoma virus is a poxvirus responsible for myxomatosis in European Rabbits (Oryctolagus cuniculus). The entire genome of the myxoma virus has been sequenced, allowing a systemic survey of the functions of a large number of putative pathogenic factors that this virus expresses to subvert the immune and inflammatory pathways of infected rabbit hosts. In Italy, industrial rabbits are mostly vaccinated against myxomatosis using the attenuated myxoma virus strains Borghi or SG33. We have identified genetic markers specific for Borghi or SG33 vaccine strains and established a PCR-based assay that could be used to: (a) rapidly diagnose the presence of myxoma virus in infected organs; (b) discriminate between field strain-infected and vaccinated rabbits and (c) differentiate between Borghi or SG33 vaccine strain. Copyright 2010 Elsevier Ltd. All rights reserved.

Nuclear transport of heat shock proteins in stressed cells

International Nuclear Information System (INIS)

Chughtai, Zahoor Saeed

2001-01-01

Nuclear import of proteins that are too large to passively enter the nucleus requires soluble factors, energy , and a nuclear localization signal (NLS). Nuclear protein transport can be regulated, and different forms of stress affect nucleocytoplasmic trafficking. As such, import of proteins containing a classical NLS is inhibited in starving yeast cells. In contrast, the heat shock protein hsp70 Ssa4p concentrates in nuclei upon starvation. Nuclear concentration of Ssa4p in starving cells is reversible, and transfer of nutrient-depleted cells to fresh medium induces Ssa4p nuclear export. This export reaction represents an active process that is sensitive to oxidative stress. Upon starvation, the N-terminal domain of Ssa4p mediates Ssa4p nuclear accumulation, and a short hydrophobic sequence, termed Star (for starvation), is sufficient to localize the reporter proteins green fluorescent protein or β-gaIactosidase to nuclei. To determine whether nuclear accumulation of Star-β-galactosidase depends on a specific nuclear carrier, I have analyzed its distribution in mutant yeast strains that carry a deletion of a single β-importin gene. With this assay I have identified Nmd5p as a β-importin required to concentrate Star-β-galactosidase in nuclei of stationary phase cells. (author)

Nuclear transport of heat shock proteins in stressed cells

Energy Technology Data Exchange (ETDEWEB)

Chughtai, Zahoor Saeed

2001-07-01

Nuclear import of proteins that are too large to passively enter the nucleus requires soluble factors, energy , and a nuclear localization signal (NLS). Nuclear protein transport can be regulated, and different forms of stress affect nucleocytoplasmic trafficking. As such, import of proteins containing a classical NLS is inhibited in starving yeast cells. In contrast, the heat shock protein hsp70 Ssa4p concentrates in nuclei upon starvation. Nuclear concentration of Ssa4p in starving cells is reversible, and transfer of nutrient-depleted cells to fresh medium induces Ssa4p nuclear export. This export reaction represents an active process that is sensitive to oxidative stress. Upon starvation, the N-terminal domain of Ssa4p mediates Ssa4p nuclear accumulation, and a short hydrophobic sequence, termed Star (for starvation), is sufficient to localize the reporter proteins green fluorescent protein or {beta}-gaIactosidase to nuclei. To determine whether nuclear accumulation of Star-{beta}-galactosidase depends on a specific nuclear carrier, I have analyzed its distribution in mutant yeast strains that carry a deletion of a single {beta}-importin gene. With this assay I have identified Nmd5p as a {beta}-importin required to concentrate Star-{beta}-galactosidase in nuclei of stationary phase cells. (author)

Oxidative stress impairs the heat stress response and delays unfolded protein recovery.

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Masaaki Adachi

2009-11-01

Full Text Available Environmental changes, air pollution and ozone depletion are increasing oxidative stress, and global warming threatens health by heat stress. We now face a high risk of simultaneous exposure to heat and oxidative stress. However, there have been few studies investigating their combined adverse effects on cell viability.Pretreatment of hydrogen peroxide (H(2O(2 specifically and highly sensitized cells to heat stress, and enhanced loss of mitochondrial membrane potential. H(2O(2 exposure impaired the HSP40/HSP70 induction as heat shock response (HSR and the unfolded protein recovery, and enhanced eIF2alpha phosphorylation and/or XBP1 splicing, land marks of ER stress. These H(2O(2-mediated effects mimicked enhanced heat sensitivity in HSF1 knockdown or knockout cells. Importantly, thermal preconditioning blocked H(2O(2-mediated inhibitory effects on refolding activity and rescued HSF1 +/+ MEFs, but neither blocked the effects nor rescued HSF1 -/- MEFs. These data strongly suggest that inhibition of HSR and refolding activity is crucial for H(2O(2-mediated enhanced heat sensitivity.H(2O(2 blocks HSR and refolding activity under heat stress, thereby leading to insufficient quality control and enhancing ER stress. These uncontrolled stress responses may enhance cell death. Our data thus highlight oxidative stress as a crucial factor affecting heat tolerance.

Analysis of syringyl and guaiacyl (S/G) ratio in lignin

CSIR Research Space (South Africa)

Spark, A

2006-12-01

Full Text Available the acidolysis products – Validate the new S/G ratio method Why are S/G ratios important? Gives a good indication of the reactivity of the lignin Experimental design Literature Review Establishing Acidolysis conditions Permanganate oxidation Lignin...-method is quick • Permanganate oxidation-method is slow but it is the standard method used at present • Nitrobenzene Oxidation, Pyrolysis, Cupric Oxidation and Thioacidolysis Lignin can be broken down to syringyl and guaiacyl subunits: OCH3 OH OCH3 OH...

Silicon Mitigates Salinity Stress by Regulating the Physiology, Antioxidant Enzyme Activities, and Protein Expression in Capsicum annuum 'Bugwang'.

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Manivannan, Abinaya; Soundararajan, Prabhakaran; Muneer, Sowbiya; Ko, Chung Ho; Jeong, Byoung Ryong

2016-01-01

Silicon- (Si-) induced salinity stress resistance was demonstrated at physiological and proteomic levels in Capsicum annuum for the first time. Seedlings of C. annuum were hydroponically treated with NaCl (50 mM) with or without Si (1.8 mM) for 15 days. The results illustrated that saline conditions significantly reduced plant growth and biomass and photosynthetic parameters and increased the electrolyte leakage potential, lipid peroxidation, and hydrogen peroxide level. However, supplementation of Si allowed the plants to recover from salinity stress by improving their physiology and photosynthesis. During salinity stress, Si prevented oxidative damage by increasing the activities of antioxidant enzymes. Furthermore, Si supplementation recovered the nutrient imbalance that had occurred during salinity stress. Additionally, proteomic analysis by two-dimensional gel electrophoresis (2DE) followed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) revealed that Si treatment upregulated the accumulation of proteins involved in several metabolic processes, particularly those associated with nucleotide binding and transferase activity. Moreover, Si modulated the expression of vital proteins involved in ubiquitin-mediated nucleosome pathway and carbohydrate metabolism. Overall, the results illustrate that Si application induced resistance against salinity stress in C. annuum by regulating the physiology, antioxidant metabolism, and protein expression.

Methods for monitoring endoplasmic reticulum stress and the unfolded protein response.

LENUS (Irish Health Repository)

Samali, Afshin

2010-01-01

The endoplasmic reticulum (ER) is the site of folding of membrane and secreted proteins in the cell. Physiological or pathological processes that disturb protein folding in the endoplasmic reticulum cause ER stress and activate a set of signaling pathways termed the Unfolded Protein Response (UPR). The UPR can promote cellular repair and sustained survival by reducing the load of unfolded proteins through upregulation of chaperones and global attenuation of protein synthesis. Research into ER stress and the UPR continues to grow at a rapid rate as many new investigators are entering the field. There are also many researchers not working directly on ER stress, but who wish to determine whether this response is activated in the system they are studying: thus, it is important to list a standard set of criteria for monitoring UPR in different model systems. Here, we discuss approaches that can be used by researchers to plan and interpret experiments aimed at evaluating whether the UPR and related processes are activated. We would like to emphasize that no individual assay is guaranteed to be the most appropriate one in every situation and strongly recommend the use of multiple assays to verify UPR activation.

Methods for Monitoring Endoplasmic Reticulum Stress and the Unfolded Protein Response

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Afshin Samali

2010-01-01

Full Text Available The endoplasmic reticulum (ER is the site of folding of membrane and secreted proteins in the cell. Physiological or pathological processes that disturb protein folding in the endoplasmic reticulum cause ER stress and activate a set of signaling pathways termed the Unfolded Protein Response (UPR. The UPR can promote cellular repair and sustained survival by reducing the load of unfolded proteins through upregulation of chaperones and global attenuation of protein synthesis. Research into ER stress and the UPR continues to grow at a rapid rate as many new investigators are entering the field. There are also many researchers not working directly on ER stress, but who wish to determine whether this response is activated in the system they are studying: thus, it is important to list a standard set of criteria for monitoring UPR in different model systems. Here, we discuss approaches that can be used by researchers to plan and interpret experiments aimed at evaluating whether the UPR and related processes are activated. We would like to emphasize that no individual assay is guaranteed to be the most appropriate one in every situation and strongly recommend the use of multiple assays to verify UPR activation.

Identification of seed proteins associated with resistance to pre-harvested aflatoxin contamination in peanut (Arachis hypogaea L

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Li Ling

2010-11-01

Full Text Available Abstract Background Pre-harvest infection of peanuts by Aspergillus flavus and subsequent aflatoxin contamination is one of the food safety factors that most severely impair peanut productivity and human and animal health, especially in arid and semi-arid tropical areas. Some peanut cultivars with natural pre-harvest resistance to aflatoxin contamination have been identified through field screening. However, little is known about the resistance mechanism, which has slowed the incorporation of resistance into cultivars with commercially acceptable genetic background. Therefore, it is necessary to identify resistance-associated proteins, and then to recognize candidate resistance genes potentially underlying the resistance mechanism. Results The objective of this study was to identify resistance-associated proteins in response to A. flavus infection under drought stress using two-dimensional electrophoresis with mass spectrometry. To identify proteins involved in the resistance to pre-harvest aflatoxin contamination, we compared the differential expression profiles of seed proteins between a resistant cultivar (YJ-1 and a susceptible cultivar (Yueyou 7 under well-watered condition, drought stress, and A. flavus infection with drought stress. A total of 29 spots showed differential expression between resistant and susceptible cultivars in response to A. flavus attack under drought stress. Among these spots, 12 protein spots that consistently exhibited an altered expression were screened by Image Master 5.0 software and successfully identified by MALDI-TOF MS. Five protein spots, including Oso7g0179400, PII protein, CDK1, Oxalate oxidase, SAP domain-containing protein, were uniquely expressed in the resistant cultivar. Six protein spots including low molecular weight heat shock protein precursor, RIO kinase, L-ascorbate peroxidase, iso-Ara h3, 50 S ribosomal protein L22 and putative 30 S ribosomal S9 were significantly up-regulated in the resistant

A Comprehensive Analysis of Chromoplast Differentiation Reveals Complex Protein Changes Associated with Plastoglobule Biogenesis and Remodeling of Protein Systems in Sweet Orange Flesh1[OPEN

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Wang, Lun; Deng, Xiuxin

2015-01-01

Globular and crystalloid chromoplasts were observed to be region specifically formed in sweet orange (Citrus sinensis) flesh and converted from amyloplasts during fruit maturation, which was associated with the composition of specific carotenoids and the expression of carotenogenic genes. Subsequent isobaric tag for relative and absolute quantitation (iTRAQ)-based quantitative proteomic analyses of purified plastids from the flesh during chromoplast differentiation and senescence identified 1,386 putative plastid-localized proteins, 1,016 of which were quantified by spectral counting. The iTRAQ values reflecting the expression abundance of three identified proteins were validated by immunoblotting. Based on iTRAQ data, chromoplastogenesis appeared to be associated with three major protein expression patterns: (1) marked decrease in abundance of the proteins participating in the translation machinery through ribosome assembly; (2) increase in abundance of the proteins involved in terpenoid biosynthesis (including carotenoids), stress responses (redox, ascorbate, and glutathione), and development; and (3) maintenance of the proteins for signaling and DNA and RNA. Interestingly, a strong increase in abundance of several plastoglobule-localized proteins coincided with the formation of plastoglobules in the chromoplast. The proteomic data also showed that stable functioning of protein import, suppression of ribosome assembly, and accumulation of chromoplast proteases are correlated with the amyloplast-to-chromoplast transition; thus, these processes may play a collective role in chromoplast biogenesis and differentiation. By contrast, the chromoplast senescence process was inferred to be associated with significant increases in stress response and energy supply. In conclusion, this comprehensive proteomic study identified many potentially new plastid-localized proteins and provides insights into the potential developmental and molecular mechanisms underlying chromoplast

Longitudinal Associations Among Pain, Posttraumatic Stress Disorder Symptoms, and Stress Appraisals.

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Vaughan, Christine A; Miles, Jeremy N V; Eisenman, David P; Meredith, Lisa S

2016-04-01

Comorbidity of posttraumatic stress disorder (PTSD) and pain is well documented, but the mechanisms underlying their comorbidity are not well understood. Cross-lagged regression models were estimated with 3 waves of longitudinal data to examine the reciprocal associations between PTSD symptom severity, as measured by the Clinician-Administered PTSD Scale (CAPS), and pain, as measured by a brief self-report measure of pain called the PEG (pain intensity [P], interference with enjoyment of life [E], and interference with general activity [G]). We evaluated stress appraisals as a mediator of these associations in a sample of low-income, underserved patients with PTSD (N = 355) at federally qualified health centers in a northeastern metropolitan area. Increases in PTSD symptom severity between baseline and 6-month and 6- and 12-month assessments were independently predicted by higher levels of pain (β = .14 for both lags) and appraisals of life stress as uncontrollable (β = .15 for both lags). Stress appraisals, however, did not mediate these associations, and PTSD symptom severity did not predict change in pain. Thus, the results did not support the role of stress appraisals as a mechanism underlying the associations between pain and PTSD. Copyright © 2016 International Society for Traumatic Stress Studies.

Isogenic FUS-eGFP iPSC Reporter Lines Enable Quantification of FUS Stress Granule Pathology that Is Rescued by Drugs Inducing Autophagy

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Lara Marrone

2018-02-01

Full Text Available Summary: Perturbations in stress granule (SG dynamics may be at the core of amyotrophic lateral sclerosis (ALS. Since SGs are membraneless compartments, modeling their dynamics in human motor neurons has been challenging, thus hindering the identification of effective therapeutics. Here, we report the generation of isogenic induced pluripotent stem cells carrying wild-type and P525L FUS-eGFP. We demonstrate that FUS-eGFP is recruited into SGs and that P525L profoundly alters their dynamics. With a screening campaign, we demonstrate that PI3K/AKT/mTOR pathway inhibition increases autophagy and ameliorates SG phenotypes linked to P525L FUS by reducing FUS-eGFP recruitment into SGs. Using a Drosophila model of FUS-ALS, we corroborate that induction of autophagy significantly increases survival. Finally, by screening clinically approved drugs for their ability to ameliorate FUS SG phenotypes, we identify a number of brain-penetrant anti-depressants and anti-psychotics that also induce autophagy. These drugs could be repurposed as potential ALS treatments. : Sterneckert and colleagues generate isogenic FUS-eGFP reporter iPSCs that enable the identification of stress granule (SG phenotypes specifically induced by the ALS mutation FUS P525L. Compound screening shows that modulation of the PI3K/AKT/mTOR pathway regulating autophagy ameliorates SG phenotypes. A second screen identifies similarly acting brain-penetrant US FDA-approved drugs that could be repurposed to treat ALS. Keywords: amyotrophic lateral sclerosis, induced pluripotent stem cells, FUS, stress granules, autophagy, gene editing, CRISPR/Cas9n

Anxiety and styles of coping with occupational stress resulting from work with 'dangerous' prisoners in prison service officers.

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Tsirigotis, Konstantinos; Gruszczyński, Wojciech; Pęczkowski, Sebastian

2015-10-01

Prisoners categorised as 'dangerous' are a category of prisoners that require and/or force into using special measures of caution, protection and security. The aim of the study was to examine the intensity of anxiety (as a state and as a trait) experienced by officers working with 'dangerous' prisoners and styles of coping with stress they adopt. A total of 40 officers working with 'dangerous' prisoners (the study group, SG) and 60 officers of the security department not working with 'dangerous' prisoners (the reference group, RG) were studied. The intensity of anxiety was assessed applying the Polish version of 'State-Trait Anxiety Inventory' (STAI); styles of coping with stress were explored employing the Polish version of 'Coping Inventory for Stressful Situations' (CISS) and the author's own questionnaire. Data were analysed using the mean, standard deviation, difference testing (the Mann-Whitney U test), correlation-regression procedure (Kendall's tau, τ correlation coefficient and forward stepwise multiple regression). Officers in the SG faced verbal and physical aggression; nevertheless, scores of officers in both the groups were within the interval of mean scores for all the studied STAI and CISS variables. Officers in the SG achieved significantly higher scores on the state-anxiety scale and the Emotion-Oriented Style (EOS), and lower scores on the Task-Oriented Style (TOS) and Social Diversion (SD). The correlation-regression procedure indicated that there were relationships between anxiety and styles of coping with stress but they differed slightly between the groups. Officers in the SG feel state anxiety stronger and display a stronger preference for the EOS than officers in the RG. Officers in the RG more strongly prefer the TOS and SD. State anxiety is a variable negatively explaining the TOS in the SG, whereas anxiety as a trait is a variable explaining the EOS in both the groups. The coping styles of warders dealing with dangerous prisoners are

Dietary live yeast alters metabolic profiles, protein biosynthesis and thermal stress tolerance of Drosophila melanogaster.

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Colinet, Hervé; Renault, David

2014-04-01

The impact of nutritional factors on insect's life-history traits such as reproduction and lifespan has been excessively examined; however, nutritional determinant of insect's thermal tolerance has not received a lot of attention. Dietary live yeast represents a prominent source of proteins and amino acids for laboratory-reared drosophilids. In this study, Drosophila melanogaster adults were fed on diets supplemented or not with live yeast. We hypothesized that manipulating nutritional conditions through live yeast supplementation would translate into altered physiology and stress tolerance. We verified how live yeast supplementation affected body mass characteristics, total lipids and proteins, metabolic profiles and cold tolerance (acute and chronic stress). Females fed with live yeast had increased body mass and contained more lipids and proteins. Using GC/MS profiling, we found distinct metabolic fingerprints according to nutritional conditions. Metabolite pathway enrichment analysis corroborated that live yeast supplementation was associated with amino acid and protein biosyntheses. The cold assays revealed that the presence of dietary live yeast greatly promoted cold tolerance. Hence, this study conclusively demonstrates a significant interaction between nutritional conditions and thermal tolerance. Copyright © 2014 Elsevier Inc. All rights reserved.

Protein oxidative stress markers in peritoneal fluids of women with deep infiltrating endometriosis are increased.

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Santulli, Pietro; Chouzenoux, Sandrine; Fiorese, Mauro; Marcellin, Louis; Lemarechal, Herve; Millischer, Anne-Elodie; Batteux, Frédéric; Borderie, Didier; Chapron, Charles

2015-01-01

when compared with controls (P endometriosis (SUP and OMA) showed non-statistically significant increases. We found positive correlations between peritoneal fluids AOPP concentrations, nitrites/nitrates levels and the total number of intestinal DIE lesions (r = 0.464; P peritoneal proteins oxidative stress markers. We demonstrate the existence of a significantly increased protein oxidative stress status in peritoneal fluid from women with endometriosis especially in cases of DIE with intestinal involvement. This study opens the way to future more mechanistics studies to determine the exact role of protein oxidative stress in the pathogenesis of endometriosis. Even if an association does not establish proof of cause and effect, these intrinsic biochemical characteristics of endometriosis may lead to the evaluation of therapeutic approaches targeting oxidative imbalance. No funding was used for this study. The authors have no conflict of interest. © The Author 2014. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Exoproteome analysis reveals higher abundance of proteins linked to alkaline stress in persistent Listeria monocytogenes strains.

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Rychli, Kathrin; Grunert, Tom; Ciolacu, Luminita; Zaiser, Andreas; Razzazi-Fazeli, Ebrahim; Schmitz-Esser, Stephan; Ehling-Schulz, Monika; Wagner, Martin

2016-02-02

surface virulence associated protein SvpA. Furthermore proteins involved in cell wall modification, such as the lipoteichonic acid primase LtaP and the N-acetylmuramoyl-l-alanine amidase (Lmo2591) are more abundant in EGDe than in the persistent strains and could indirectly contribute to virulence. In conclusion this study provides information about a set of proteins that could potentially support survival of L. monocytogenes in abiotic niches in food processing environments. Based on these data, a more detailed analysis of the role of the identified proteins under stresses mimicking conditions in food producing environment is essential for further elucidate the mechanism of the phenomenon of persistence of L. monocytogenes. Copyright © 2015 Elsevier B.V. All rights reserved.

