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Sample records for hsp90 orchestrates stress

  1. Stressing Out Hsp90 in Neurotoxic Proteinopathies.

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    Inda, Carmen; Bolaender, Alexander; Wang, Tai; Gandu, Srinivasa R; Koren, John

    2016-01-01

    A toxic accumulation of proteins is the hallmark pathology of several neurodegenerative disorders. Protein accumulation is regularly prevented by the network of molecular chaperone proteins, including and especially Hsp90. For reasons not yet elucidated, Hsp90 and the molecular chaperones interact with, but do not degrade, these toxic proteins resulting in the pathogenic accumulation of proteins such as tau, in Alzheimer's Disease, and α-synuclein, in Parkinson's Disease. In this review, we describe the associations between Hsp90 and the pathogenic and driver proteins of several neurodegenerative disorders. We additionally describe how the inhibition of Hsp90 promotes the degradation of both mutant and pathogenic protein species in models of neurodegenerative diseases. We also examine the current state of Hsp90 inhibitors capable of crossing the blood-brain barrier; compounds which may be capable of slowing, preventing, and possible reversing neurodegenerative diseases.

  2. Overexpression of GmHsp90s, a heat shock protein 90 (Hsp90 gene family cloning from soybean, decrease damage of abiotic stresses in Arabidopsis thaliana.

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    Jinyan Xu

    Full Text Available Hsp90 is one of the most conserved and abundant molecular chaperones and is an essential component of the protective stress response; however, its roles in abiotic stress responses in soybean (Glycine max remain obscure. Here, 12 GmHsp90 genes from soybean were identified and found to be expressed and to function differentially under abiotic stresses. The 12 GmHsp90 genes were isolated and named GmHsp90A1-GmHsp90A6, GmHsp90B1, GmHsp90B2, GmHsp90C1.1, GmHsp90C1.2, GmHsp90C2.1 and GmHsp90C2.2 based on their characteristics and high homology to other Hsp90s according to a new nomenclature system. Quantitative real-time PCR expression data revealed that all the genes exhibited higher transcript levels in leaves and could be strongly induced under heat, osmotic and salt stress but not cold stress. Overexpression of five typical genes (GmHsp90A2, GmHsp90A4, GmHsp90B1, GmHsp90C1.1 and GmHsp90C2.1 in Arabidopsis thaliana provided useful evidences that GmHsp90 genes can decrease damage of abiotic stresses. In addition, an abnormal accumulation of proline was detected in some transgenic Arabidopsis plants suggested overexpressing GmHsp90s may affect the synthesis and response system of proline. Our work represents a systematic determination of soybean genes encoding Hsp90s, and provides useful evidence that GmHsp90 genes function differently in response to abiotic stresses and may affect the synthesis and response system of proline.

  3. Hsp90 Is Essential under Heat Stress in the Bacterium Shewanella oneidensis

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    Flora Ambre Honoré

    2017-04-01

    Full Text Available The Hsp90 chaperone is essential in eukaryotes and activates a large array of client proteins. In contrast, its role is still elusive in bacteria, and only a few Hsp90 bacterial clients are known. Here, we found that Hsp90 is essential in the model bacterium Shewanella oneidensis under heat stress. A genetic screen for Hsp90 client proteins identified TilS, an essential protein involved in tRNA maturation. Overexpression of TilS rescued the growth defect of the hsp90 deletion strain under heat stress. In vivo, the activity and the amount of TilS were significantly reduced in the absence of Hsp90 at high temperature. Furthermore, we showed that Hsp90 interacts with TilS, and Hsp90 prevents TilS aggregation in vitro at high temperature. Together, our results indicate that TilS is a client of Hsp90 in S. oneidensis. Therefore, our study links the essentiality of bacterial Hsp90 at high temperature with the identification of a client.

  4. Effect of thermal stress on HSP90 expression of Bali cattle in Barru district, South Sulawesi

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    Aritonang, S. B.; Yuniati, R.; Abinawanto, Imron, M.; Bowolaksono, A.