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

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Tomokazu Matsuda

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

High intake of heterocyclic amines from meat is associated with oxidative stress.

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Carvalho, A M; Miranda, A M; Santos, F A; Loureiro, A P M; Fisberg, R M; Marchioni, D M

2015-04-28

High meat intake has been related to chronic diseases such as cancer and CVD. One hypothesis is that heterocyclic amines (HCA), which are formed during the cooking process of meat, can generate reactive species. These compounds can cause oxidation of lipids, proteins and DNA, resulting in oxidative stress, cell damage and loss of biological function. This association has been seen in vitro; however, it remains unclear in vivo. The aim of the present study was to investigate the association between oxidative stress and HCA intake, and oxidative stress and meat intake. Data were from the Health Survey for Sao Paulo--ISA-Capital (561 adult and elderly). Food intake was estimated by one 24-h dietary recall (24HR) complemented by a detailed FFQ with preferences of cooking methods and level of doneness for meat. HCA intake was estimated linking the meat from the 24HR to a database of HCA. Oxidative stress was estimated by malondialdehyde (MDA) concentration in the plasma, after derivatisation with thiobarbituric acid and quantification by HPLC/diode array. Analyses were performed using multivariate logistic regressions adjusted for smoking, sex, age, BMI, skin colour, energy intake, fruit and vegetable intake, and physical activity. A positive association between HCA intake and MDA concentration (OR 1·17; 95% CI 1·01, 1·38) was observed, showing that HCA from meat may contribute to increase oxidative stress, and may consequently increase the risk of chronic diseases.

The stress protein heat shock cognate 70 (Hsc70) inhibits the Transient Receptor Potential Vanilloid type 1 (TRPV1) channel.

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Iftinca, Mircea; Flynn, Robyn; Basso, Lilian; Melo, Helvira; Aboushousha, Reem; Taylor, Lauren; Altier, Christophe

2016-01-01

Specialized cellular defense mechanisms prevent damage from chemical, biological, and physical hazards. The heat shock proteins have been recognized as key chaperones that maintain cell survival against a variety of exogenous and endogenous stress signals including noxious temperature. However, the role of heat shock proteins in nociception remains poorly understood. We carried out an expression analysis of the constitutively expressed 70 kDa heat-shock cognate protein, a member of the stress-induced HSP70 family in lumbar dorsal root ganglia from a mouse model of Complete Freund's Adjuvant-induced chronic inflammatory pain. We used immunolabeling of dorsal root ganglion neurons, behavioral analysis and patch clamp electrophysiology in both dorsal root ganglion neurons and HEK cells transfected with Hsc70 and Transient Receptor Potential Channels to examine their functional interaction in heat shock stress condition. We report an increase in protein levels of Hsc70 in mouse dorsal root ganglia, 3 days post Complete Freund's Adjuvant injection in the hind paw. Immunostaining of Hsc70 was observed in most of the dorsal root ganglion neurons, including the small size nociceptors immunoreactive to the TRPV1 channel. Standard whole-cell patch-clamp technique was used to record Transient Receptor Potential Vanilloid type 1 current after exposure to heat shock. We found that capsaicin-evoked currents are inhibited by heat shock in dorsal root ganglion neurons and transfected HEK cells expressing Hsc70 and TRPV1. Blocking Hsc70 with matrine or spergualin compounds prevented heat shock-induced inhibition of the channel. We also found that, in contrast to TRPV1, both the cold sensor channels TRPA1 and TRPM8 were unresponsive to heat shock stress. Finally, we show that inhibition of TRPV1 depends on the ATPase activity of Hsc70 and involves the rho-associated protein kinase. Our work identified Hsc70 and its ATPase activity as a central cofactor of TRPV1 channel function

Effects of heat shock protein 90 expression on pectoralis major oxidation in broilers exposed to acute heat stress.

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Hao, Y; Gu, X H

2014-11-01

This study was conducted to determine the effects of heat shock protein 90 (HSP90) expression on pH, lipid peroxidation, heat shock protein 70 (HSP70), and glucocorticoid receptor (GR) expression of pectoralis major in broilers exposed to acute heat stress. In total, 90 male broilers were randomly allocated to 3 groups: control (CON), heat stress (HS), or geldanamycin treatment (GA). On d 41, the broilers in the GA group were injected intraperitoneally with GA (5 μg/kg of BW), and the broilers in the CON and HS groups were injected intraperitoneally with saline. Twenty-four hours later, the broilers in the CON group were moved to environmental chambers controlled at 22°C for 2 h, and the broilers in the HS and GA groups were moved to environmental chambers controlled at 40°C for 2 h. The pH values of the pectoralis major after 30 min and 24 h of chilling after slaughter of HS and GA broilers were significantly lower (P stress caused significant increases in sera corticosterone and lactic dehydrogenase, the activity of malondialdehyde and superoxide dismutase, the expression of HSP90 and HSP70, and nuclear expression of GR protein in the pectoralis major (P stress induced a significant decrease in GR protein expression in the cytoplasm and GR mRNA expression. Furthermore, the low expression of HSP90 significantly increased levels of lactic dehydrogenase and malondialdehyde and GR protein expression in the cytoplasm under heat stress (P shock protein 90 was positively correlated with corticosterone and superoxide dismutase activities (P < 0.01), and HSP90 mRNA was negatively correlated with pH after chilling for 24 h. The results demonstrated that HSP90 plays a pivotal role in protecting cells from oxidation. ©2014 Poultry Science Association Inc.

A liver stress-endocrine nexus promotes metabolic integrity during dietary protein dilution

DEFF Research Database (Denmark)

Maida, Adriano; Zota, Annika; Sjøberg, Kim Anker

2016-01-01

of impaired glucose homeostasis independently of obesity and food intake. DPD-mediated metabolic inefficiency and improvement of glucose homeostasis were independent of uncoupling protein 1 (UCP1), but required expression of liver-derived fibroblast growth factor 21 (FGF21) in both lean and obese mice. FGF21...... expression and secretion as well as the associated metabolic remodeling induced by DPD also required induction of liver-integrated stress response-driven nuclear protein 1 (NUPR1). Insufficiency of select nonessential amino acids (NEAAs) was necessary and adequate for NUPR1 and subsequent FGF21 induction...... and secretion in hepatocytes in vitro and in vivo. Taken together, these data indicate that DPD promotes improved glucose homeostasis through an NEAA insufficiency-induced liver NUPR1/FGF21 axis....

Folding propensity of intrinsically disordered proteins by osmotic stress

International Nuclear Information System (INIS)

Mansouri, Amanda L.; Grese, Laura N.; Rowe, Erica L.

2016-01-01

Proteins imparted with intrinsic disorder conduct a range of essential cellular functions. To better understand the folding and hydration properties of intrinsically disordered proteins (IDPs), we used osmotic stress to induce conformational changes in nuclear co-activator binding domain (NCBD) and activator for thyroid hormone and retinoid receptor (ACTR). Osmotic stress was applied by the addition of small and polymeric osmolytes, where we discovered that water contributions to NCBD folding always exceeded those for ACTR. Both NCBD and ACTR were found to gain a-helical structure with increasing osmotic stress, consistent with their folding upon NCBD/ACTR complex formation. Using small-angle neutron scattering (SANS), we further characterized NCBD structural changes with the osmolyte ethylene glycol. Here a large reduction in overall size initially occurred before substantial secondary structural change. In conclusion, by focusing on folding propensity, and linked hydration changes, we uncover new insights that may be important for how IDP folding contributes to binding.

Aggregation of SND1 in Stress Granules is Associated with the Microtubule Cytoskeleton During Heat Shock Stimulus.

Science.gov (United States)

Shao, Jie; Gao, Fei; Zhang, Bingbing; Zhao, Meng; Zhou, Yunli; He, Jinyan; Ren, Li; Yao, Zhi; Yang, Jie; Su, Chao; Gao, Xingjie

2017-12-01

Stress granules (SGs) are dynamic dense structures in the cytoplasm that form in response to a variety of environmental stress stimuli. Staphylococcal nuclease and Tudor domain containing 1 (SND1) is a type of RNA-binding protein and has been identified as a transcriptional co-activator. Our previous studies have shown that SND1 is a component of the stress granule, which forms under stress conditions. Here, we observed that SND1 granules were often surrounded by ɑ-tubulin-microtubules in 45°C-treated HeLa cells at 15 min or colocalized with microtubules at 30 or 45 min. Furthermore, Nocodazole-mediated microtubule depolymerization could significantly affect the efficient recruitment of SND1 proteins to the SGs during heat shock stress. In addition, the 45°C heat shock mediated the enhancement of eIF2α phosphorylation, which was not affected by treatment with Nocodazole, an agent that disrupts the cytoskeleton. The intact microtubule cytoskeletal tracks are important for the efficient assembly of SND1 granules under heat shock stress and may facilitate SND1 shuttling between cytoplasmic RNA foci. Anat Rec, 300:2192-2199, 2017. © 2017 The Authors The Anatomical Record published by Wiley Periodicals, Inc. on behalf of American Association of Anatomists. Copyright © 2017 The Authors The Anatomical Record published by Wiley Periodicals, Inc. on behalf of American Association of Anatomists.

Lower fetuin-A, retinol binding protein 4 and several metabolites after gastric bypass compared to sleeve gastrectomy in patients with type 2 diabetes.

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Mia Jüllig

Full Text Available BACKGROUND: Bypass of foregut secreted factors promoting insulin resistance is hypothesized to be one of the mechanisms by which resolution of type 2 diabetes (T2D follows roux-en-y gastric bypass (GBP surgery. AIM: To identify insulin resistance-associated proteins and metabolites which decrease more after GBP than after sleeve gastrectomy (SG prior to diabetes remission. METHODS: Fasting plasma from 15 subjects with T2D undergoing GBP or SG was analyzed by proteomic and metabolomic methods 3 days before and 3 days after surgery. Subjects were matched for age, BMI, metformin therapy and glycemic control. Insulin resistance was calculated using homeostasis model assessment (HOMA-IR. For proteomics, samples were depleted of abundant plasma proteins, digested with trypsin and labeled with iTRAQ isobaric tags prior to liquid chromatography-tandem mass spectrometry analysis. Metabolomic analysis was performed using gas chromatography-mass spectrometry. The effect of the respective bariatric surgery on identified proteins and metabolites was evaluated using two-way analysis of variance and appropriate post-hoc tests. RESULTS: HOMA-IR improved, albeit not significantly, in both groups after surgery. Proteomic analysis yielded seven proteins which decreased significantly after GBP only, including Fetuin-A and Retinol binding protein 4, both previously linked to insulin resistance. Significant decrease in Fetuin-A and Retinol binding protein 4 after GBP was confirmed using ELISA and immunoassay. Metabolomic analysis identified significant decrease of citrate, proline, histidine and decanoic acid specifically after GBP. CONCLUSION: Greater early decrease was seen for Fetuin-A, Retinol binding protein 4, and several metabolites after GBP compared to SG, preceding significant weight loss. This may contribute to enhanced T2D remission observed following foregut bypass procedures.

Oxidative stress and lowered total antioxidant status are associated with a history of suicide attempts.

Science.gov (United States)

Vargas, Heber Odebrecht; Nunes, Sandra Odebrecht Vargas; Pizzo de Castro, Marcia; Bortolasci, Chiara Cristina; Sabbatini Barbosa, Décio; Kaminami Morimoto, Helena; Venugopal, Kamalesh; Dodd, Seetal; Maes, Michael; Berk, Michael

2013-09-25

There is evidence that depression is accompanied by inflammation, oxidative and nitrosative stress (O&NS) and metabolic disorders. However links between oxidative stress and suicide attempts in depressed patients are poorly understood. This study examines whether a history of suicide attempts is associated with inflammation, O&NS and metabolic disorders. Blood specimens were collected from study participants aged 18-60 (N=342) recruited at the State University of Londrina, Brazil, and measured for oxidative stress biomarkers: nitric oxide metabolites (NOx), lipid hydroperoxides, malondialdehyde, advanced oxidation protein products and plasma total antioxidant potential (TRAP); inflammatory biomarkers: fibrinogen, high-sensitivity C-reactive protein, erythrocyte sedimentation rate, interleukin-6 and tumor necrosis factor-α; and metabolic variables. Subjects were divided into those with (n=141) and without (n=201) a history of suicidal attempts. Individuals with a history of suicide attempts had significantly higher levels of NOx and lipid hydroperoxides and lowered TRAP as compared to individuals without suicide attempts. There were no significant associations between a history of suicide attempts and inflammatory and metabolic biomarkers and metabolic syndrome. Logistic regression showed that both unipolar and bipolar disorder, female gender, smoking behavior and lipid hydroperoxides were significantly associated with a history of suicide attempts. The combined effects of oxidative stress, smoking, depression, female gender were independent from classical risk factors, including marital status, years of education and anxiety. O&NS as well as lowered antioxidant levels may play a role in the pathophysiology of suicidal behavior independently from the effects of depression and smoking, both of which are associated with increased O&NS, and classical suicide predictors, such as years of education and marital status. © 2013 Published by Elsevier B.V.

PsMPK7, a stress-associated mitogen-activated protein kinase (MAPK) in Phytophthora sojae, is required for stress tolerance, reactive oxygenated species detoxification, cyst germination, sexual reproduction and infection of soybean.

Science.gov (United States)

Gao, Jian; Cao, Mingna; Ye, Wenwu; Li, Haiyang; Kong, Liang; Zheng, Xiaobo; Wang, Yuanchao

2015-01-01

The sensing of stress signals and their transduction into appropriate responses are crucial for the adaptation, survival and infection of phytopathogenic fungi and oomycetes. Amongst evolutionarily conserved pathways, mitogen-activated protein kinase (MAPK) cascades function as key signal transducers that use phosphorylation to convey information. In this study, we identified a gene, designated PsMPK7, one of 14 predicted genes encoding MAPKs in Phytophthora sojae. PsMPK7 was highly transcribed in each tested stage, but was up-regulated in the zoospore, cyst and cyst germination stages. Silencing of PsMPK7 affected the growth of germinated cysts, oospore production and the pathogenicity of soybean. PsMPK7 transcription was induced by stresses from sorbitol, NaCl and hydrogen peroxide. Transformants in which PsMPK7 expression was silenced (PsMPK7-silenced) were significantly more sensitive to osmotic and oxidative stress. Aniline blue and diaminobenzidine staining revealed that the silenced lines did not suppress the host reactive oxygen species (ROS) burst, indicating that either the inoculated plants activated stronger defence responses to the transformants and/or the PsMPK7-silenced transformants failed to overcome plant defences. In addition, extracellular secretion of laccase decreased in the silenced lines. Overall, our results indicate that the PsMPK7 gene encodes a stress-associated MAPK in P. sojae that is important not only for responses to various stresses, but also for ROS detoxification, cyst germination, sexual oospore production and infection of soybean. © 2014 BSPP AND JOHN WILEY & SONS LTD.

Endogenous stress proteins as targets for anti-inflammatory T cells

NARCIS (Netherlands)

Wieten, L.

2009-01-01

Stress proteins such as heat shock proteins (Hsp) are important controllers of both cellular and immune homeostasis. Enhanced Hsp expression can be observed in virtually every inflammatory condition and has been proposed by us and others to lead to local activation of Hsp-specific anti-inflammatory

Beyond the ENDF format: A modern nuclear database structure. SG38 meeting, NEA Headquarters, 29-30 November 2012

International Nuclear Information System (INIS)

McNabb, D.; Zerkin, V.; Mattoon, C.; Koning, A.; Brown, D.; Leal, L.; Sublet, J.C.; Coste-Delclaux, M.; Capote, R.; Forrest, R.; Kodeli, I.; Trkov, A.; Beck, B.; Haeck, W.; Fukahori, T.; Mills, R.W.; White, M.C.; Cullen, D.E.

2012-11-01

WPEC subgroup 38 (SG38) was formed to develop a new structure for storing nuclear reaction data, that is meant to eventually replace ENDF-6 as the standard way to store and share evaluations. The work of SG38 covers the following tasks: Designing flexible, general-purpose data containers; Determining a logical and easy-to-understand top-level hierarchy for storing evaluated nuclear reaction data; Creating a particle database for storing particles, masses and level schemes; Specifying the infrastructure (plotting, processing, etc.) that must accompany the new structure; Developing an Application Programming Interface or API to allow other codes to access data stored in the new structure; Specifying what tests need to be implemented for quality assurance of the new structure and associated infrastructure; Ensuring documentation and governance of the structure and associated infrastructure. This document is the proceedings of the SG38 meeting, held at the NEA Headquarters on 29-30 November 2012. It comprises all the available presentations (slides) given by the participants as well as 3 reports: A - Welcome and Introduction: - Purpose and goals for SG38 (D. McNabb); - Lessons from ENDF, EXFOR and other formats (V. Zerkin); - Lessons from first LLNL attempt at defining a new nuclear data structure (C. Mattoon); - Example of "2"3"9Pu data B - Purpose of the new data structure: - GND: Purpose of the new data structure (A. Koning); - Purpose of the new data structure: Dave's Perspective (D. Brown); C - Nuclear Data System Overview: - ENDF File uses in AMPX (L. Leal); D - Benefits and requirements for data evaluation and processing: - Benefits and requirements (J.C. Sublet); - CEA/DEN contribution (M. Coste-Delclaux); - Proposals from the IAEA-NDS (V. Zerkin, R. Capote, R. Forrest); - User View on the ENDF Formats and Data Processing (I. Kodeli); - On the ENDF Formats and Data Processing - report (A. Trkov); E - Format perspective, organization and requirements for basic

Identification and subcellular localization of porcine deltacoronavirus accessory protein NS6

International Nuclear Information System (INIS)

Fang, Puxian; Fang, Liurong; Liu, Xiaorong; Hong, Yingying; Wang, Yongle; Dong, Nan; Ma, Panpan; Bi, Jing; Wang, Dang; Xiao, Shaobo

2016-01-01

Porcine deltacoronavirus (PDCoV) is an emerging swine enteric coronavirus. Accessory proteins are genus-specific for coronavirus, and two putative accessory proteins, NS6 and NS7, are predicted to be encoded by PDCoV; however, this remains to be confirmed experimentally. Here, we identified the leader-body junction sites of NS6 subgenomic RNA (sgRNA) and found that the actual transcription regulatory sequence (TRS) utilized by NS6 is non-canonical and is located upstream of the predicted TRS. Using the purified NS6 from an Escherichia coli expression system, we obtained two anti-NS6 monoclonal antibodies that could detect the predicted NS6 in cells infected with PDCoV or transfected with NS6-expressing plasmids. Further studies revealed that NS6 is always localized in the cytoplasm of PDCoV-infected cells, mainly co-localizing with the endoplasmic reticulum (ER) and ER-Golgi intermediate compartments, as well as partially with the Golgi apparatus. Together, our results identify the NS6 sgRNA and demonstrate its expression in PDCoV-infected cells. -- Highlights: •The leader-body fusion site of NS6 sgRNA is identified. •NS6 sgRNA uses a non-canonical transcription regulatory sequence (TRS). •NS6 can be expressed in PDCoV-infected cell. •NS6 predominantly localize to the ER complex and ER-Golgi intermediate compartment.

Identification and subcellular localization of porcine deltacoronavirus accessory protein NS6

Energy Technology Data Exchange (ETDEWEB)

Fang, Puxian; Fang, Liurong; Liu, Xiaorong; Hong, Yingying; Wang, Yongle; Dong, Nan; Ma, Panpan [State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070 (China); The Cooperative Innovation Center for Sustainable Pig Production, Wuhan 430070 (China); Bi, Jing [State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070 (China); Department of Immunology and Aetology, College of Basic Medicine, Hubei University of Chinese Medicine, Wuhan 430065 (China); Wang, Dang [State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070 (China); The Cooperative Innovation Center for Sustainable Pig Production, Wuhan 430070 (China); Xiao, Shaobo, E-mail: vet@mail.hzau.edu.cn [State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070 (China); The Cooperative Innovation Center for Sustainable Pig Production, Wuhan 430070 (China)

2016-12-15

Porcine deltacoronavirus (PDCoV) is an emerging swine enteric coronavirus. Accessory proteins are genus-specific for coronavirus, and two putative accessory proteins, NS6 and NS7, are predicted to be encoded by PDCoV; however, this remains to be confirmed experimentally. Here, we identified the leader-body junction sites of NS6 subgenomic RNA (sgRNA) and found that the actual transcription regulatory sequence (TRS) utilized by NS6 is non-canonical and is located upstream of the predicted TRS. Using the purified NS6 from an Escherichia coli expression system, we obtained two anti-NS6 monoclonal antibodies that could detect the predicted NS6 in cells infected with PDCoV or transfected with NS6-expressing plasmids. Further studies revealed that NS6 is always localized in the cytoplasm of PDCoV-infected cells, mainly co-localizing with the endoplasmic reticulum (ER) and ER-Golgi intermediate compartments, as well as partially with the Golgi apparatus. Together, our results identify the NS6 sgRNA and demonstrate its expression in PDCoV-infected cells. -- Highlights: •The leader-body fusion site of NS6 sgRNA is identified. •NS6 sgRNA uses a non-canonical transcription regulatory sequence (TRS). •NS6 can be expressed in PDCoV-infected cell. •NS6 predominantly localize to the ER complex and ER-Golgi intermediate compartment.

Are stress proteins induced during PUVA therapy?

Energy Technology Data Exchange (ETDEWEB)

Al-Masaud, A.S. [Leeds Univ. (United Kingdom); Cunliffe, W.J.; Holland, D.B. [Leeds General Infirmary (United Kingdom)

1996-05-01

Heat shock or stress proteins are produced in practically all cell types when they are exposed to temperatures a few degrees above normal. Measurement of the skin temperature of patients undergoing psoralen and ultraviolet A (PUVA) cabinet treatment for psoriasis revealed that the outer layers of the skin experience a mean temperature rise of 5.3{sup o}C. However, this did not produce a detectable stress response in epidermal samples taken after PUVA treatment. In vitro exposure of epidermis from biopsies or of cultured keratinocytes to a 5-7{sup o}C temperature rise produced a heat shock response, as measured by an increase in the production of proteins of the HSP90 and HSP70 families. These results were confirmed by the use of specific monoclonal antibodies. The corresponding mRNAs were also analysed using labelled probes. In an in vitro system, following simulated PUVA treatment of cultured keratinocytes, increases in the synthesis of HSP90 and HSP70 were detected but these increases did not correlate with changes in mRNA levels. (author).

Are stress proteins induced during PUVA therapy?

International Nuclear Information System (INIS)

Al-Masaud, A.S.; Cunliffe, W.J.; Holland, D.B.

1996-01-01

Heat shock or stress proteins are produced in practically all cell types when they are exposed to temperatures a few degrees above normal. Measurement of the skin temperature of patients undergoing psoralen and ultraviolet A (PUVA) cabinet treatment for psoriasis revealed that the outer layers of the skin experience a mean temperature rise of 5.3 o C. However, this did not produce a detectable stress response in epidermal samples taken after PUVA treatment. In vitro exposure of epidermis from biopsies or of cultured keratinocytes to a 5-7 o C temperature rise produced a heat shock response, as measured by an increase in the production of proteins of the HSP90 and HSP70 families. These results were confirmed by the use of specific monoclonal antibodies. The corresponding mRNAs were also analysed using labelled probes. In an in vitro system, following simulated PUVA treatment of cultured keratinocytes, increases in the synthesis of HSP90 and HSP70 were detected but these increases did not correlate with changes in mRNA levels. (author)

Silicon Mitigates Salinity Stress by Regulating the Physiology, Antioxidant Enzyme Activities, and Protein Expression in Capsicum annuum ‘Bugwang'

Science.gov (United States)

Manivannan, Abinaya; Soundararajan, Prabhakaran; Muneer, Sowbiya; Ko, Chung Ho

2016-01-01

Silicon- (Si-) induced salinity stress resistance was demonstrated at physiological and proteomic levels in Capsicum annuum for the first time. Seedlings of C. annuum were hydroponically treated with NaCl (50 mM) with or without Si (1.8 mM) for 15 days. The results illustrated that saline conditions significantly reduced plant growth and biomass and photosynthetic parameters and increased the electrolyte leakage potential, lipid peroxidation, and hydrogen peroxide level. However, supplementation of Si allowed the plants to recover from salinity stress by improving their physiology and photosynthesis. During salinity stress, Si prevented oxidative damage by increasing the activities of antioxidant enzymes. Furthermore, Si supplementation recovered the nutrient imbalance that had occurred during salinity stress. Additionally, proteomic analysis by two-dimensional gel electrophoresis (2DE) followed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) revealed that Si treatment upregulated the accumulation of proteins involved in several metabolic processes, particularly those associated with nucleotide binding and transferase activity. Moreover, Si modulated the expression of vital proteins involved in ubiquitin-mediated nucleosome pathway and carbohydrate metabolism. Overall, the results illustrate that Si application induced resistance against salinity stress in C. annuum by regulating the physiology, antioxidant metabolism, and protein expression. PMID:27088085

Identification of Proteins Using iTRAQ and Virus-Induced Gene Silencing Reveals Three Bread Wheat Proteins Involved in the Response to Combined Osmotic-Cold Stress.

Science.gov (United States)

Zhang, Ning; Zhang, Lingran; Shi, Chaonan; Zhao, Lei; Cui, Dangqun; Chen, Feng

2018-05-25

Crops are often subjected to a combination of stresses in the field. To date, studies on the physiological and molecular responses of common wheat to a combination of osmotic and cold stresses, however, remain unknown. In this study, wheat seedlings exposed to osmotic-cold stress for 24 h showed inhibited growth, as well as increased lipid peroxidation, relative electrolyte leakage, and soluble sugar contents. iTRAQ-based quantitative proteome method was employed to determine the proteomic profiles of the roots and leaves of wheat seedlings exposed to osmotic-cold stress conditions. A total of 250 and 258 proteins with significantly altered abundance in the roots and leaves were identified, respectively, and the majority of these proteins displayed differential abundance, thereby revealing organ-specific differences in adaptation to osmotic-cold stress. Yeast two hybrid assay examined five pairs of stress/defense-related protein-protein interactions in the predicted protein interaction network. Furthermore, quantitative real-time PCR analysis indicated that abiotic stresses increased the expression of three candidate protein genes, i.e., TaGRP2, CDCP, and Wcor410c in wheat leaves. Virus-induced gene silencing indicated that three genes TaGRP2, CDCP, and Wcor410c were involved in modulating osmotic-cold stress in common wheat. Our study provides useful information for the elucidation of molecular and genetics bases of osmotic-cold combined stress in bread wheat.

Molecular characterization of a proteolysis-resistant lipase from Bacillus pumilus SG2

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R. Sangeetha

2014-06-01

Full Text Available Proteolysis-resistant lipases can be well exploited by industrial processes which employ both lipase and protease as biocatalysts. A proteolysis resistant lipase from Bacillus pumilus SG2 was isolated, purified and characterized earlier. The lipase was resistant to native and commercial proteases. In the present work, we have characterized the lip gene which encodes the proteolysis-resistant lipase from Bacillus pumilus SG2. The parameters and structural details of lipase were analysed. The lip gene consisted of 650 bp. The experimental molecular weight of SG2 lipase was nearly double that of its theoretical molecular weight, thus suggesting the existence of the functional lipase as a covalent dimer. The proteolytic cleavage sites of the lipase would have been made inaccessible by dimerisation, thus rendering the lipase resistant to protease.

Gamma-sarcoglycan is required for the response of archvillin to mechanical stimulation in skeletal muscle

Science.gov (United States)

Spinazzola, Janelle M.; Smith, Tara C.; Liu, Min; Luna, Elizabeth J.; Barton, Elisabeth R.

2015-01-01

Loss of gamma-sarcoglycan (γ-SG) induces muscle degeneration and signaling defects in response to mechanical load, and its absence is common to both Duchenne and limb girdle muscular dystrophies. Growing evidence suggests that aberrant signaling contributes to the disease pathology; however, the mechanisms of γ-SG-mediated mechanical signaling are poorly understood. To uncover γ-SG signaling pathway components, we performed yeast two-hybrid screens and identified the muscle-specific protein archvillin as a γ-SG and dystrophin interacting protein. Archvillin protein and message levels were significantly upregulated at the sarcolemma of murine γ-SG-null (gsg−/−) muscle but delocalized in dystrophin-deficient mdx muscle. Similar elevation of archvillin protein was observed in human quadriceps muscle lacking γ-SG. Reintroduction of γ-SG in gsg−/− muscle by rAAV injection restored archvillin levels to that of control C57 muscle. In situ eccentric contraction of tibialis anterior (TA) muscles from C57 mice caused ERK1/2 phosphorylation, nuclear activation of P-ERK1/2 and stimulus-dependent archvillin association with P-ERK1/2. In contrast, TA muscles from gsg−/− and mdx mice exhibited heightened P-ERK1/2 and increased nuclear P-ERK1/2 localization following eccentric contractions, but the archvillin–P-ERK1/2 association was completely ablated. These results position archvillin as a mechanically sensitive component of the dystrophin complex and demonstrate that signaling defects caused by loss of γ-SG occur both at the sarcolemma and in the nucleus. PMID:25605665

Microsecond molecular dynamics simulations of intrinsically disordered proteins involved in the oxidative stress response.

Directory of Open Access Journals (Sweden)

Elio A Cino

Full Text Available Intrinsically disordered proteins (IDPs are abundant in cells and have central roles in protein-protein interaction networks. Interactions between the IDP Prothymosin alpha (ProTα and the Neh2 domain of Nuclear factor erythroid 2-related factor 2 (Nrf2, with a common binding partner, Kelch-like ECH-associated protein 1(Keap1, are essential for regulating cellular response to oxidative stress. Misregulation of this pathway can lead to neurodegenerative diseases, premature aging and cancer. In order to understand the mechanisms these two disordered proteins employ to bind to Keap1, we performed extensive 0.5-1.0 microsecond atomistic molecular dynamics (MD simulations and isothermal titration calorimetry experiments to investigate the structure/dynamics of free-state ProTα and Neh2 and their thermodynamics of bindings. The results show that in their free states, both ProTα and Neh2 have propensities to form bound-state-like β-turn structures but to different extents. We also found that, for both proteins, residues outside the Keap1-binding motifs may play important roles in stabilizing the bound-state-like structures. Based on our findings, we propose that the binding of disordered ProTα and Neh2 to Keap1 occurs synergistically via preformed structural elements (PSEs and coupled folding and binding, with a heavy bias towards PSEs, particularly for Neh2. Our results provide insights into the molecular mechanisms Neh2 and ProTα bind to Keap1, information that is useful for developing therapeutics to enhance the oxidative stress response.

Prostaglandin E synthase interacts with inducible heat shock protein 70 after heat stress in bovine primary dermal fibroblast cells.

Science.gov (United States)

Richter, Constanze; Viergutz, Torsten; Schwerin, Manfred; Weitzel, Joachim M

2015-01-01

Exposure to heat stress in dairy cows leads to undesired side effects that are reflected by complex alterations in endocrine parameters, such as reduced progesterone, estradiol, and thyroid hormone concentrations. These endocrine maladaptation leads to failure to resume cyclicity, a poor uterine environment and inappropriate immune responses in postpartum dairy cows. Prostaglandins (PG's) are lipid mediators, which serve as signal molecules in response to various external stimuli as well as to cell-specific internal signal molecules. A central role in PG synthesis plays prostaglandin E synthase (PGES) that catalyzes the isomerization of PGH2 to PGE2 .The present study was conducted to investigate heat stress associated PGES expression. Expression of PGES and inducible heat shock protein 70 (HSP70), as a putative chaperonic protein, was studied in bovine primary fibroblasts under different heat shock conditions. Bovine primary fibroblasts produce PGE2 at homoiothermical norm temperature (38.5°C in bovine), but reduce PGE2 production rates under extreme heat stress (at 45°C for 6 h). By contrast, PGE2 production rates are maintained after a milder heat stress (at 41.5°C for 6 h). PGE2 synthesis is abolished by application of cyclooxygenase inhibitor indomethacin, indicating de novo synthesis. Heat stress increases HSP70 but not PGES protein concentrations. HSP70 physically interacts with PGES and the PGES-HSP70 complex did not dissociate upon heat stress at 45°C even after returning the cells to 37°C. The PGE2 production negatively correlates with the portion of PGES-HSP70 complex. These results suggest a protein interaction between HSP70 and PGES in dermal fibroblast cells. Blockage of PGES protein by HSP70 seems to interfere with the regulatory processes essential for cellular adaptive protection. © 2014 International Society for Advancement of Cytometry. © 2014 International Society for Advancement of Cytometry.

Oxidative stress and CCN1 protein in human skin connective tissue aging

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Zhaoping Qin

2016-06-01

Full Text Available Reactive oxygen species (ROS is an important pathogenic factor involved in human aging. Human skin is a primary target of oxidative stress from ROS generated from both extrinsic and intrinsic sources, like ultraviolet irradiation (UV and endogenous oxidative metabolism. Oxidative stress causes the alterations of collagen-rich extracellular matrix (ECM, the hallmark of skin connective tissue aging. Age-related alteration of dermal collagenous ECM impairs skin structural integrity and creates a tissue microenvironment that promotes age-related skin diseases, such as poor wound healing and skin cancer. Here, we review recent advances in our understanding of oxidative stress and CCN1 protein (first member of CCN family proteins, a critical mediator of oxidative stress-induced skin connective tissue aging.

Acute Heat Stress Changes Protein Expression in the Testes of a Broiler-Type Strain of Taiwan Country Chickens.

Science.gov (United States)

Wang, Shih-Han; Cheng, Chuen-Yu; Chen, Chao-Jung; Chan, Hong-Lin; Chen, Hsin-Hsin; Tang, Pin-Chi; Chen, Chih-Feng; Lee, Yen-Pai; Huang, San-Yuan

2018-03-19

Heat stress leads to decreased fertility in roosters. This study investigated the global protein expression in response to acute heat stress in the testes of a broiler-type strain of Taiwan country chickens (TCCs). Twelve 45-week-old roosters were randomly allocated to the control group maintained at 25°C, and three groups subjected to acute heat stress at 38°C for 4 h, with 0, 2, and 6 h of recovery, respectively. Testis samples were collected for hematoxylin and eosin staining, apoptosis assay, and protein analysis. The results revealed 101 protein spots that differed significantly from the control following exposure to acute heat stress. The proteins that were differentially expressed participated mainly in protein metabolism and other metabolic processes, responses to stimuli, apoptosis, cellular organization, and spermatogenesis. Proteins that negatively regulate apoptosis were downregulated and proteins involved in autophagy and major heat shock proteins (HSP90α, HSPA5, and HSPA8) were upregulated in the testes of heat-stressed chickens. In conclusion, acute heat stress causes a change in protein expression in the testes of broiler-type B strain TCCs and may thus impair cell morphology, spermatogenesis, and apoptosis. The expression of heat shock proteins increased to attenuate the testicular injury induced by acute heat stress.

The 'tubulin-like' S1 protein of Spirochaeta is a member of the hsp65 stress protein family

Science.gov (United States)

Munson, D.; Obar, R.; Tzertzinis, G.; Margulis, L.

1993-01-01

A 65-kDa protein (called S1) from Spirochaeta bajacaliforniensis was identified as 'tubulin-like' because it cross-reacted with at least four different antisera raised against tubulin and was isolated, with a co-polymerizing 45-kDa protein, by warm-cold cycling procedures used to purify tubulin from mammalian brain. Furthermore, at least three genera of non-cultivable symbiotic spirochetes (Pillotina, Diplocalyx, and Hollandina) that contain conspicuous 24-nm cytoplasmic tubules displayed a strong fluorescence in situ when treated with polyclonal antisera raised against tubulin. Here we summarize results that lead to the conclusion that this 65-kDa protein has no homology to tubulin. S1 is an hsp65 stress protein homologue. Hsp65 is a highly immunogenic family of hsp60 proteins which includes the 65-kDa antigens of Mycobacterium tuberculosis (an active component of Freund's complete adjuvant), Borrelia, Treponema, Chlamydia, Legionella, and Salmonella. The hsp60s, also known as chaperonins, include E. coli GroEL, mitochondrial and chloroplast chaperonins, the pea aphid 'symbionin' and many other proteins involved in protein folding and the stress response.

Learning from EDF investigations on SG divider plates and vessel head nozzles. Evidence of prior deformation effect on stress corrosion cracking

International Nuclear Information System (INIS)

Deforge, D.; Duisabeau, L.; Miloudi, S.; Thebault, Y.; Couvant, T.; Vaillant, F.; Lemaire, E.

2011-01-01

Nickel Based alloys Stress Corrosion Cracking (SCC) has been a major concern for all the Nuclear Power Plants (NPP) utilities since the beginning of the seventies. At EDF, the nineties were marked by the occurrence of cracks on vessel head nozzles. These cracks were responsible for a leak at Bugey 3 vessel head, which was the precursor leading to the replacement of all vessel heads. From 2002, new cases of Stress Corrosion Cracking were reported on Steam Generator (SG) Divider Plates (SGDP) welded junctions. These cracks are periodically inspected inservice and reparations could be performed in case of a significant evolution of the phenomenon even if the safety issue is less relevant than for the vessel head nozzles. Both issues have led to an important non-destructive testing (NDT) program and to destructive investigations campaigns. NDT were performed on an exhaustive basis for all vessel head nozzles and for all the divider plates of 900 MWe plants. Destructive investigations were performed on more than 30 vessel head nozzles and on 6 divider plates. The last investigations were performed on samples from two decommissioned Steam Generators of Chinon B1 which present SCC cracks. In this paper, the main conclusions driven from the analysis of both NDT and destructive investigation results are reported and a comparison of the behaviours of divider plates and vessel head nozzles is given. Results give evidence that prior plastic deformation of the components before operation is fundamental for the further environmental behaviour of the material. Analysis of field experience based on parameters characteristics of prior deformation and parameters characteristics of material microstructure can be used to account for the components which are the most sensitive to SCC cracking. Some perspectives on SCC predictive models are also presented. (authors)

Information Flow Analysis for Human-System Interaction in the SG Level Control

International Nuclear Information System (INIS)

Kim, Jong Hyun; Shin, Yeong Cheol

2008-01-01

Interaction between automatic control and operators is one of main issues in the application of automation technology. Inappropriate information from automatic control systems causes unexpected problems in human-automation collaboration. Poor information becomes critical, especially when the operator takes over the control from an automation system. Operators cannot properly handle the situation transferred from the automatic mode because of inadequate situation awareness, if the operator is out-of-the loop and the automatic control system fails. Some cases of unplanned reactor trips during the transition between the manual mode and the automatic mode are reported in nuclear power plants (NPPs). Among unplanned reactor trips since 2002, two cases were partially caused by automation-related failures of steam generator (SG) level control. This paper conducts information flow analysis to identify information and control requirement for human-system interaction of SG level control. At first, this paper identifies the level of automation in SG level control systems and then function allocation between system control and human operators. Then information flow analysis for monitoring and transition of automation is performed by adapting job process chart. Information and control requirements will be useful as an input for the human-system interface (HSI) design of SG level control

Identification of interacting proteins of retinoid-related orphan nuclear receptor gamma in HepG2 cells

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Ze-Min Huang1,#, Jun Wu2,#, Zheng-Cai Jia1, Yi Tian1, Jun Tang3, Yan Tang1, Ying Wang2, Yu-Zhang Wu1,* & Bing Ni1,*

2012-06-01

Full Text Available The retinoid-related orphan nuclear receptor gamma (RORγplays critical roles in regulation of development, immunity andmetabolism. As transcription factor usually forms a proteincomplex to function, thus capturing and dissecting of theRORγ protein complex will be helpful for exploring themechanisms underlying those functions. After construction ofthe recombinant tandem affinity purification (TAP plasmid,pMSCVpuro RORγ-CTAP(SG, the nuclear localization ofRORγ-CTAP(SG fusion protein was verified. Followingisolation of RORγ protein complex by TAP strategy, sevencandidate interacting proteins were identified. Finally, the heatshock protein 90 (HSP90 and receptor-interacting protein 140(RIP140 were confirmed to interplay with RORγ byco-immunoprecipitation. Interference of HSP90 or/and RIP140genes resulted in dramatically decreased expression ofCYP2C8 gene, the RORγ target gene. Data from this studydemonstrate that HSP90 and RIP140 proteins interact withRORγ protein in a complex format and function asco-activators in the RORγ-mediated regulatory processes ofHepG2 cells.

Momordica charantia (bitter melon attenuates high-fat diet-associated oxidative stress and neuroinflammation

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Feher Domonkos

2011-06-01

Full Text Available Abstract Background The rising epidemic of obesity is associated with cognitive decline and is considered as one of the major risk factors for neurodegenerative diseases. Neuroinflammation is a critical component in the progression of several neurological and neurodegenerative diseases. Increased metabolic flux to the brain during overnutrition and obesity can orchestrate stress response, blood-brain barrier (BBB disruption, recruitment of inflammatory immune cells from peripheral blood and microglial cells activation leading to neuroinflammation. The lack of an effective treatment for obesity-associated brain dysfunction may have far-reaching public health ramifications, urgently necessitating the identification of appropriate preventive and therapeutic strategies. The objective of our study was to investigate the neuroprotective effects of Momordica charantia (bitter melon on high-fat diet (HFD-associated BBB disruption, stress and neuroinflammatory cytokines. Methods C57BL/6 female mice were fed HFD with and without bitter melon (BM for 16 weeks. BBB disruption was analyzed using Evans blue dye. Phosphate-buffered saline (PBS perfused brains were analyzed for neuroinflammatory markers such as interleukin-22 (IL-22, IL-17R, IL-16, NF-κB1, and glial cells activation markers such as Iba1, CD11b, GFAP and S100β. Additionally, antioxidant enzymes, ER-stress proteins, and stress-resistant transcription factors, sirtuin 1 (Sirt1 and forkhead box class O transcription factor (FoxO were analyzed using microarray, quantitative real-time RT-PCR, western immunoblotting and enzymatic assays. Systemic inflammation was analyzed using cytokine antibody array. Results BM ameliorated HFD-associated changes in BBB permeability as evident by reduced leakage of Evans blue dye. HFD-induced glial cells activation and expression of neuroinflammatory markers such as NF-κB1, IL-16, IL-22 as well as IL-17R were normalized in the brains of mice supplemented with BM

[Screening differentially expressed plasma proteins in cold stress rats based on iTRAQ combined with mass spectrometry technology].

Science.gov (United States)

Liu, Yan-zhi; Guo, Jing-ru; Peng, Meng-ling; Ma, Li; Zhen, Li; Ji, Hong; Yang, Huan-min

2015-09-01

Isobaric tags for relative and absolute quantitation (iTRAQ) combined with mass spectrometry were used to screen differentially expressed plasma proteins in cold stress rats. Thirty health SPF Wistar rats were randomly divided into cold stress group A and control group B, then A and B were randomly divided into 3 groups (n = 5): A1, A2, A3 and B1, B2, B3. The temperature of room raising was (24.0 +/- 0.1) degrees C, and the cold stress temperature was (4.0 +/- 0.1) degrees C. The rats were treated with different temperatures until 12 h. The abdominal aortic blood was collected with heparin anticoagulation suction tube. Then, the plasma was separated for protein extraction, quantitative, enzymolysis, iTHAQ labeling, scx fractionation and mass spectrometry analysis. Totally, 1085 proteins were identified in the test, 39 differentially expressed proteins were screened, including 29 up-regulated proteins and 10 down-regulated proteins. Three important differentially expressed proteins related to cold stress were screened by bioinfonnatics analysis (Minor histocompatihility protein HA-1, Has-related protein Rap-1b, Integrin beta-1). In the experiment, the differentially expressed plasma proteins were successfully screened in cold stress rats. iTRAQ technology provided a good platform to screen protein diaguostic markers on cold stress rats, and laid a good foundation for further. study on animal cold stress mechanism.

Cross-lagged associations between children's stress and adiposity: the Children's Body Composition and Stress study.

Science.gov (United States)

Michels, Nathalie; Sioen, Isabelle; Boone, Liesbet; Clays, Els; Vanaelst, Barbara; Huybrechts, Inge; De Henauw, Stefaan

2015-01-01

The public health threats stress and adiposity have previously been associated with each other. Longitudinal studies are needed to reveal whether this association is bidirectional and the moderating factors. In the longitudinal Children's Body Composition and Stress study, 316 children (aged 5-12 years) had measures of stress (questionnaires concerning negative life events, problem behavior, and emotions) and adiposity (body mass index, waist-to-height ratio, and fat percentage) in three waves at 1-year intervals. The bidirectionality of the association between stress and adiposity was examined using cross-lagged analyses. We tested moderation by cortisol and life-style (physical activity, screen time, food consumption, eating behavior and sleep duration). Adiposity (body mass index: β = 0.48 and fat percentage: β = 0.18; p stress levels, but stress was not directly related to subsequent increases in adiposity indices. Cortisol and life-style factors displayed a moderating effect on the association between stress and adiposity. Stress was positively associated with adiposity in children with high cortisol awakening patterns (β = 0.204; p = .020) and high sweet food consumption (β = 0.190; p = .031), whereas stress was associated with lower adiposity in the most active children (β = -0.163; p = .022). Stress is associated with the development of children's adiposity, but the effects depend on cortisol levels and life-style factors. This creates new perspectives for multifactorial obesity prevention programs. Our results also highlight the adverse effect of an unhealthy body composition on children's psychological well-being.

A Molecular Web: Endoplasmic Reticulum Stress, Inflammation and Oxidative Stress

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Namrata eChaudhari

2014-07-01

Full Text Available Execution of fundamental cellular functions demands regulated protein folding homeostasis. Endoplasmic reticulum (ER is an active organelle existing to implement this function by folding and modifying secretory and membrane proteins. Loss of protein folding homeostasis is central to various diseases and budding evidences suggest ER stress as being a major contributor in the development or pathology of a diseased state besides other cellular stresses. The trigger for diseases may be diverse but, inflammation and/or ER stress may be basic mechanisms increasing the severity or complicating the condition of the disease. Chronic ER stress and activation of the unfolded protein response (UPR through endogenous or exogenous insults may result in impaired calcium and redox homeostasis, oxidative stress via protein overload thereby also influencing vital mitochondrial functions. Calcium released from the ER augments the production of mitochondrial Reactive Oxygen Species (ROS. Toxic accumulation of ROS within ER and mitochondria disturb fundamental organelle functions. Sustained ER stress is known to potentially elicit inflammatory responses via UPR pathways. Additionally, ROS generated through inflammation or mitochondrial dysfunction could accelerate ER malfunction. Dysfunctional UPR pathways has been associated with a wide range of diseases including several neurodegenerative diseases, stroke, metabolic disorders, cancer, inflammatory disease, diabetes mellitus, cardiovascular disease and others. In this review we have discussed the UPR signaling pathways, and networking between ER stress induced inflammatory pathways, oxidative stress and mitochondrial signaling events which further induce or exacerbate ER stress.

Estimation of S/G ratio in woods using 1064 nm FT-Raman spectroscopy

Science.gov (United States)

Umesh P. Agarwal; Sally A. Ralph; Dharshana Padmakshan; Sarah Liu; Steven D. Karlen; Cliff Foster; John Ralph

2015-01-01

Two simple methods based on the 370 cm-1 Raman band intensity were developed for estimation of syringyl-to-guaiacyl (S/G) ratio in woods. The methods, in principle, are representative of the whole cell wall lignin and not just the portion of lignin that gets cleaved to release monomers, for example, during certain S/G chemical analyses. As such,...

Oxidative stress and Ramadan observance; a possible influence of associated dieting

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RJ Shephard

2017-06-01

Full Text Available Introduction: The effects of Ramadan observance and any associated dietary restriction upon oxidative stress are not well known. The topic has thus been examined in a brief systematic review of available literature concerning non-athletic but otherwise healthy subjects, patients with selected clinical conditions, and in athletes. Methods: Ovid/Medline and Google searches were supplemented by a perusal of reference lists in papers thus identified. Results: Ramadan observance and associated dietary restrictions are generally associated with a decrease of body mass in non-athletic adults, and in patients with conditions such as obesity, metabolic syndrome, diabetes mellitus and hypertension. During Ramadan, measures of oxidative stress (particularly malondialdehyde and F2 isoprostanes are consistently decreased, antioxidant status (particularly levels of peroxidases, uric acid and reduced glutathione are enhanced and inflammatory reactions (particularly c-reactive protein, IL-6 and TNF-a are decreased in association with decreases in body mass. Perhaps because of lower initial body weights and greater dietary control during Ramadan, changes of oxidant status are more variable in athletes; in 3 of 7 studies, Ramadan observance had little effect on oxidant status, and in 2 reports there was some deterioration. In 3 of 4 studies where athletes underwent short-term dieting, there was also no improvement of antioxidant status. Conclusion: Ramadan observance and any associated dieting reduce oxidative stress in non-athletic individuals, apparently in association with decreases of body mass. In athletes, oxidant levels are generally unchanged during Ramadan, and if food intake is maintained they may even increase. More information is needed upon possible adverse health consequences, but chronic risks are probably small because any changes are limited to one month per year.

Ribosomal elongation factor 4 promotes cell death associated with lethal stress.

Science.gov (United States)

Li, Liping; Hong, Yuzhi; Luan, Gan; Mosel, Michael; Malik, Muhammad; Drlica, Karl; Zhao, Xilin

2014-12-09

Ribosomal elongation factor 4 (EF4) is highly conserved among bacteria, mitochondria, and chloroplasts. However, the EF4-encoding gene, lepA, is nonessential and its deficiency shows no growth or fitness defect. In purified systems, EF4 back-translocates stalled, posttranslational ribosomes for efficient protein synthesis; consequently, EF4 has a protective role during moderate stress. We were surprised to find that EF4 also has a detrimental role during severe stress: deletion of lepA increased Escherichia coli survival following treatment with several antimicrobials. EF4 contributed to stress-mediated lethality through reactive oxygen species (ROS) because (i) the protective effect of a ΔlepA mutation against lethal antimicrobials was eliminated by anaerobic growth or by agents that block hydroxyl radical accumulation and (ii) the ΔlepA mutation decreased ROS levels stimulated by antimicrobial stress. Epistasis experiments showed that EF4 functions in the same genetic pathway as the MazF toxin, a stress response factor implicated in ROS-mediated cell death. The detrimental action of EF4 required transfer-messenger RNA (tmRNA, which tags truncated proteins for degradation and is known to be inhibited by EF4) and the ClpP protease. Inhibition of a protective, tmRNA/ClpP-mediated degradative activity would allow truncated proteins to indirectly perturb the respiratory chain and thereby provide a potential link between EF4 and ROS. The connection among EF4, MazF, tmRNA, and ROS expands a pathway leading from harsh stress to bacterial self-destruction. The destructive aspect of EF4 plus the protective properties described previously make EF4 a bifunctional factor in a stress response that promotes survival or death, depending on the severity of stress. Translation elongation factor 4 (EF4) is one of the most conserved proteins in nature, but it is dispensable. Lack of strong phenotypes for its genetic knockout has made EF4 an enigma. Recent biochemical work has

Membrane-associated proteomics of chickpea identifies Sad1/UNC-84 protein (CaSUN1), a novel component of dehydration signaling

Science.gov (United States)

Jaiswal, Dinesh Kumar; Mishra, Poonam; Subba, Pratigya; Rathi, Divya; Chakraborty, Subhra; Chakraborty, Niranjan

2014-02-01

Dehydration affects almost all the physiological processes including those that result in the accumulation of misfolded proteins in the endoplasmic reticulum (ER), which in turn elicits a highly conserved signaling, the unfolded protein response (UPR). We investigated the dehydration-responsive membrane-associated proteome of a legume, chickpea, by 2-DE coupled with mass spectrometry. A total of 184 protein spots were significantly altered over a dehydration treatment of 120 h. Among the differentially expressed proteins, a non-canonical SUN domain protein, designated CaSUN1 (Cicer arietinum Sad1/UNC-84), was identified. CaSUN1 localized to the nuclear membrane and ER, besides small vacuolar vesicles. The transcripts were downregulated by both abiotic and biotic stresses, but not by abscisic acid treatment. Overexpression of CaSUN1 conferred stress tolerance in transgenic Arabidopsis. Furthermore, functional complementation of the yeast mutant, slp1, could rescue its growth defects. We propose that the function of CaSUN1 in stress response might be regulated via UPR signaling.

Factors Associated with Urinary Stress Incontinence in Primiparas

OpenAIRE

Pei-Ling Chou; Fang-Ping Chen; Li-Fen Teng

2005-01-01

Objective: To evaluate obstetric and maternal risk factors for stress urinary incontinence in primiparas. Materials and Methods: From January 2001 to August 2002, 378 primiparas were interviewed about stress urinary incontinence 1 year after delivery. The association between symptoms of urinary stress incontinence and obstetric factors was assessed. Results: Twenty-four (6%) primiparas had urinary stress incontinence after delivery. Maternal age was positively associated with urinary st...

Protein Thiols as an Indication of Oxidative Stress

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Yousef Rezaei Chianeh

2014-06-01

Full Text Available Thiol is an organic compound that contain sulphhydryl group that have a critical role in preventing any involvement of oxidative stress in the cell. These defensive functions are generally considered to be carried out by the low molecular weight thiol glutathione and by cysteine residues in the active sites of proteins such as thioredoxin and peroxiredoxin. In addition, there are thiols exposed on protein surfaces that are not directly involved with protein function, although they can interact with the intracellular environment.The process of protection of the cell against an oxidative damage occur by thiol and cystein residue that has a low molecular weight. These residue are present in the active sites of a protein like, peroxiredoxin and thioredoxin. Apart from intracellular antioxidant defense mechanism by protein thiol, there are presence of thiol in outer surface of protein that are not involved with the function of protein, even though they can interact with intracellular part of the cell. [Archives Medical Review Journal 2014; 23(3.000: 443-456

Beyond the ENDF format: A modern nuclear database structure. SG38 meeting, NEA Headquarters, 20-22 May 2015

International Nuclear Information System (INIS)

Brown, David; Jouanne, Cedric; Malvagi, Fausto; Coste-Delclaux, Mireille; Hawari, Ayman I.; Mattoon, Caleb; Zerkin, Viktor; ); Cornock, Mark; Conlin, Jeremy; Lloydge, Zhigang; Oleynik, Dmitry S.; Cabellos, Oscar; Mills, Robert W.; Kim, Do Heon; Leal, Luiz Carlos; White, Morgan c.; Dunn, Michael; Kahler, Albert C. skip; Mcnabb, Dennis P.; Roubtsov, Danila; Cho, Young-Sik; Beck, Bret; Haeck, Wim

2015-05-01

WPEC subgroup 38 (SG38) was formed to develop a new structure for storing nuclear reaction data, that is meant to eventually replace ENDF-6 as the standard way to store and share evaluations. The work of SG38 covers the following tasks: Designing flexible, general-purpose data containers; Determining a logical and easy-to-understand top-level hierarchy for storing evaluated nuclear reaction data; Creating a particle database for storing particles, masses and level schemes; Specifying the infrastructure (plotting, processing, etc.) that must accompany the new structure; Developing an Application Programming Interface or API to allow other codes to access data stored in the new structure; Specifying what tests need to be implemented for quality assurance of the new structure and associated infrastructure; Ensuring documentation and governance of the structure and associated infrastructure. This document is the proceedings of the SG38 meeting which took place at the NEA Headquarters, on 20-22 May 2015. It comprises all the available presentations (slides) given by the participants: - 1) Recent updates to the top-level hierarchy, review draft requirements document (D. Brown); - 2) Properties of Particles (PoPs) database (C.M. Mattoon); - 3) Proposed General Purpose Data Containers (B. Beck); - 4) Feedback on GND Specifications (C. Jouanne); - 5) Using XML in the IAEA-NDS: status, feedback and proposals (V. Zerkin); - 6) Observations Related to Thermal Neutron Scattering Law Data (A.I. Hawari); - 7) Meeting Notes (D.P. Mcnabb)

Life prediction of steam generator tubing due to stress corrosion crack using Monte Carlo Simulation

International Nuclear Information System (INIS)

Hu Jun; Liu Fei; Cheng Guangxu; Zhang Zaoxiao

2011-01-01

Highlights: → A life prediction model for SG tubing was proposed. → The initial crack length for SCC was determined. → Two failure modes called rupture mode and leak mode were considered. → A probabilistic life prediction code based on Monte Carlo method was developed. - Abstract: The failure of steam generator tubing is one of the main accidents that seriously affects the availability and safety of a nuclear power plant. In order to estimate the probability of the failure, a probabilistic model was established to predict the whole life-span and residual life of steam generator (SG) tubing. The failure investigated was stress corrosion cracking (SCC) after the generation of one through-wall axial crack. Two failure modes called rupture mode and leak mode based on probabilistic fracture mechanics were considered in this proposed model. It took into account the variance in tube geometry and material properties, and the variance in residual stresses and operating conditions, all of which govern the propagations of cracks. The proposed model was numerically calculated by using Monte Carlo Simulation (MCS). The plugging criteria were first verified and then the whole life-span and residual life of the SG tubing were obtained. Finally, important sensitivity analysis was also carried out to identify the most important parameters affecting the life of SG tubing. The results will be useful in developing optimum strategies for life-cycle management of the feedwater system in nuclear power plants.

Comparative proteomics analysis of oral cancer cell lines: identification of cancer associated proteins

Science.gov (United States)

2014-01-01

Background A limiting factor in performing proteomics analysis on cancerous cells is the difficulty in obtaining sufficient amounts of starting material. Cell lines can be used as a simplified model system for studying changes that accompany tumorigenesis. This study used two-dimensional gel electrophoresis (2DE) to compare the whole cell proteome of oral cancer cell lines vs normal cells in an attempt to identify cancer associated proteins. Results Three primary cell cultures of normal cells with a limited lifespan without hTERT immortalization have been successfully established. 2DE was used to compare the whole cell proteome of these cells with that of three oral cancer cell lines. Twenty four protein spots were found to have changed in abundance. MALDI TOF/TOF was then used to determine the identity of these proteins. Identified proteins were classified into seven functional categories – structural proteins, enzymes, regulatory proteins, chaperones and others. IPA core analysis predicted that 18 proteins were related to cancer with involvements in hyperplasia, metastasis, invasion, growth and tumorigenesis. The mRNA expressions of two proteins – 14-3-3 protein sigma and Stress-induced-phosphoprotein 1 – were found to correlate with the corresponding proteins’ abundance. Conclusions The outcome of this analysis demonstrated that a comparative study of whole cell proteome of cancer versus normal cell lines can be used to identify cancer associated proteins. PMID:24422745

KvLEA, a New Isolated Late Embryogenesis Abundant Protein Gene from Kosteletzkya virginica Responding to Multiabiotic Stresses

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Xiaoli Tang

2016-01-01

Full Text Available The LEA proteins are a kind of hydrophilic proteins, playing main functions in desiccation tolerance. However, their importance as a kind of stress proteins in abiotic stress is being clarified little by little. In this study we isolated, cloned, and identified the first KvLEA gene in Kosteletzkya virginica. Bioinformatic analysis showed that the protein encoded by this gene had common properties of LEA proteins and the multiple sequences alignment and phylogenetic analysis further showed that this protein had high homology with two Arabidopsis LEA proteins. Gene expression analysis revealed that this gene had a higher expression in root and it was induced obviously by salt stress. Moreover, the transcripts of KvLEA were also induced by other abiotic stresses including drought, high temperature, chilling, and ABA treatment. Among these abiotic stresses, ABA treatment brought about the biggest changes to this gene. Collectively, our research discovered a novel LEA gene and uncovered its involvement in multiabiotic stresses in K. virginica. This research not only enriched studies on LEA gene in plant but also would accelerate more studies on K. virginica in the future.

Heat shock protein 27 phosphorylation state is associated with cancer progression

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Maria eKatsogiannou

2014-10-01

Full Text Available Understanding the mechanisms that control stress-induced survival is critical to explain how tumors frequently resist to treatment and to improve current anti-cancer therapies. Cancer cells are able to cope with stress and escape drug toxicity by regulating heat shock proteins (Hsps expression and function. Hsp27 (HSPB1, a member of the small Hsp family, represents one of the key players of many signaling pathways contributing to tumorigenicity, treatment resistance and apoptosis inhibition. Hsp27 is overexpressed in many types of cancer and its functions are regulated by post-translational modifications, such as phosphorylation. Protein phosphorylation is the most widespread signaling mechanism in eukaryotic cells, and it is involved in all fundamental cellular processes. Aberrant phosphorylation of Hsp27 has been associated with several diseases such as cancer but the molecular mechanisms by which it is implicated in cancer development and progression remain undefined. This review focuses on the role of phosphorylation in Hsp27 functions in cancer cells and its potential usefulness as therapeutic target in cancer.

Biochemical analysis of cerebrospinal fluid in the laboratories of deployed medical treatment facilities: are Multistix 10 SG strip and iSTAT useful?

Science.gov (United States)

Lefrere, Bertrand; Plantamura, J; Renard, C; Ceppa, F; Delacour, H

2017-12-01

During military deployment, the diagnosis and the management of acute bacterial meningitis can be problematic, as deployed Medical Treatment Facilities (MTFs) often have a limited laboratory diagnostic capability. However, French Role 2 and 3 MTFs have point-of-care (POC) testing to perform urinary (Multistix 10 SG strip) and blood (iSTAT handheld analyser) biochemical testing mentioned in AMedP8.5. The purpose of this study was to compare the accuracy of this urine test strip and of the iSTAT CHEM8 and CG4 cartridges with a standard hospital bench top analyser in order to determine if these POC devices have a potential role in the biochemical analysis of cerebrospinal fluid (CSF protein, CSF glucose and CSF lactate, respectively). Agreement between the index methods and the reference methods (suitable kits on the Cobas 6 000 System) was evaluated by parallel testing of 30 CSF samples by both techniques. For CSF protein, agreement between the strip and the reference method was evaluated determining the κ coefficient. For CSF glucose and CSF lactate subgroups, least squares linear regressions were calculated and Bland-Altman analyses were performed. The Multistix 10 SG strip can be used to make a semiquantitative determination of CSF protein. A good agreement between the strip and the reference method was observed (κ coefficient: 0.93 (IC 95 0.82 to 1)). This strip is thus well adapted to demonstrate an elevation of CSF protein level as observed in acute bacterial meningitis. The iSTAT CHEM8 and CG4+ cartridges correlated well with the reference methods for the determination of CSF glucose and CSF lactate, respectively (r 2 >0.98) but exhibited a negative bias (∼ -7% and ∼ -15%, respectively). The combined use of the Multistix 10 SG strip and of the iSTAT system appears to be an attractive solution for the biochemical investigation of CSF in medical treatment facilities with limited laboratory diagnostic capability. Published by the BMJ Publishing Group

Involvement of Calmodulin and Calmodulin-like Proteins in Plant Responses to Abiotic Stresses

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B W Poovaiah

2015-08-01

Full Text Available Transient changes in intracellular Ca2+ concentration have been well recognized to act as cell signals coupling various environmental stimuli to appropriate physiological responses with accuracy and specificity in plants. Calmodulin (CaM and calmodulin-like proteins (CMLs are major Ca2+ sensors, playing critical roles in interpreting encrypted Ca2+ signals. Ca2+-loaded CaM/CMLs interact and regulate a broad spectrum of target proteins such as channels/pumps/antiporters for various ions, transcription factors, protein kinases, protein phosphatases, metabolic enzymes and proteins with unknown biochemical functions. Many of the target proteins of CaM/CMLs directly or indirectly regulate plant responses to environmental stresses. Basic information about stimulus-induced Ca2+ signal and overview of Ca2+ signal perception and transduction are briefly discussed in the beginning of this review. How CaM/CMLs are involved in regulating plant responses to abiotic stresses are emphasized in this review. Exciting progress has been made in the past several years, such as the elucidation of Ca2+/CaM-mediated regulation of AtSR1/CAMTA3 and plant responses to chilling and freezing stresses, Ca2+/CaM-mediated regulation of CAT3, MAPK8 and MKP1 in homeostasis control of ROS signals, discovery of CaM7 as a DNA-binding transcription factor regulating plant response to light signals. However, many key questions in Ca2+/CaM-mediated signaling warrant further investigation. Ca2+/CaM-mediated regulation of most of the known target proteins is presumed based on their interaction. The downstream targets of CMLs are mostly unknown, and how specificity of Ca2+ signaling could be realized through the actions of CaM/CMLs and their target proteins is largely unknown. Future breakthroughs in Ca2+/CaM-mediated signaling will not only improve our understanding of how plants respond to environmental stresses, but also provide the knowledge base to improve stress-tolerance of crops.

Hypothesis: NDL proteins function in stress responses by regulating microtubule organization

OpenAIRE

Khatri, Nisha; Mudgil, Yashwanti

2015-01-01

N-MYC DOWNREGULATED-LIKE proteins (NDL), members of the alpha/beta hydrolase superfamily were recently rediscovered as interactors of G-protein signaling in Arabidopsis thaliana. Although the precise molecular function of NDL proteins is still elusive, in animals these proteins play protective role in hypoxia and expression is induced by hypoxia and nickel, indicating role in stress. Homology of NDL1 with animal counterpart N-MYC DOWNREGULATED GENE (NDRG) suggests similar functions in animals...

The Zinc-Finger Thylakoid-Membrane Protein FIP Is Involved With Abiotic Stress Response in Arabidopsis thaliana

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Karina L. Lopes

2018-04-01

Full Text Available Many plant genes have their expression modulated by stress conditions. Here, we used Arabidopsis FtsH5 protease, which expression is regulated by light stress, as bait in a yeast two-hybrid screen to search for new proteins involved in the stress response. As a result, we found FIP (FtsH5 Interacting Protein, which possesses an amino proximal cleavable transit peptide, a hydrophobic membrane-anchoring region, and a carboxyl proximal C4-type zinc-finger domain. In vivo experiments using FIP fused to green fluorescent protein (GFP showed a plastid localization. This finding was corroborated by chloroplast import assays that showed FIP inserted in the thylakoid membrane. FIP expression was down-regulated in plants exposed to high light intensity, oxidative, salt, and osmotic stresses, whereas mutant plants expressing low levels of FIP were more tolerant to these abiotic stresses. Our data shows a new thylakoid-membrane protein involved with abiotic stress response in Arabidopsis thaliana.

The Hydrogen Detection Technique for SG Protection System

International Nuclear Information System (INIS)

Lv Mingyu; Pei Zhiyong; Yu Huajin

2015-01-01

SG that is pressure boundary between secondary loop and triple loop is the key equipment of fast reactor, in which heat in secondary loop is transferred to water or steam in triple loop. According to data from IAEA, SG is the highest failure rate equipment in fast reactor, especially because of failure of heat transfer tube. In order to monitor failure of heat transfer tube, Fast Reactor Engineering Department develops diffusion type hydrogen detection system, which is used to detect sodium-water reaction in time. This paper firstly introduces experimental research scheme and results of this hydrogen detection technique; Subsequently, it is described that how this technique can be engineering realized in CEFR; Moreover, through developing a series of calibration tests and hydrogen injection tests, it is obtained that sensitivity, response time and calibration curse for hydrogen detection system of CEFR. (author)

Comparative proteome analysis of metabolic proteins from seeds of durum wheat (cv. Svevo) subjected to heat stress

DEFF Research Database (Denmark)

Laino, Paolo; Shelton, Dale; Finnie, Christine

2010-01-01

of nonprolamin proteins were monitored to identify polypeptides affected by heat stress during grain fill. This study shows that heat stress alters significantly the durum wheat seed proteome, although the changes range is only between 1.2- and 2.2-fold. This analysis revealed 132 differentially expressed...... include proteins with metabolic activity or structural function. In order to investigate the consequences of heat stress on the accumulation of nonprolamin proteins in mature durum wheat kernels, the Italian cultivar Svevo was subjected to two thermal regimes (heat stress versus control). The 2-D patterns...... polypeptides, 47 of which were identified by MALDI-TOF and MALDI-TOF-TOF MS and included HSPs, proteins involved in the glycolysis and carbohydrate metabolism, as well as stress-related proteins. Many of the heat-induced polypeptides are considered to be allergenic for sensitive individuals....

The role of the open-quotes stress protein responseclose quotes in hormesis

International Nuclear Information System (INIS)

Smith-Sonneborn, J.

1992-01-01

Hormesis refers to the phenomenon of induction of beneficial effects by low doses of otherwise harmful physical or chemical agents: 'a little bit of bad can be good for you.' That the hormetic response may operate by a common mechanism has already been proposed, but this review is the first to propose the hypothesis that the common pathway is a heat shock-like response. The heat shock response is a model for a more general phenomenon, called the stress response. The stress response is characterized by increased synthesis of a family of stressor specific proteins with concomitant reduction of synthesis of most of the proteins transcribed prior to the exposure to the toxic agent. The stress response has been characterized using heat, radiation, heavy metals, and oxidizing agents as the stressors. This chapter includes: Identification of agents known to induce both the stress response and hormetic phenomena; A description of the unique and common pathways in the stress response to three stressors - heat, DNA-damaging agents, and teratogens; The stress response as a model for teratogen-induced damage; A theory explaining the paradoxical beneficial response to low doses of an otherwise harmful agent via a stress-response pathway

Study on Temperature Control System Based on SG3525

Science.gov (United States)

Cheng, Cong; Zhu, Yifeng; Wu, Junfeng

2017-12-01

In this paper, it uses the way of dry bath temperature to heat the microfluidic chip directly by the heating plate and the liquid sample in microfluidic chip is heated through thermal conductivity, thus the liquid sample will maintain at target temperature. In order to improve the reliability of the whole machine, a temperature control system based on SG3525 is designed.SG3525 is the core of the system which uses PWM wave produced by itself to drive power tube to heat the heating plate. The bridge circuit consisted of thermistor and PID regulation ensure that the temperature can be controlled at 37 °C with a correctness of ± 0.2 °C and a fluctuation of ± 0.1 °C.

Exploring the Plant–Microbe Interface by Profiling the Surface-Associated Proteins of Barley Grains

DEFF Research Database (Denmark)

Sultan, Abida; Andersen, Birgit; Svensson, Birte

2016-01-01

Cereal grains are colonized by a microbial community that actively interacts with the plant via secretion of various enzymes, hormones, and metabolites. Microorganisms decompose plant tissues by a collection of depolymerizing enzymes, including β-1,4-xylanases, that are in turn inhibited by plant...... xylanase inhibitors. To gain insight into the importance of the microbial consortia and their interaction with barley grains, we used a combined gel-based (2-DE coupled to MALDI-TOF-TOF MS) and gel-free (LC–MS/MS) proteomics approach complemented with enzyme activity assays to profile the surface......-associated proteins and xylanolytic activities of two barley cultivars. The surface-associated proteome was dominated by plant proteins with roles in defense and stress-responses, while the relatively less abundant microbial (bacterial and fungal) proteins were involved in cell-wall and polysaccharide degradation...

The cardiokine story unfolds: ischemic stress-induced protein secretion in the heart.

Science.gov (United States)

Doroudgar, Shirin; Glembotski, Christopher C

2011-04-01

Intercellular communication depends on many factors, including proteins released via the classical or non-classical secretory pathways, many of which must be properly folded to be functional. Owing to their adverse effects on the secretion machinery, stresses such as ischemia can impair the folding of secreted proteins. Paradoxically, cells rely on secreted proteins to mount a response designed to resist stress-induced damage. This review examines this paradox using proteins secreted from the heart, cardiokines, as examples, and focuses on how the ischemic heart maintains or even increases the release of select cardiokines that regulate important cellular processes in the heart, including excitation-contraction coupling, hypertrophic growth, myocardial remodeling and stem cell function, in ways that moderate ischemic damage and enhance cardiac repair. Copyright © 2010 Elsevier Ltd. All rights reserved.

Multi-frequencies ECT algorithms to remove sodium noise in ISI of ferromagnetic SG tubes of FBR

International Nuclear Information System (INIS)

Mihalache, Ovidiu

2012-01-01

The paper presents developments and application of multi-frequency eddy current to be used during In-Service Inspection (ISI) of ferromagnetic steam generator (SG) tubes of Fast Breeder Reactors (FBR). Signal enhancement by means of multi-frequency ECT techniques are validated through 3D simulations of both signals and noise due to sodium forms around SG tube or SP. The purpose of such algorithms is to remove from ECT signal the electromagnetic noise resulting from sodium accumulated outside of SG tubes after SG vessel draining. Finite element method (FEM) simulations are used to analyse different sodium build-up scenarios observed experimentally, and to determine optimal multi-frequency ECT algorithms to suppress the most efficiently sodium noise. Also a new 'window multi-frequency' algorithm is applied and validated using 3-dimensional FEM simulations of SP and sodium forms. (author)

Proteome-Level Analysis of Metabolism- and Stress-Related Proteins during Seed Dormancy and Germination in Gnetum parvifolium.

Science.gov (United States)

Chang, Ermei; Deng, Nan; Zhang, Jin; Liu, Jianfeng; Chen, Lanzhen; Zhao, Xiulian; Abbas, M; Jiang, Zeping; Shi, Shengqing

2018-03-21

Gnetum parvifolium is a rich source of materials for traditional medicines, food, and oil, but little is known about the mechanism underlying its seed dormancy and germination. In this study, we analyzed the proteome-level changes in its seeds during germination using isobaric tags for relative and absolute quantitation. In total, 1,040 differentially expressed proteins were identified, and cluster analysis revealed the distinct time points during which signal transduction and oxidation-reduction activity changed. Gene Ontology analysis showed that "carbohydrate metabolic process" and "response to oxidative stress" were the main enriched terms. Proteins associated with starch degradation and antioxidant enzymes were important for dormancy-release, while proteins associated with energy metabolism and protein synthesis were up-regulated during germination. Moreover, protein-interaction networks were mainly associated with heat-shock proteins. Furthermore, in accord with changes in the energy metabolism- and antioxidant-related proteins, indole-3-acetic acid, Peroxidase, and soluble sugar content increased, and the starch content decreased in almost all six stages of dormancy and germination analyzed (S1-S6). The activity of superoxide dismutase, abscisic acid, and malondialdehyde content increased in the dormancy stages (S1-S3) and then decreased in the germination stages (S4-S6). Our results provide new insights into G. parvifolium seed dormancy and germination at the proteome and physiological levels, with implications for improving seed propagation.

Severe Burn-Induced Intestinal Epithelial Barrier Dysfunction Is Associated With Endoplasmic Reticulum Stress and Autophagy in Mice

Science.gov (United States)

Huang, Yalan; Feng, Yanhai; Wang, Yu; Wang, Pei; Wang, Fengjun; Ren, Hui

2018-01-01

The disruption of intestinal barrier plays a vital role in the pathophysiological changes after severe burn injury, however, the underlying mechanisms are poorly understood. Severe burn causes the disruption of intestinal tight junction (TJ) barrier. Previous studies have shown that endoplasmic reticulum (ER) stress and autophagy are closely associated with the impairment of intestinal mucosa. Thus, we hypothesize that ER stress and autophagy are likely involved in burn injury-induced intestinal epithelial barrier dysfunction. Mice received a 30% total body surface area (TBSA) full-thickness burn, and were sacrificed at 0, 1, 2, 6, 12 and 24 h postburn. The results showed that intestinal permeability was increased significantly after burn injury, accompanied by the damage of mucosa and the alteration of TJ proteins. Severe burn induced ER stress, as indicated by increased intraluminal chaperone binding protein (BIP), CCAAT/enhancer-binding protein homologous protein (CHOP) and inositol-requiring enzyme 1(IRE1)/X-box binding protein 1 splicing (XBP1). Autophagy was activated after burn injury, as evidenced by the increase of autophagy related protein 5 (ATG5), Beclin 1 and LC3II/LC3I ratio and the decrease of p62. Besides, the number of autophagosomes was also increased after burn injury. The levels of p-PI3K(Ser191), p-PI3K(Ser262), p-AKT(Ser473), and p-mTOR were decreased postburn, suggesting that autophagy-related PI3K/AKT/mTOR pathway is involved in the intestinal epithelial barrier dysfunction following severe burn. In summary, severe burn injury induces the ER stress and autophagy in intestinal epithelia, leading to the disruption of intestinal barrier. PMID:29740349

Redox stress proteins are involved in adaptation response of the hyperthermoacidophilic archaeon Sulfolobus solfataricus to nickel challenge

Directory of Open Access Journals (Sweden)

Scaloni Andrea

2007-08-01

Full Text Available Abstract Background Exposure to nickel (Ni and its chemical derivatives has been associated with severe health effects in human. On the contrary, poor knowledge has been acquired on target physiological processes or molecular mechanisms of this metal in model organisms, including Bacteria and Archaea. In this study, we describe an analysis focused at identifying proteins involved in the recovery of the archaeon Sulfolobus solfataricus strain MT4 from Ni-induced stress. Results To this purpose, Sulfolobus solfataricus was grown in the presence of the highest nickel sulphate concentration still allowing cells to survive; crude extracts from treated and untreated cells were compared at the proteome level by using a bi-dimensional chromatography approach. We identified several proteins specifically repressed or induced as result of Ni treatment. Observed up-regulated proteins were largely endowed with the ability to trigger recovery from oxidative and osmotic stress in other biological systems. It is noteworthy that most of the proteins induced following Ni treatment perform similar functions and a few have eukaryal homologue counterparts. Conclusion These findings suggest a series of preferential gene expression pathways activated in adaptation response to metal challenge.

Studies on protein turnover and energy expenditure in chronically undernourished adults during stress of infection

International Nuclear Information System (INIS)

Kurpad, A.V.; Shetty, P.S.; Reeds, P.J.

1994-01-01

Chronic undernutrition in man leads to adaptive responses which could reduce the requirements for dietary energy and protein. It is also possible that these adaptive responses, which are economical in nature, could lead to a decreased capacity for combating stress. Undernourished people are more susceptible to infections, and during these stresses, show different patterns of protein and energy metabolism from well-nourished subjects. Animal models have clearly shown a diminished response to tissue injury, in terms of the anabolic acute phase response. It is proposed to study the effect of prior nutritional status on the degree to which an infective stress stimulates the acute phase protein synthesis by the liver. In addition, the supply of amino acids to the liver in conditions of stress could come from the breakdown of body tissue proteins, particularly muscle. It is intended to study muscle protein turnover by the use of 13 C-leucine in undernourished subjects under conditions of stress. Since whole body protein turnover can be measured by two methods, using 15 N-glycine and 13 C-leucine, a comparison of these two methods will initially be made in chronically undernourished subjects. It is also intended to study daily energy expenditure in the subject by an isotopic method, i.e. the appearance of 13 CO 2 in the breath after the administration of 13 C-bicarbonate. (author). 8 refs

Associations between circadian and stress response cortisol in children.

Science.gov (United States)

Simons, Sterre S H; Cillessen, Antonius H N; de Weerth, Carolina

2017-01-01

Hypothalamic-pituitary-adrenal (HPA) axis functioning is characterized by the baseline production of cortisol following a circadian rhythm, as well as by the superimposed production of cortisol in response to a stressor. However, it is relatively unknown whether the basal cortisol circadian rhythm is associated with the cortisol stress response in children. Since alterations in cortisol stress responses have been associated with mental and physical health, this study investigated whether the cortisol circadian rhythm is associated with cortisol stress responses in 6-year-old children. To this end, 149 normally developing children (M age  = 6.09 years; 70 girls) participated in an innovative social evaluative stress test that effectively provoked increases in cortisol. To determine the cortisol stress response, six cortisol saliva samples were collected and two cortisol stress response indices were calculated: total stress cortisol and cortisol stress reactivity. To determine children's cortisol circadian rhythm eight cortisol circadian samples were collected during two days. Total diurnal cortisol and diurnal cortisol decline scores were calculated as indices of the cortisol circadian rhythm. Hierarchical regression analyses indicated that higher total diurnal cortisol as well as a smaller diurnal cortisol decline, were both uniquely associated with higher total stress cortisol. No associations were found between the cortisol circadian rhythm indices and cortisol stress reactivity. Possible explanations for the patterns found are links with children's self-regulatory capacities and parenting quality.

Cytoskeleton-interacting LIM-domain protein CRP1 suppresses cell proliferation and protects from stress-induced cell death

International Nuclear Information System (INIS)

Latonen, Leena; Jaervinen, Paeivi M.; Laiho, Marikki

2008-01-01

Members of the cysteine-rich protein (CRP) family are actin cytoskeleton-interacting LIM-domain proteins known to act in muscle cell differentiation. We have earlier found that CRP1, a founding member of this family, is transcriptionally induced by UV radiation in human diploid fibroblasts [M. Gentile, L. Latonen, M. Laiho, Cell cycle arrest and apoptosis provoked by UV radiation-induced DNA damage are transcriptionally highly divergent responses, Nucleic Acids Res. 31 (2003) 4779-4790]. Here we show that CRP1 is induced by growth-inhibitory signals, such as increased cellular density, and cytotoxic stress induced by UV radiation or staurosporine. We found that high levels of CRP1 correlate with differentiation-associated morphology towards the myofibroblast lineage and that expression of ectopic CRP1 suppresses cell proliferation. Following UV- and staurosporine-induced stresses, expression of CRP1 provides a survival advantage evidenced by decreased cellular death and increased cellular metabolic activity and attachment. Our studies identify that CRP1 is a novel stress response factor, and provide evidence for its growth-inhibitory and cytoprotective functions

Protein complexes associated with the Kaposi's sarcoma-associated herpesvirus-encoded LANA

International Nuclear Information System (INIS)

Kaul, Rajeev; Verma, Subhash C.; Robertson, Erle S.

2007-01-01

Kaposi's sarcoma-associated herpesvirus (KSHV) is the major biological cofactor contributing to development of Kaposi's sarcoma. KSHV establishes a latent infection in human B cells expressing the latency-associated nuclear antigen (LANA), a critical factor in the regulation of viral latency. LANA is known to modulate viral and cellular gene expression. We report here on some initial proteomic studies to identify cellular proteins associated with the amino and carboxy-terminal domains of LANA. The results of these studies show an association of known cellular proteins which support LANA functions and have identified additional LANA-associated proteins. These results provide new evidence for complexes involving LANA with a number of previously unreported functional classes of proteins including DNA polymerase, RNA helicase and cell cycle control proteins. The results also indicate that the amino terminus of LANA can interact with its carboxy-terminal domain. This interaction is potentially important for facilitating associations with other cell cycle regulatory proteins which include CENP-F identified in association with both the amino and carboxy-termini. These novel associations add to the diversity of LANA functions in relation to the maintenance of latency and subsequent transformation of KSHV infected cells

Development of a light weighted mobile robot for SG tube inspection in NPP

International Nuclear Information System (INIS)

Seo, Yong Chil; Jeong, Kyung Min; Shin, Hochul; Gweng, Jung Ju; Lee, Sung Uk; Jeong, Seung Ho; Choi, Young Soo; Kim, Seung Ho; Shin, Chun Sup; Park, Ki Tae

2012-01-01

Steam generators (SG) are among the most critical components of pressurized water Nuclear Power Plants (NPP). SG tubes must provide a reliable pressure boundary between the primary and secondary cooling water, because any leakage from tube defects could result in the release of radioactivity to the environment. Thus degradations of steam generators tubes should be monitored and inspected periodically under nuclear regulation. In service inspections of SG tubes are carried out using eddy current test (ECT) and the defected tubes are usually plugged. Because the radioactivity in the internal SG chambers limits free access of human workers, remote manipulators are required. In South Korea, Manipulators such as the Zet ec SM series and the Westinghouse ROSA series have bee used. Such manipulators are rigidly mounted to man ways or tube sheets of SG. Confusions of the inspected tubes may occur from deflection of the manipulators. To reduce the deflections of the manipulators for covering the large working areas of tube sheets, sufficient rigidity is required and that leads to an increase of the weight. Such weight increase results in some difficulties for handling and more radiation exposure of human workers. Recently light weighed mobile robots have been introduced by Westinghouse and Zet ec. The robots can move keeping in contact with the tube sheets using devices which are commonly called cam locks. They are easier to handle and provide no confusion for the position of the inspected tubes. But when the clamping forces are loosed accidentally, they can be fall down and light repair works can be performed. This paper provides the design results for a lightweight mobile robot which is being developed in cooperation of our institutes

Development of a light weighted mobile robot for SG tube inspection in NPP

Energy Technology Data Exchange (ETDEWEB)

Seo, Yong Chil; Jeong, Kyung Min; Shin, Hochul; Gweng, Jung Ju; Lee, Sung Uk; Jeong, Seung Ho; Choi, Young Soo; Kim, Seung Ho [KAERI, Daejeon (Korea, Republic of); Shin, Chun Sup; Park, Ki Tae [Korea Plant Service and Engineering, Busan (Korea, Republic of)

2012-10-15

Steam generators (SG) are among the most critical components of pressurized water Nuclear Power Plants (NPP). SG tubes must provide a reliable pressure boundary between the primary and secondary cooling water, because any leakage from tube defects could result in the release of radioactivity to the environment. Thus degradations of steam generators tubes should be monitored and inspected periodically under nuclear regulation. In service inspections of SG tubes are carried out using eddy current test (ECT) and the defected tubes are usually plugged. Because the radioactivity in the internal SG chambers limits free access of human workers, remote manipulators are required. In South Korea, Manipulators such as the Zet ec SM series and the Westinghouse ROSA series have bee used. Such manipulators are rigidly mounted to man ways or tube sheets of SG. Confusions of the inspected tubes may occur from deflection of the manipulators. To reduce the deflections of the manipulators for covering the large working areas of tube sheets, sufficient rigidity is required and that leads to an increase of the weight. Such weight increase results in some difficulties for handling and more radiation exposure of human workers. Recently light weighed mobile robots have been introduced by Westinghouse and Zet ec. The robots can move keeping in contact with the tube sheets using devices which are commonly called cam locks. They are easier to handle and provide no confusion for the position of the inspected tubes. But when the clamping forces are loosed accidentally, they can be fall down and light repair works can be performed. This paper provides the design results for a lightweight mobile robot which is being developed in cooperation of our institutes.

Rubella virus capsid protein modulation of viral genomic and subgenomic RNA synthesis

International Nuclear Information System (INIS)

Tzeng, W.-P.; Frey, Teryl K.

2005-01-01

The ratio of the subgenomic (SG) to genome RNA synthesized by rubella virus (RUB) replicons expressing the green fluorescent protein reporter gene (RUBrep/GFP) is substantially higher than the ratio of these species synthesized by RUB (4.3 for RUBrep/GFP vs. 1.3-1.4 for RUB). It was hypothesized that this modulation of the viral RNA synthesis was by one of the virus structural protein genes and it was found that introduction of the capsid (C) protein gene into the replicons as an in-frame fusion with GFP resulted in an increase of genomic RNA production (reducing the SG/genome RNA ratio), confirming the hypothesis and showing that the C gene was the moiety responsible for the modulation effect. The N-terminal one-third of the C gene was required for the effect of be exhibited. A similar phenomenon was not observed with the replicons of Sindbis virus, a related Alphavirus. Interestingly, modulation was not observed when RUBrep/GFP was co-transfected with either other RUBrep or plasmid constructs expressing the C gene, demonstrating that modulation could occur only when the C gene was provided in cis. Mutations that prevented translation of the C protein failed to modulate RNA synthesis, indicating that the C protein was the moiety responsible for modulation; consistent with this conclusion, modulation of RNA synthesis was maintained when synonymous codon mutations were introduced at the 5' end of the C gene that changed the C gene sequence without altering the amino acid sequence of the C protein. These results indicate that C protein translated in proximity of viral replication complexes, possibly from newly synthesized SG RNA, participate in regulating the replication of viral RNA

Homeodomain-Interacting Protein Kinase (HPK-1) regulates stress responses and ageing in C. elegans.

Science.gov (United States)

Berber, Slavica; Wood, Mallory; Llamosas, Estelle; Thaivalappil, Priya; Lee, Karen; Liao, Bing Mana; Chew, Yee Lian; Rhodes, Aaron; Yucel, Duygu; Crossley, Merlin; Nicholas, Hannah R

2016-01-21

Proteins of the Homeodomain-Interacting Protein Kinase (HIPK) family regulate an array of processes in mammalian systems, such as the DNA damage response, cellular proliferation and apoptosis. The nematode Caenorhabditis elegans has a single HIPK homologue called HPK-1. Previous studies have implicated HPK-1 in longevity control and suggested that this protein may be regulated in a stress-dependent manner. Here we set out to expand these observations by investigating the role of HPK-1 in longevity and in the response to heat and oxidative stress. We find that levels of HPK-1 are regulated by heat stress, and that HPK-1 contributes to survival following heat or oxidative stress. Additionally, we show that HPK-1 is required for normal longevity, with loss of HPK-1 function leading to a faster decline of physiological processes that reflect premature ageing. Through microarray analysis, we have found that HPK-1-regulated genes include those encoding proteins that serve important functions in stress responses such as Phase I and Phase II detoxification enzymes. Consistent with a role in longevity assurance, HPK-1 also regulates the expression of age-regulated genes. Lastly, we show that HPK-1 functions in the same pathway as DAF-16 to regulate longevity and reveal a new role for HPK-1 in development.

Wheat F-Box Protein Gene TaFBA1 Is Involved in Plant Tolerance to Heat Stress

Directory of Open Access Journals (Sweden)

Qinxue Li

2018-04-01

Full Text Available Adverse environmental conditions, including high temperature, often affect the growth and production of crops worldwide. F-box protein, a core component of the Skp1-Cullin-F-box (SCF E3 ligase complex, plays an important role in abiotic stress responses. A previously cloned gene from wheat, TaFBA1, encodes a homologous F-box protein. A Yeast two-Hybrid (Y2H assay showed that TaFBA1 interacted with other SCF proteins. We found that the expression of TaFBA1 could be induced by heat stress (45°C. Overexpression of TaFBA1 enhanced heat stress tolerance in transgenic tobacco, because growth inhibition was reduced and photosynthesis increased as compared with those in the wild type (WT plants. Furthermore, the accumulation of H2O2, O2-, and carbonyl protein decreased and cell damage was alleviated in transgenic plants under heat stress, which resulted in less oxidative damage. However, the transgenic plants contained more enzymatic antioxidants after heat stress, which might be related to the regulation of some antioxidant gene expressions. The qRT-PCR analysis showed that the overexpression of TaFBA1 upregulated the expression of genes involved in reactive oxygen species (ROS scavenging, proline biosynthesis, and abiotic stress responses. We identified the interaction of TaFBA1 with Triticum aestivum stress responsive protein 1 (TaASRP1 by Y2H assay and bimolecular fluorescence complementation (BiFC assay. The results suggested that TaFBA1 may improve enzymatic antioxidant levels and regulate gene expression by interacting with other proteins, such as TaASRP1, which leads to the enhanced heat stress tolerance seen in the transgenic plants.

Methodology for failure assessment of SMART SG tube with once-through helical-coiled type

International Nuclear Information System (INIS)

Kim, Young Jin; Choi, Shin Beom; Cho, Doo Ho; Chang, Yoon Suk

2010-09-01

In this research project, existing integrity evaluation method for SMART steam generator tube with crack-like flaw was reviewed to determine subject analysis model and investigate possibility of failure under crack closure behavior. Furthermore, failure pressure estimation was proposed for SMART steam generator tubes containing wear-type defects. For each subject, the following issues are addressed: 1. Determination of subject analysis model for SMART SG tube contaning crack-like flaw 2. Applicability review on existing integrity evaluation method and investigation of failure possibility for SMART SG tube containing crack-like flaw 3. Development of failure pressure estimation model for SMART SG tube with wear type defect It is anticipated that if the technologies developed in this study are applied, structural integrity can be estimated accurately

The tomato DWD motif-containing protein DDI1 interacts with the CUL4–DDB1-based ubiquitin ligase and plays a pivotal role in abiotic stress responses

Energy Technology Data Exchange (ETDEWEB)

Miao, Min [Ministry of Education Key Laboratory for Bio-resource and Eco-environment, College of Life Science, State Key Laboratory of Hydraulics and Mountain River Engineering, Sichuan University, Chengdu 610064 (China); School of Biotechnology and Food Engineering, Hefei University of Technology, Hefei 230009 (China); Department of Plant, Soil and Entomological Sciences, University of Idaho, Moscow, ID 83844-2339 (United States); Zhu, Yunye [School of Biotechnology and Food Engineering, Hefei University of Technology, Hefei 230009 (China); Qiao, Maiju [Ministry of Education Key Laboratory for Bio-resource and Eco-environment, College of Life Science, State Key Laboratory of Hydraulics and Mountain River Engineering, Sichuan University, Chengdu 610064 (China); Tang, Xiaofeng [Ministry of Education Key Laboratory for Bio-resource and Eco-environment, College of Life Science, State Key Laboratory of Hydraulics and Mountain River Engineering, Sichuan University, Chengdu 610064 (China); School of Biotechnology and Food Engineering, Hefei University of Technology, Hefei 230009 (China); Zhao, Wei [School of Biotechnology and Food Engineering, Hefei University of Technology, Hefei 230009 (China); Xiao, Fangming [Department of Plant, Soil and Entomological Sciences, University of Idaho, Moscow, ID 83844-2339 (United States); Liu, Yongsheng, E-mail: liuyongsheng1122@hfut.edu.cn [Ministry of Education Key Laboratory for Bio-resource and Eco-environment, College of Life Science, State Key Laboratory of Hydraulics and Mountain River Engineering, Sichuan University, Chengdu 610064 (China); School of Biotechnology and Food Engineering, Hefei University of Technology, Hefei 230009 (China)

2014-08-08

Highlights: • We identify DDI1 as a DAMAGED DNA BINDING PROTEIN1 (DDB1)-interacting protein. • DDI1 interacts with the CUL4–DDB1-based ubiquitin ligase in the nucleus. • DDI1 plays a positive role in regulating abiotic stress response in tomato. - Abstract: CULLIN4(CUL4)–DAMAGED DNA BINDING PROTEIN1 (DDB1)-based ubiquitin ligase plays significant roles in multiple physiological processes via ubiquitination-mediated degradation of relevant target proteins. The DDB1–CUL4-associated factor (DCAF) acts as substrate receptor in the CUL4–DDB1 ubiquitin ligase complex and determines substrate specificity. In this study, we identified a tomato (Solanum lycopersicum) DDB1-interacting (DDI1) protein as a DCAF protein involved in response to abiotic stresses, including UV radiation, high salinity and osmotic stress. Co-immunoprecipitation and bimolecular fluorescence complementation assay indicated that DDI1 associates with CUL4–DDB1 in the nucleus. Quantitative RT-PCR analysis indicated the DDI1 gene is induced by salt, mannitol and UV-C treatment. Moreover, transgenic tomato plants with overexpression or knockdown of the DDI1 gene exhibited enhanced or attenuated tolerance to salt/mannitol/UV-C, respectively. Thus, our data suggest that DDI1 functions as a substrate receptor of the CUL4–DDB1 ubiquitin ligase, positively regulating abiotic stress response in tomato.

The tomato DWD motif-containing protein DDI1 interacts with the CUL4–DDB1-based ubiquitin ligase and plays a pivotal role in abiotic stress responses

International Nuclear Information System (INIS)

Miao, Min; Zhu, Yunye; Qiao, Maiju; Tang, Xiaofeng; Zhao, Wei; Xiao, Fangming; Liu, Yongsheng

2014-01-01

Highlights: • We identify DDI1 as a DAMAGED DNA BINDING PROTEIN1 (DDB1)-interacting protein. • DDI1 interacts with the CUL4–DDB1-based ubiquitin ligase in the nucleus. • DDI1 plays a positive role in regulating abiotic stress response in tomato. - Abstract: CULLIN4(CUL4)–DAMAGED DNA BINDING PROTEIN1 (DDB1)-based ubiquitin ligase plays significant roles in multiple physiological processes via ubiquitination-mediated degradation of relevant target proteins. The DDB1–CUL4-associated factor (DCAF) acts as substrate receptor in the CUL4–DDB1 ubiquitin ligase complex and determines substrate specificity. In this study, we identified a tomato (Solanum lycopersicum) DDB1-interacting (DDI1) protein as a DCAF protein involved in response to abiotic stresses, including UV radiation, high salinity and osmotic stress. Co-immunoprecipitation and bimolecular fluorescence complementation assay indicated that DDI1 associates with CUL4–DDB1 in the nucleus. Quantitative RT-PCR analysis indicated the DDI1 gene is induced by salt, mannitol and UV-C treatment. Moreover, transgenic tomato plants with overexpression or knockdown of the DDI1 gene exhibited enhanced or attenuated tolerance to salt/mannitol/UV-C, respectively. Thus, our data suggest that DDI1 functions as a substrate receptor of the CUL4–DDB1 ubiquitin ligase, positively regulating abiotic stress response in tomato

Water and Salt Stresses, Kinetin and Protein Synthesis in Tobacco Leaves 1

Science.gov (United States)

Ben-Zioni, Aliza; Itai, C.; Vaadia, Y.

1967-01-01

The capacity of tobacco (Nicotiana rustica) leaf discs to incorporate l-leucine 14C into proteins was measured. Leaf discs were obtained from plants which experienced soil water depletion, or which were exposed to a saline or osmotic stress in the root medium. The stresses were brief of relatively short duration and water potential did not decrease below 4 bars in the root media. Leaf discs were sampled 2 hours after stress removal, achieved by reirrigation, or replacement of saline and osmotic solutions with normal nutrient solution. Plants were always turgid when leaves were sampled. All stressed tissues showed reduced capacity to incorporate l-leucine 14C into protein. The reduction was about 50% and could not be attributed either to reduced uptake into the discs, or to possible isotopic dilution. Incorporation decreased progressively with leaf age in control discs as well as in stressed leaf discs. At all ages tested, incorporation in stressed discs was lower than that of the control. Full recovery of incorporation capacity in stressed discs was obtained when discs were sampled 72 hours after stress removal but not earlier. Kinetin pretreatment prior to incubation with labelled leucine partially restored incorporation in stressed discs. The differences in response to kinetin of stressed and control discs suggest a lower endogenous level of cytokinins in the stressed discs. The results were qualitatively similar regardless of the kind of stress given to the plants during pretreatment. This supports the hypothesis that the normal supply of root cytokinins is important in shoot metabolism. PMID:16656515

The transcription elongation factor Bur1-Bur2 interacts with replication protein A and maintains genome stability during replication stress

DEFF Research Database (Denmark)

Clausing, Emanuel; Mayer, Andreas; Chanarat, Sittinan

2010-01-01

Multiple DNA-associated processes such as DNA repair, replication, and recombination are crucial for the maintenance of genome integrity. Here, we show a novel interaction between the transcription elongation factor Bur1-Bur2 and replication protein A (RPA), the eukaryotic single-stranded DNA......-binding protein with functions in DNA repair, recombination, and replication. Bur1 interacted via its C-terminal domain with RPA, and bur1-¿C mutants showed a deregulated DNA damage response accompanied by increased sensitivity to DNA damage and replication stress as well as increased levels of persisting Rad52...... foci. Interestingly, the DNA damage sensitivity of an rfa1 mutant was suppressed by bur1 mutation, further underscoring a functional link between these two protein complexes. The transcription elongation factor Bur1-Bur2 interacts with RPA and maintains genome integrity during DNA replication stress....

Quantitative phosphoproteomics reveals the role of protein arginine phosphorylation in the bacterial stress response.

Science.gov (United States)

Schmidt, Andreas; Trentini, Débora Broch; Spiess, Silvia; Fuhrmann, Jakob; Ammerer, Gustav; Mechtler, Karl; Clausen, Tim

2014-02-01

Arginine phosphorylation is an emerging protein modification implicated in the general stress response of Gram-positive bacteria. The modification is mediated by the arginine kinase McsB, which phosphorylates and inactivates the heat shock repressor CtsR. In this study, we developed a mass spectrometric approach accounting for the peculiar chemical properties of phosphoarginine. The improved methodology was used to analyze the dynamic changes in the Bacillus subtilis arginine phosphoproteome in response to different stress situations. Quantitative analysis showed that a B. subtilis mutant lacking the YwlE arginine phosphatase accumulated a strikingly large number of arginine phosphorylations (217 sites in 134 proteins), however only a minor fraction of these sites was increasingly modified during heat shock or oxidative stress. The main targets of McsB-mediated arginine phosphorylation comprise central factors of the stress response system including the CtsR and HrcA heat shock repressors, as well as major components of the protein quality control system such as the ClpCP protease and the GroEL chaperonine. These findings highlight the impact of arginine phosphorylation in orchestrating the bacterial stress response.

Selection of number of SG modules per secondary sodium loop for a Prototype SFR

International Nuclear Information System (INIS)

Kim, Eui Kwang; Yoo, Jae-Woon; Joo, Hyung-Kook; Lee, Tae-Ho

2014-01-01

Steam generator is a vertical counter flow shell and tube heat exchanger with sodium on shell side and water-steam in tubes. The concept of steam generator modularization is discussed and the advantages of large and small steam generator modules are presented. To estimate the economics of the steam generator modularization for the PGSFR, detailed optimization studies were carried out for 1, 2, 3, 4 and 5 steam generator modules per loop, taking into consideration the effect of capital cost, operating cost, and outage cost. For the economics of the steam generator modularization for a PGSFR, detailed optimization studies were carried out for 1, 2, 3, 4 and 5 steam generator modules per loop. The effect of capital cost, operating cost, and outage cost were taken into consideration. The studies indicated that the design with less SG modules is found to be more economical, if the weld failure rate is below 2.0x1E-5 failure/year. From the economics of the modularization, thermal rate of SG, reliability, operability considerations, appropriate number of SG modules required for PGSFR will be 1 SG module per loop

Cmr1/WDR76 defines a nuclear genotoxic stress body linking genome integrity and protein quality control

DEFF Research Database (Denmark)

Gallina, Irene; Colding, Camilla Skettrup; Henriksen, Peter

2015-01-01

DNA replication stress is a source of genomic instability. Here we identify changed mutation rate 1 (Cmr1) as a factor involved in the response to DNA replication stress in Saccharomyces cerevisiae and show that Cmr1-together with Mrc1/Claspin, Pph3, the chaperonin containing TCP1 (CCT) and 25...... other proteins-define a novel intranuclear quality control compartment (INQ) that sequesters misfolded, ubiquitylated and sumoylated proteins in response to genotoxic stress. The diversity of proteins that localize to INQ indicates that other biological processes such as cell cycle progression...... propose that Cmr1/WDR76 plays a role in the recovery from genotoxic stress through regulation of the turnover of sumoylated and phosphorylated proteins....

A new protein-protein interaction sensor based on tripartite split-GFP association.

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Cabantous, Stéphanie; Nguyen, Hau B; Pedelacq, Jean-Denis; Koraïchi, Faten; Chaudhary, Anu; Ganguly, Kumkum; Lockard, Meghan A; Favre, Gilles; Terwilliger, Thomas C; Waldo, Geoffrey S

2013-10-04

Monitoring protein-protein interactions in living cells is key to unraveling their roles in numerous cellular processes and various diseases. Previously described split-GFP based sensors suffer from poor folding and/or self-assembly background fluorescence. Here, we have engineered a micro-tagging system to monitor protein-protein interactions in vivo and in vitro. The assay is based on tripartite association between two twenty amino-acids long GFP tags, GFP10 and GFP11, fused to interacting protein partners, and the complementary GFP1-9 detector. When proteins interact, GFP10 and GFP11 self-associate with GFP1-9 to reconstitute a functional GFP. Using coiled-coils and FRB/FKBP12 model systems we characterize the sensor in vitro and in Escherichia coli. We extend the studies to mammalian cells and examine the FK-506 inhibition of the rapamycin-induced association of FRB/FKBP12. The small size of these tags and their minimal effect on fusion protein behavior and solubility should enable new experiments for monitoring protein-protein association by fluorescence.

Removal of Secondary Side Deposit and Foreign Objects in SG of Yonggwang Nuclear Power Plant, Unit 2

Energy Technology Data Exchange (ETDEWEB)

Jeong, Wootae [KHNP-CRI, Daejeon (Korea, Republic of); Yun, Sangjung; Choi, Yongseok; Kim, Taehwa; Seo, Hongchang [Byucksan Digital Valley II, Seoul (Korea, Republic of)

2014-05-15

Mock-up test and training before the service were made to minimize operation time and radiation dose. 8 bag filters and 49 cartridge filters were consumed for filtering of soft and hard sludge in SG A, B, and C. Total 21.05 kilograms of sludge was removed. Five foreign objects were found and removed in tube bundle of SG A, in annulus and tube bundle of SG B, and in tube bundle of SG C. During the 20{sup th} OH of Yonggwang NPP unit 2, we removed 21.05 kilograms of sludge and five foreign objects from three steam generators. Gradual increase in weight of sludge removed is assumed to show us that more sludge is created as operation time of steam generator increases. Low level chemical cleaning at 19{sup th} OH removed 517 kilograms of sludge which was dominant of all the sludge removal weight. We could assume, from this fact, that high level chemical cleaning could remove significant amount of sludge compared to mechanical lancing method. Five foreign objects which were removed from inside of SG showed us that more thorough inside cleaning and inspection is necessary during fabrication of steam generator.

Candidate hippocampal biomarkers of susceptibility and resilience to stress in a rat model of depression

DEFF Research Database (Denmark)

Henningsen, Kim; Palmfeldt, Johan; Christiansen, Sofie Friis

2012-01-01

-scale proteomics was used to map hippocampal protein alterations in different stress states. Membrane proteins were successfully captured by two-phase separation and peptide based proteomics. Using iTRAQ labeling coupled with mass spectrometry, more than 2000 proteins were quantified and 73 proteins were found......Susceptibility to stress plays a crucial role in the development of psychiatric disorders such as unipolar depression and post-traumatic stress disorder. In the present study the chronic mild stress rat model of depression was used to reveal stress-susceptible and stress-resilient rats. Large...... to be differentially expressed. Stress susceptibility was associated with increased expression of a sodium-channel protein (SCN9A) currently investigated as a potential antidepressant target. Differential protein profiling also indicated stress susceptibility to be associated with deficits in synaptic vesicle release...

Overexpression of a Pathogenesis-Related Protein 10 Enhances Biotic and Abiotic Stress Tolerance in Rice

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Jingni Wu

2016-12-01

Full Text Available Pathogenesis-related proteins play multiple roles in plant development and biotic and abiotic stress tolerance. Here, we characterize a rice defense related gene named “jasmonic acid inducible pathogenesis-related class 10” (JIOsPR10 to gain an insight into its functional properties. Semi-quantitative RT-PCR analysis showed up-regulation of JIOsPR10 under salt and drought stress conditions. Constitutive over-expression JIOsPR10 in rice promoted shoot and root development in transgenic plants, however, their productivity was unaltered. Further experiments exhibited that the transgenic plants showed reduced susceptibility to rice blast fungus, and enhanced salt and drought stress tolerance as compared to the wild type. A comparative proteomic profiling of wild type and transgenic plants showed that overexpression of JIOsPR10 led to the differential modulation of several proteins mainly related with oxidative stresses, carbohydrate metabolism, and plant defense. Taken together, our findings suggest that JIOsPR10 plays important roles in biotic and abiotic stresses tolerance probably by activation of stress related proteins.

[sgRNA design for the CRISPR/Cas9 system and evaluation of its off-target effects].

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Xie, Sheng-song; Zhang, Yi; Zhang, Li-sheng; Li, Guang-lei; Zhao, Chang-zhi; Ni, Pan; Zhao, Shu-hong

2015-11-01

The third generation of CRISPR/Cas9-mediated genome editing technology has been successfully applied to genome modification of various species including animals, plants and microorganisms. How to improve the efficiency of CRISPR/Cas9 genome editing and reduce its off-target effects has been extensively explored in this field. Using sgRNA (Small guide RNA) with high efficiency and specificity is one of the critical factors for successful genome editing. Several software have been developed for sgRNA design and/or off-target evaluation, which have advantages and disadvantages respectively. In this review, we summarize characters of 16 kinds online and standalone software for sgRNA design and/or off-target evaluation and conduct a comparative analysis of these different kinds of software through developing 38 evaluation indexes. We also summarize 11 experimental approaches for testing genome editing efficiency and off-target effects as well as how to screen highly efficient and specific sgRNA.

Emery-Dreifuss Muscular Dystrophy-Associated Mutant Forms of Lamin A Recruit the Stress Responsive Protein Ankrd2 into the Nucleus, Affecting the Cellular Response to Oxidative Stress

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Silvia Angori

2017-05-01

Full Text Available Background: Ankrd2 is a stress responsive protein mainly expressed in muscle cells. Upon the application of oxidative stress, Ankrd2 translocates into the nucleus where it regulates the activity of genes involved in cellular response to stress. Emery-Dreifuss Muscular Dystrophy 2 (EDMD2 is a muscular disorder caused by mutations of the gene encoding lamin A, LMNA. As well as many phenotypic abnormalities, EDMD2 muscle cells also feature a permanent basal stress state, the underlying molecular mechanisms of which are currently unclear. Methods: Experiments were performed in EDMD2-lamin A overexpressing cell lines and EDMD2-affected human myotubes. Oxidative stress was produced by H2O2 treatment. Co-immunoprecipitation, cellular subfractionation and immunofluorescence analysis were used to validate the relation between Ankrd2 and forms of lamin A; cellular sensibility to stress was monitored by the analysis of Reactive Oxygen Species (ROS release and cell viability. Results: Our data demonstrate that oxidative stress induces the formation of a complex between Ankrd2 and lamin A. However, EDMD2-lamin A mutants were able to bind and mislocalize Ankrd2 in the nucleus even under basal conditions. Nonetheless, cells co-expressing Ankrd2 and EDMD2-lamin A mutants were more sensitive to oxidative stress than the Ankrd2-wild type lamin A counterpart. Conclusions: For the first time, we present evidence that in muscle fibers from patients affected by EDMD2, Ankrd2 has an unusual nuclear localization. By introducing a plausible mechanism ruling this accumulation, our data hint at a novel function of Ankrd2 in the pathogenesis of EDMD2-affected cells.

Context Specificity of Stress-activated Mitogen-activated Protein (MAP) Kinase Signaling: The Story as Told by Caenorhabditis elegans*

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Andrusiak, Matthew G.; Jin, Yishi

2016-01-01

Stress-associated p38 and JNK mitogen-activated protein (MAP) kinase signaling cascades trigger specific cellular responses and are involved in multiple disease states. At the root of MAP kinase signaling complexity is the differential use of common components on a context-specific basis. The roundworm Caenorhabditis elegans was developed as a system to study genes required for development and nervous system function. The powerful genetics of C. elegans in combination with molecular and cellular dissections has led to a greater understanding of how p38 and JNK signaling affects many biological processes under normal and stress conditions. This review focuses on the studies revealing context specificity of different stress-activated MAPK components in C. elegans. PMID:26907690

Genotypic variability in faba bean (vicia faba L.) for seed yield and protein content under drought stress during vegetative and Reproductive Stages

International Nuclear Information System (INIS)

Abdelmula, A. A.; Gasim, S. M.; Link, W.; Mohamed, A. A.; Khalifa, J. E.

2012-01-01

Faba bean (viciafaba L.) is subjected to drought stress during different growth stages. In this study, variability in seed yield and protein content was investigated when drought occurred during the vegetative and reproductive stages. Twenty two genotypes of faba bean were field evaluated under three levels of drought stress at two locations in the Sudan. The three levels of drought were normal watering (non-stress), drought during the vegetative stage and drought during the reproductive stage. Data were collected on yield and vegetative traits and protein content. The results showed that yield, as well as other traits, were reduced by drought. The genotypes exhibited significant differences for 100 seed weight, plant height and protein content. The interaction between the genotypes and drought was significant for yield/plant. Some genotypes were more sensitive when drought occurred during the vegetative stage, some when drought occurred during the vegetative stage, and others were more stable under the three levels of drought. yield/plant showed significant covariance with pods/plant and plant height. The association between different characters varied according to trait and the time of drought incidence. The correlation of yield/plant with protein content was negative under all drought levels, and the average correlation coefficient was 0.32. It could be concluded that the specific adaptation and the wide adaptation have great implication for improving faba bean under drought. To select for high seed yield under drought, secondary characters, such as pods/plant and plant height could be of great importance. Drought could reduce protein content and affect its association with yield/plant.(Author)

Acute psychophysiological stress impairs human associative learning.

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Ehlers, M R; Todd, R M

2017-11-01

Addiction is increasingly discussed asa disorder of associative learning processes, with both operant and classical conditioning contributing to the development of maladaptive habits. Stress has long been known to promote drug taking and relapse and has further been shown to shift behavior from goal-directed actions towards more habitual ones. However, it remains to be investigated how acute stress may influence simple associative learning processes that occur before a habit can be established. In the present study, healthy young adults were exposed to either acute stress or a control condition half an hour before performing simple classical and operant conditioning tasks. Psychophysiological measures confirmed successful stress induction. Results of the operant conditioning task revealed reduced instrumental responding under delayed acute stress that resembled behavioral responses to lower levels of reward. The classical conditioning experiment revealed successful conditioning in both experimental groups; however, explicit knowledge of conditioning as indicated by stimulus ratings differentiated the stress and control groups. These findings suggest that operant and classical conditioning are differentially influenced by the delayed effects of acute stress with important implications for the understanding of how new habitual behaviors are initially established. Copyright © 2017 Elsevier Inc. All rights reserved.

Limbic system associated membrane protein as a potential target for neuropsychiatric disorders

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Eero eVasar

2013-03-01

Full Text Available The studies performed in laboratory animals and psychiatric patients suggest a possible role of limbic system associated membrane protein (LAMP in the mechanisms of psychiatric disorders. Stressful manipulations and genetic invalidation have revealed a role of the Lsamp gene in the regulation of anxiety in rodents. Besides that, Lsamp deficient mice display reduced aggressiveness and impaired adaptation in novel and stressful environments. The behavioural effects of amphetamine were blunted in genetically modified mice. Recent pharmacological and biochemical studies point towards altered function of GABA-, 5-hydroxytryptamine- and dopaminergic systems in Lsamp deficient mice. Moreover, we found an association between the gene polymorphisms of LSAMP and major depressive disorder. Patients suffering from major depressive disorder had significantly increased ratio between risk and protective haplotypes of the LSAMP gene compared to healthy volunteers. However, the impact of these haplotypes for the function of LAMP is not clear and remains to be elucidated in future studies.

Inferring microbial interaction networks from metagenomic data using SgLV-EKF algorithm.

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Alshawaqfeh, Mustafa; Serpedin, Erchin; Younes, Ahmad Bani

2017-03-27

Inferring the microbial interaction networks (MINs) and modeling their dynamics are critical in understanding the mechanisms of the bacterial ecosystem and designing antibiotic and/or probiotic therapies. Recently, several approaches were proposed to infer MINs using the generalized Lotka-Volterra (gLV) model. Main drawbacks of these models include the fact that these models only consider the measurement noise without taking into consideration the uncertainties in the underlying dynamics. Furthermore, inferring the MIN is characterized by the limited number of observations and nonlinearity in the regulatory mechanisms. Therefore, novel estimation techniques are needed to address these challenges. This work proposes SgLV-EKF: a stochastic gLV model that adopts the extended Kalman filter (EKF) algorithm to model the MIN dynamics. In particular, SgLV-EKF employs a stochastic modeling of the MIN by adding a noise term to the dynamical model to compensate for modeling uncertainties. This stochastic modeling is more realistic than the conventional gLV model which assumes that the MIN dynamics are perfectly governed by the gLV equations. After specifying the stochastic model structure, we propose the EKF to estimate the MIN. SgLV-EKF was compared with two similarity-based algorithms, one algorithm from the integral-based family and two regression-based algorithms, in terms of the achieved performance on two synthetic data-sets and two real data-sets. The first data-set models the randomness in measurement data, whereas, the second data-set incorporates uncertainties in the underlying dynamics. The real data-sets are provided by a recent study pertaining to an antibiotic-mediated Clostridium difficile infection. The experimental results demonstrate that SgLV-EKF outperforms the alternative methods in terms of robustness to measurement noise, modeling errors, and tracking the dynamics of the MIN. Performance analysis demonstrates that the proposed SgLV-EKF algorithm

Cloning, characterization, and heat stress-induced redistribution of a protein homologous to human hsp27 in the zebrafish Danio rerio

International Nuclear Information System (INIS)

Mao Li; Bryantsev, Anton L.; Chechenova, Maria B.; Shelden, Eric A.

2005-01-01

Hsp27 is a small heat shock protein (shsp) regulating stress tolerance and increasingly thought to play roles in tissue homeostasis and differentiation. The zebrafish Danio rerio is an important model for the study of developmental processes, but little is known regarding shsps in this animal. Here, we report the sequence, expression, regulation, and function of a zebrafish protein (zfHsp27) homologous to human Hsp27. zfHsp27 contains three conserved phosphorylatable serines and a cysteine important for regulation of apoptosis, but it lacks much of a C-terminal tail domain and shows low homology in two putative actin interacting domains that are features of mammalian Hsp27. zfHsp27 mRNA is most abundant in adult skeletal muscle and heart and is upregulated during early embryogenesis. zfHsp27 expressed in mammalian fibroblasts was phosphorylated in response to heat stress and anisomycin, and this phosphorylation was prevented by treatment with SB202190, an inhibitor of p38 MAPK. Expression of zfHsp27 and human Hsp27 in mammalian fibroblasts promoted a similar degree of tolerance to heat stress. zfHsp27 fusion proteins entered the nucleus and associated with the cytoskeleton of heat stressed cells in vitro and in zebrafish embryos. These results reveal conservation in regulation and function of mammalian and teleost Hsp27 proteins and define zebrafish as a new model for the study of Hsp27 function

Organ-specific gene expression: the bHLH protein Sage provides tissue specificity to Drosophila FoxA.

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Fox, Rebecca M; Vaishnavi, Aria; Maruyama, Rika; Andrew, Deborah J

2013-05-01

FoxA transcription factors play major roles in organ-specific gene expression, regulating, for example, glucagon expression in the pancreas, GLUT2 expression in the liver, and tyrosine hydroxylase expression in dopaminergic neurons. Organ-specific gene regulation by FoxA proteins is achieved through cooperative regulation with a broad array of transcription factors with more limited expression domains. Fork head (Fkh), the sole Drosophila FoxA family member, is required for the development of multiple distinct organs, yet little is known regarding how Fkh regulates tissue-specific gene expression. Here, we characterize Sage, a bHLH transcription factor expressed exclusively in the Drosophila salivary gland (SG). We show that Sage is required for late SG survival and normal tube morphology. We find that many Sage targets, identified by microarray analysis, encode SG-specific secreted cargo, transmembrane proteins, and the enzymes that modify these proteins. We show that both Sage and Fkh are required for the expression of Sage target genes, and that co-expression of Sage and Fkh is sufficient to drive target gene expression in multiple cell types. Sage and Fkh drive expression of the bZip transcription factor Senseless (Sens), which boosts expression of Sage-Fkh targets, and Sage, Fkh and Sens colocalize on SG chromosomes. Importantly, expression of Sage-Fkh target genes appears to simply add to the tissue-specific gene expression programs already established in other cell types, and Sage and Fkh cannot alter the fate of most embryonic cell types even when expressed early and continuously.

Changes in protein expression in testes of L2 strain Taiwan country chickens in response to acute heat stress.

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Wang, Shih-Han; Cheng, Chuen-Yu; Chen, Chao-Jung; Chen, Hsin-Hsin; Tang, Pin-Chi; Chen, Chih-Feng; Lee, Yen-Pai; Huang, San-Yuan

2014-07-01

Heat stress causes a decrease of fertility in roosters. Yet, the way acute heat stress affects protein expression remains poorly understood. This study investigated differential protein expression in testes of the L2 strain of Taiwan country chickens following acute heat stress. Twelve 45-week-old roosters were allocated into four groups, including control roosters kept at 25 °C, roosters subjected to 38 °C acute heat stress for 4 hours without recovery, with 2 hours of recovery, and with 6 hours of recovery. Testis samples were collected for morphologic assay and protein analysis. Some of the differentially expressed proteins were validated by Western blot and immunohistochemistry. Abnormal and apoptotic spermatogenic cells were observed at 2 hours of recovery after acute heat stress, especially among the spermatocytes. Two-dimensional difference gel electrophoresis revealed that 119 protein spots were differentially expressed in chicken testes following heat stress, and peptide mass fingerprinting revealed that these spots contained 92 distinct proteins. In the heat-stressed samples, the heat shock proteins, chaperonin containing t-complex, and proteasome subunits were downregulated, and glutathione S-transferase, transgelin, and DJ-1 were upregulated. Our results demonstrate that acute heat stress impairs the processes of translation, protein folding, and protein degradation, and thus results in apoptosis and interferes with spermatogenesis. On the other hand, the increased expression of antioxidant enzymes, including glutathione S-transferase and DJ-1, may attenuate heat-induced damage. These findings may have implications for breeding chickens that can tolerate more extreme conditions. Copyright © 2014 Elsevier Inc. All rights reserved.

Identification and profiling of salinity stress-responsive proteins in Sorghum bicolor seedlings

DEFF Research Database (Denmark)

Ngara, Rudo; Ndimba, Roya; Borch-Jensen, Jonas

2012-01-01

Sorghum bicolor, a drought tolerant cereal crop, is not only an important food source in the semi arid/arid regions but also a potential model for studying and gaining a better understanding of the molecular mechanisms of drought and salt stress tolerance in cereals. In this study, seeds of a sweet...... sorghum variety, MN1618, were planted and grown on solid MS growth medium with or without 100mM NaCl. Heat shock protein expression immunoblotting assays demonstrated that this salt treatment induced stress within natural physiological parameters for our experimental material. 2D PAGE in combination...... with MS/MS proteomics techniques were used to separate, visualise and identify salinity stress responsive proteins in young sorghum leaves. Out of 281 Coomassie stainable spots, 118 showed statistically significant responses (p...

Mutations that alter a conserved element upstream of the potato virus X triple block and coat protein genes affect subgenomic RNA accumulation.

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Kim, K H; Hemenway, C

1997-05-26

The putative subgenomic RNA (sgRNA) promoter regions upstream of the potato virus X (PVX) triple block and coat protein (CP) genes contain sequences common to other potexviruses. The importance of these sequences to PVX sgRNA accumulation was determined by inoculation of Nicotiana tabacum NT1 cell suspension protoplasts with transcripts derived from wild-type and modified PVX cDNA clones. Analyses of RNA accumulation by S1 nuclease digestion and primer extension indicated that a conserved octanucleotide sequence element and the spacing between this element and the start-site for sgRNA synthesis are critical for accumulation of the two major sgRNA species. The impact of mutations on CP sgRNA levels was also reflected in the accumulation of CP. In contrast, genomic minus- and plus-strand RNA accumulation were not significantly affected by mutations in these regions. Studies involving inoculation of tobacco plants with the modified transcripts suggested that the conserved octanucleotide element functions in sgRNA accumulation and some other aspect of the infection process.

Cardiac protein expression patterns are associated with distinct inborn exercise capacity in non-selectively bred rats

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L.P. Ribeiro

2018-01-01

Full Text Available In the present study, we successfully demonstrated for the first time the existence of cardiac proteomic differences between non-selectively bred rats with distinct intrinsic exercise capacities. A proteomic approach based on two-dimensional gel electrophoresis coupled to mass spectrometry was used to study the left ventricle (LV tissue proteome of rats with distinct intrinsic exercise capacity. Low running performance (LRP and high running performance (HRP rats were categorized by a treadmill exercise test, according to distance run to exhaustion. The running capacity of HRPs was 3.5-fold greater than LRPs. Protein profiling revealed 29 differences between HRP and LRP rats (15 proteins were identified. We detected alterations in components involved in metabolism, antioxidant and stress response, microfibrillar and cytoskeletal proteins. Contractile proteins were upregulated in the LVs of HRP rats (α-myosin heavy chain-6, myosin light chain-1 and creatine kinase, whereas the LVs of LRP rats exhibited upregulation in proteins associated with stress response (aldehyde dehydrogenase 2, α-crystallin B chain and HSPβ-2. In addition, the cytoskeletal proteins desmin and α-actin were upregulated in LRPs. Taken together, our results suggest that the increased contractile protein levels in HRP rats partly accounted for their improved exercise capacity, and that proteins considered risk factors to the development of cardiovascular disease were expressed in higher amounts in LRP animals.

Aniline-induced nitrosative stress in rat spleen: Proteomic identification of nitrated proteins

International Nuclear Information System (INIS)

Fan Xiuzhen; Wang Jianling; Soman, Kizhake V.; Ansari, G.A.S.; Khan, M. Firoze

2011-01-01

Aniline exposure is associated with toxicity to the spleen which is characterized by splenomegaly, hyperplasia, fibrosis, and a variety of sarcomas on chronic exposure in rats. However, mechanisms by which aniline elicits splenotoxic responses are not well understood. Earlier we have shown that aniline exposure leads to increased nitration of proteins in the spleen. However, nitrated proteins remain to be characterized. Therefore, in the current study using proteomic approaches, we focused on characterizing the nitrated proteins in the spleen of aniline-exposed rats. Aniline exposure led to increased tyrosine nitration of proteins, as determined by 2D Western blotting with anti-3-nitrotyrosine specific antibody, compared to the controls. The analyzed nitrated proteins were found in the molecular weight range of 27.7 to 123.6 kDa. A total of 37 nitrated proteins were identified in aniline-treated and control spleens. Among them, 25 were found only in aniline-treated rats, 11 were present in both aniline-treated and control rats, while one was found in controls only. The nitrated proteins identified mainly represent skeletal proteins, chaperones, ferric iron transporter, enzymes, nucleic acids binding protein, and signaling and protein synthesis pathways. Furthermore, aniline exposure led to significantly increased iNOS mRNA and protein expression in the spleen, suggesting its role in increased reactive nitrogen species formation and contribution to increased nitrated proteins. The identified nitrated proteins provide a global map to further investigate alterations in their structural and functional properties, which will lead to a better understanding of the role of protein nitration in aniline-mediated splenic toxicity. - Highlights: → Proteomic approaches are used to identify nitrated proteins in the spleen. → Twenty five nitrated proteins were found only in the spleen of aniline-treated rats. → Aniline exposure led to increased iNOS mRNA and protein

Proteomics analysis of maize (Zea mays L.) grain based on iTRAQ reveals molecular mechanisms of poor grain filling in inferior grains.

Science.gov (United States)

Yu, Tao; Li, Geng; Liu, Peng; Dong, Shuting; Zhang, Jiwang; Zhao, Bin

2017-06-01

In maize, inferior grains (IG) located on the upper part of the ear have poor grain filling process compared to superior grains (SG) located on the middle and lower parts of the ear. This difference limits satisfactory yield and quality; however, the underlying molecular mechanisms remain unknown. Here, using the isobaric tag for relative and absolute quantification (iTRAQ) technology, the proteomes of IG and SG during early and middle grain filling stages were investigated. In total, 4720 proteins were identified in maize grain and 305 differentially accumulated proteins (DiAPs) were detected between IG and SG. These DiAPs were involved in diverse cellular and metabolic processes with preferred distribution in protein synthesis/destination and metabolism. Compared to SG, DiAPs related to cell growth/division and starch synthesis were lag-accumulated and down-regulated in IG, respectively, resulting in smaller sink sizes and lower sink activities in IG. Meanwhile, impediment of the glycolysis pathway in IG may lead to reduce energy supply and building materials for substance synthesis. Additionally, reactive oxygen species (ROS) homeostasis and the defense system were disturbed in IG, which might lead to reduce protection against various environmental stresses. The present study provides new information on the proteomic differences between IG and SG, and explains possible molecular mechanisms for poor grain filling in IG. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Effect of Phosphorus Fertilizer and Water Stress on Protein and Phenolic Contents in Cotton (Gossypium Hirsutum L.)

International Nuclear Information System (INIS)

Abbas, Z.; Muhammad, S.; Murtaza, G.; Ahmad, I.; Shakeel, A.; Islam, M.; Ahmad, M.; Abdullah, M.

2015-01-01

Crop quality and production are affected by various fertilizers and water stress. In present research, the response of cotton variety CIM-496 to water stress and phosphorus fertilizer was investigated. Samples were collected after 90 days of planting. Kjeldahl method and thin layer chromatography (TLC) were used for the quantitative and qualitative analysis of total protein and phenolic compounds, respectively. Proteins were greatly affected by fertilizer treatment and water stress, but phenolic compounds remained unchanged upon fertilizer treatment. However, they were greatly affected by irrigation and water stress. Crop treated with 100 kg ha/sup -1/ P/sub 2/O/sub 5/ under water stress maintained high protein content as compared to unfertilized and no water stress treatments. However, phenolic compounds were found higher in fully irrigated plants as compared to water stress ones. Fertilizer treatments had no considerable effect on phenolic compounds. (author)

Mutations of C19orf12, coding for a transmembrane glycine zipper containing mitochondrial protein, cause mis-localization of the protein, inability to respond to oxidative stress and increased mitochondrial Ca2+

DEFF Research Database (Denmark)

Venco, Paola; Bonora, Massimo; Giorgi, Carlotta

2015-01-01

19orf12 protein was not exclusively present in mitochondria, but also in the Endoplasmic Reticulum (ER) and MAM (Mitochondria Associated Membrane), while mutant C19orf12 variants presented a different localization. Moreover, after induction of oxidative stress, a GFP-tagged C19orf12 wild-type protein...... was able to relocate to the cytosol. On the contrary, mutant isoforms were not able to respond to oxidative stress. High mitochondrial calcium concentration and increased H2O2 induced apoptosis were found in fibroblasts derived from one patient as compared to controls. C19orf12 protein is a 17 k...... to rearrange in a structural domain, which is homologs to the N-terminal regulatory domain of the magnesium transporter MgtE, suggesting that C19orf12 may act as a regulatory protein for human MgtE transporters. The mutations here described affect respectively one glycine residue of the glycine zipper motifs...

Factors associated with protein consumption in elderly

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Natália GASPARETO

Full Text Available ABSTRACT Objective We evaluated factors associated with protein consumption by the elderly. Methods We performed a cross-sectional study in a sample of 295 elderly consumers of health facilities in São Caetano do Sul, São Paulo, Brazil. Protein consumption data (g and g/kg were obtained through 24-hour dietary recalls, which was reapplied in a 30% sub-sample to estimate habitual consumption, with an interval of two weeks. The association between protein consumption and sociodemographic, economic, health, and dietary variables was tested using multiple linear regression. Results There was a positive association between protein consumption (g and g/kg and better Brazilian Healthy Eating Index-Revised, between protein consumption (g and male sex, and a negative association between protein consumption (g/kg and greater calf circumference. Higher average protein consumption (g or g/kg was observed among married elderly, individuals with higher income and schooling, who were economically active, eutrophic, without dyslipidemia and symptoms of dysphagia, who consumed three main meals and an intermediate snack. Conclusion The results showed that protein consumption was associated with diet quality, sex, and calf circumference. The identification of elderly groups prone to protein inadequacy may direct individual and collective interventions to prevent muscle mass reduction and its implications, such as sarcopenia and other adverse outcomes.

Increased Expression of the Innate Immune Receptor TLR10 in Obesity and Type-2 Diabetes: Association with ROS-Mediated Oxidative Stress

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Sardar Sindhu

2018-01-01

Full Text Available Background/Aims: Metabolic diseases such as obesity and type-2 diabetes (T2D are known to be associated with chronic low-grade inflammation called metabolic inflammation together with an oxidative stress milieu found in the expanding adipose tissue. The innate immune Toll-like receptors (TLR such as TLR2 and TLR4 have emerged as key players in metabolic inflammation; nonetheless, TLR10 expression in the adipose tissue and its significance in obesity/T2D remain unclear. Methods: TLR10 gene expression was determined in the adipose tissue samples from healthy non-diabetic and T2D individuals, 13 each, using real-time RT-PCR. TLR10 protein expression was determined by immunohistochemistry, confocal microscopy, and flow cytometry. Regarding in vitro studies, THP-1 cells, peripheral blood mononuclear cells (PBMC, or primary monocytes were treated with hydrogen peroxide (H2O2 for induction of reactive oxygen species (ROS-mediated oxidative stress. Superoxide dismutase (SOD activity was measured using a commercial kit. Data (mean±SEM were compared using unpaired student’s t-test and P<0.05 was considered significant. Results: The adipose tissue TLR10 gene/protein expression was found to be significantly upregulated in obesity as well as T2D which correlated with body mass index (BMI. ROS-mediated oxidative stress induced high levels of TLR10 gene/protein expression in monocytic cells and PBMC. In these cells, oxidative stress induced a time-dependent increase in SOD activity. Pre-treatment of cells with anti-oxidants/ROS scavengers diminished the expression of TLR10. ROS-induced TLR10 expression involved the nuclear factor-kappaB (NF-κB/mitogen activated protein kinase (MAPK signaling as well as endoplasmic reticulum (ER stress. H2O2-induced oxidative stress interacted synergistically with palmitate to trigger the expression of TLR10 which associated with enhanced expression of proinflammatory cytokines/chemokine. Conclusion: Oxidative stress

Socioeconomic Status, Subjective Social Status, and Perceived Stress: Associations with Stress Physiology and Executive Functioning.

Science.gov (United States)

Ursache, Alexandra; Noble, Kimberly G; Blair, Clancy

2015-01-01

Several studies have investigated associations between socioeconomic status (SES) and indicators of children's physiological and cognitive self-regulation. Although objective measures of family SES may be good proxies for families' experiences of disadvantage, less is known about subjective aspects of families' experiences. We hypothesize that subjective social status (SSS) and perceived stress may be important independent predictors of children's stress physiology and executive functioning (EF). Eighty-two children from diverse SES backgrounds were administered EF measures and provided saliva samples for cortisol assay. Caregivers reported on objective SES, SSS, and perceived stress. Results suggest that SES and SSS are both independently and positively related to EF. In models predicting stress physiology, higher perceived stress was associated with lower baseline cortisol. Moreover, SES and age interacted to predict cortisol levels such that among younger children, lower SES was associated with higher cortisol, whereas among older children, lower SES was associated with lower cortisol. Results highlight the importance of considering both objective and subjective indicators of families' SES and stressful experiences in relation to multiple aspects of children's self-regulation.

Exposure to tributyltin induces endoplasmic reticulum stress and the unfolded protein response in zebrafish.

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Komoike, Yuta; Matsuoka, Masato

2013-10-15

Tributyltin (TBT) is a major marine contaminant and causes endocrine disruption, hepatotoxicity, immunotoxicity, and neurotoxicity. However, the molecular mechanisms underlying the toxicity of TBT have not been fully elucidated. We examined whether exposure to TBT induces the endoplasmic reticulum (ER) stress response in zebrafish, a model organism. Zebrafish-derived BRF41 fibroblast cells were exposed to 0.5 or 1 μM TBT for 0.5-16 h and subsequently lysed and immunoblotted to detect ER stress-related proteins. Zebrafish embryos, grown until 32 h post fertilization (hpf), were exposed to 1 μM TBT for 16 h and used in whole mount in situ hybridization and immunohistochemistry to visualize the expression of ER chaperones and an ER stress-related apoptosis factor. Exposure of the BRF41 cells to TBT caused phosphorylation of the zebrafish homolog of protein kinase RNA-activated-like ER kinase (PERK), eukaryotic translation initiation factor 2 alpha (eIF2α), and inositol-requiring enzyme 1 (IRE1), characteristic splicing of X-box binding protein 1 (XBP1) mRNA, and enhanced expression of activating transcription factor 4 (ATF4) protein. In TBT-exposed zebrafish embryos, ectopic expression of the gene encoding zebrafish homolog of the 78 kDa glucose-regulating protein (GRP78) and gene encoding CCAAT/enhancer-binding protein homologous protein (CHOP) was detected in the precursors of the neuromast, which is a sensory organ for detecting water flow and vibration. Our in vitro and in vivo studies revealed that exposure of zebrafish to TBT induces the ER stress response via activation of both the PERK-eIF2α and IRE1-XBP1 pathways of the unfolded protein response (UPR) in an organ-specific manner. Copyright © 2013 Elsevier B.V. All rights reserved.

Anionic lipids and the cytoskeletal proteins MreB and RodZ define the spatio-temporal distribution and function of membrane stress controller PspA in Escherichia coli.

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Jovanovic, Goran; Mehta, Parul; Ying, Liming; Buck, Martin

2014-11-01

All cell types must maintain the integrity of their membranes. The conserved bacterial membrane-associated protein PspA is a major effector acting upon extracytoplasmic stress and is implicated in protection of the inner membrane of pathogens, formation of biofilms and multi-drug-resistant persister cells. PspA and its homologues in Gram-positive bacteria and archaea protect the cell envelope whilst also supporting thylakoid biogenesis in cyanobacteria and higher plants. In enterobacteria, PspA is a dual function protein negatively regulating the Psp system in the absence of stress and acting as an effector of membrane integrity upon stress. We show that in Escherichia coli the low-order oligomeric PspA regulatory complex associates with cardiolipin-rich, curved polar inner membrane regions. There, cardiolipin and the flotillin 1 homologue YqiK support the PspBC sensors in transducing a membrane stress signal to the PspA-PspF inhibitory complex. After stress perception, PspA high-order oligomeric effector complexes initially assemble in polar membrane regions. Subsequently, the discrete spatial distribution and dynamics of PspA effector(s) in lateral membrane regions depend on the actin homologue MreB and the peptidoglycan machinery protein RodZ. The consequences of loss of cytoplasmic membrane anionic lipids, MreB, RodZ and/or YqiK suggest that the mode of action of the PspA effector is closely associated with cell envelope organization. © 2014 The Authors.

Overexpressed cyclophilin B suppresses apoptosis associated with ROS and Ca2+ homeostasis after ER stress.

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Kim, Jinhwan; Choi, Tae Gyu; Ding, Yan; Kim, Yeonghwan; Ha, Kwon Soo; Lee, Kyung Ho; Kang, Insug; Ha, Joohun; Kaufman, Randal J; Lee, Jinhwa; Choe, Wonchae; Kim, Sung Soo

2008-11-01

Prolonged accumulation of misfolded proteins in the endoplasmic reticulum (ER) results in ER stress-mediated apoptosis. Cyclophilins are protein chaperones that accelerate the rate of protein folding through their peptidyl-prolyl cis-trans isomerase (PPIase) activity. In this study, we demonstrated that ER stress activates the expression of the ER-localized cyclophilin B (CypB) gene through a novel ER stress response element. Overexpression of wild-type CypB attenuated ER stress-induced cell death, whereas overexpression of an isomerase activity-defective mutant, CypB/R62A, not only increased Ca(2+) leakage from the ER and ROS generation, but also decreased mitochondrial membrane potential, resulting in cell death following exposure to ER stress-inducing agents. siRNA-mediated inhibition of CypB expression rendered cells more vulnerable to ER stress. Finally, CypB interacted with the ER stress-related chaperones, Bip and Grp94. Taken together, we concluded that CypB performs a crucial function in protecting cells against ER stress via its PPIase activity.

Accumulation and localization of extensin protein in apoplast of pea root nodule under aluminum stress.

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Sujkowska-Rybkowska, Marzena; Borucki, Wojciech

2014-12-01

Cell wall components such as hydroxyproline-rich glycoproteins (HRGPs, extensins) have been proposed to be involved in aluminum (Al) resistance mechanisms in plants. We have characterized the distribution of extensin in pea (Pisum sativum L.) root nodules apoplast under short (for 2 and 24h) Al stress. Monoclonal antibodie LM1 have been used to locate extensin protein epitope by immunofluorescence and immunogold labeling. The nodules were shown to respond to Al stress by thickening of plant and infection thread (IT) walls and disturbances in threads growth and bacteria endocytosis. Immunoblot results indicated the presence of a 17-kDa band specific for LM1. Irrespective of the time of Al stress, extensin content increased in root nodules. Further observation utilizing fluorescence and transmission electron microscope showed that LM1 epitope was localized in walls and intercellular spaces of nodule cortex tissues and in the infection threads matrix. Al stress in nodules appears to be associated with higher extensin accumulation in matrix of enlarged thick-walled ITs. In addition to ITs, thickened walls and intercellular spaces of nodule cortex were also associated with intense extensin accumulation. These data suggest that Al-induced extensin accumulation in plant cell walls and ITs matrix may have influence on the process of IT growth and tissue and cell colonization by Rhizobium bacteria. Copyright © 2014 Elsevier Ltd. All rights reserved.

Transient phosphorylation of tumor associated microtubule associated protein (TMAP)/cytoskeleton associated protein 2 (CKAP2) at Thr-596 during early phases of mitosis

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Hong, Kyung Uk; Choi, Yong-Bock; Lee, Jung-Hwa; Kim, Hyun-Jun; Kwon, Hye-Rim; Seong, Yeon-Sun; Kim, Heung Tae; Park, Joobae; Bae, Chang-Dae; Hong, Kyeong-Man

2008-01-01

Tumor associated microtubule associated protein (TMAP), also known as cytoskeleton associated protein 2 (CKAP2) is a mitotic spindle-associated protein whose expression is cell cycle-regulated and also frequently deregulated in cancer cells. Two monoclonal antibodies (mAbs) against TMAP/CKAP2 were produced: B-1-13 and D-12-3. Interestingly, the reactivity of mAb D-12-3 to TMAP/CKAP2 was markedly decreased specifically in mitotic cell lysate. The epitope mapping study showed that mAb D-12-3 re...

The MCM-associated protein MCM-BP is important for human nuclear morphology.

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Jagannathan, Madhav; Sakwe, Amos M; Nguyen, Tin; Frappier, Lori

2012-01-01

Mini-chromosome maintenance complex-binding protein (MCM-BP) was discovered as a protein that is strongly associated with human MCM proteins, known to be crucial for DNA replication in providing DNA helicase activity. The Xenopus MCM-BP homologue appears to play a role in unloading MCM complexes from chromatin after DNA synthesis; however, the importance of MCM-BP and its functional contribution to human cells has been unclear. Here we show that depletion of MCM-BP by sustained expression of short hairpin RNA (shRNA) results in highly abnormal nuclear morphology and centrosome amplification. The abnormal nuclear morphology was not seen with depletion of other MCM proteins and was rescued with shRNA-resistant MCM-BP. MCM-BP depletion was also found to result in transient activation of the G2 checkpoint, slowed progression through G2 and increased replication protein A foci, indicative of replication stress. In addition, MCM-BP depletion led to increased cellular levels of MCM proteins throughout the cell cycle including soluble MCM pools. The results suggest that MCM-BP makes multiple contributions to human cells that are not limited to unloading of the MCM complex.

The Candida albicans stress response gene Stomatin-Like Protein 3 is implicated in ROS-induced apoptotic-like death of yeast phase cells.

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Karen A Conrad

Full Text Available The ubiquitous presence of SPFH (Stomatin, Prohibitin, Flotillin, HflK/HflC proteins in all domains of life suggests that their function would be conserved. However, SPFH functions are diverse with organism-specific attributes. SPFH proteins play critical roles in physiological processes such as mechanosensation and respiration. Here, we characterize the stomatin ORF19.7296/SLP3 in the opportunistic human pathogen Candida albicans. Consistent with the localization of stomatin proteins, a Slp3p-Yfp fusion protein formed visible puncta along the plasma membrane. We also visualized Slp3p within the vacuolar lumen. Slp3p primary sequence analyses identified four putative S-palmitoylation sites, which may facilitate membrane localization and are conserved features of stomatins. Plasma membrane insertion sequences are present in mammalian and nematode SPFH proteins, but are absent in Slp3p. Strikingly, Slp3p was present in yeast cells, but was absent in hyphal cells, thus categorizing it as a yeast-phase specific protein. Slp3p membrane fluorescence significantly increased in response to cellular stress caused by plasma membrane, cell wall, oxidative, or osmotic perturbants, implicating SLP3 as a general stress-response gene. A slp3Δ/Δ homozygous null mutant had no detected phenotype when slp3Δ/Δ mutants were grown in the presence of a variety of stress agents. Also, we did not observe a defect in ion accumulation, filamentation, endocytosis, vacuolar structure and function, cell wall structure, or cytoskeletal structure. However, SLP3 over-expression triggered apoptotic-like death following prolonged exposure to oxidative stress or when cells were induced to form hyphae. Our findings reveal the cellular localization of Slp3p, and for the first time associate Slp3p function with the oxidative stress response.