    2017-07-01

    Heat shock protein 90-kDa is induced stress protein that expressed in response to stress and play crucial roles in environmental stress tolerance and adaptation. This study aimed to determine effect of environmental heat stress on the HSP90 expression of Bali cattle. Heat stress was measured by temperature humidity index in the morning and evening across 5-days on August 2016. The blood samples of Bali cattle were taken from venous jungularis. HSP90 was derived from RNA isolation of whole blood then was followed reverse transcription two steps. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) was performed to analyze the transcript variants of HSP90, followed by comparative ΔΔCt to determine HSP90 expression. The results of temperature and humidity index (THI) measurement indicated THI on afternoon was higher than in the morning. The difference in environmental conditions in the morning and afternoon effected changes on rectal temperature but neither did on Hsp90 expression.

  5. Client Proteins and Small Molecule Inhibitors Display Distinct Binding Preferences for Constitutive and Stress-Induced HSP90 Isoforms and Their Conformationally Restricted Mutants.

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    Thomas L Prince

    Full Text Available The two cytosolic/nuclear isoforms of the molecular chaperone HSP90, stress-inducible HSP90α and constitutively expressed HSP90β, fold, assemble and maintain the three-dimensional structure of numerous client proteins. Because many HSP90 clients are important in cancer, several HSP90 inhibitors have been evaluated in the clinic. However, little is known concerning possible unique isoform or conformational preferences of either individual HSP90 clients or inhibitors. In this report, we compare the relative interaction strength of both HSP90α and HSP90β with the transcription factors HSF1 and HIF1α, the kinases ERBB2 and MET, the E3-ubiquitin ligases KEAP1 and RHOBTB2, and the HSP90 inhibitors geldanamycin and ganetespib. We observed unexpected differences in relative client and drug preferences for the two HSP90 isoforms, with HSP90α binding each client protein with greater apparent affinity compared to HSP90β, while HSP90β bound each inhibitor with greater relative interaction strength compared to HSP90α. Stable HSP90 interaction was associated with reduced client activity. Using a defined set of HSP90 conformational mutants, we found that some clients interact strongly with a single, ATP-stabilized HSP90 conformation, only transiently populated during the dynamic HSP90 chaperone cycle, while other clients interact equally with multiple HSP90 conformations. These data suggest different functional requirements among HSP90 clientele that, for some clients, are likely to be ATP-independent. Lastly, the two inhibitors examined, although sharing the same binding site, were differentially able to access distinct HSP90 conformational states.

  6. Hsp90 orchestrates transcriptional regulation by Hsf1 and cell wall remodelling by MAPK signalling during thermal adaptation in a pathogenic yeast.

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    Michelle D Leach

    2012-12-01

    Full Text Available Thermal adaptation is essential in all organisms. In yeasts, the heat shock response is commanded by the heat shock transcription factor Hsf1. Here we have integrated unbiased genetic screens with directed molecular dissection to demonstrate that multiple signalling cascades contribute to thermal adaptation in the pathogenic yeast Candida albicans. We show that the molecular chaperone heat shock protein 90 (Hsp90 interacts with and down-regulates Hsf1 thereby modulating short term thermal adaptation. In the longer term, thermal adaptation depends on key MAP kinase signalling pathways that are associated with cell wall remodelling: the Hog1, Mkc1 and Cek1 pathways. We demonstrate that these pathways are differentially activated and display cross talk during heat shock. As a result ambient temperature significantly affects the resistance of C. albicans cells to cell wall stresses (Calcofluor White and Congo Red, but not osmotic stress (NaCl. We also show that the inactivation of MAP kinase signalling disrupts this cross talk between thermal and cell wall adaptation. Critically, Hsp90 coordinates this cross talk. Genetic and pharmacological inhibition of Hsp90 disrupts the Hsf1-Hsp90 regulatory circuit thereby disturbing HSP gene regulation and reducing the resistance of C. albicans to proteotoxic stresses. Hsp90 depletion also affects cell wall biogenesis by impairing the activation of its client proteins Mkc1 and Hog1, as well as Cek1, which we implicate as a new Hsp90 client in this study. Therefore Hsp90 modulates the short term Hsf1-mediated activation of the classic heat shock response, coordinating this response with long term thermal adaptation via Mkc1- Hog1- and Cek1-mediated cell wall remodelling.

  7. Sequence features and phylogenetic analysis of the stress protein Hsp90α in chinook salmon Oncorhynchus tshawytscha, a poikilothermic vertebrate

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    Palmisano, Aldo N.; Winton, James R.; Dickhoff, Walton W.

    1999-01-01

    We cloned and sequenced a chinook salmon Hsp90 cDNA; sequence analysis shows it to be Hsp90??. Phylogenetic analysis supports the hypothesis that ?? and ?? paralogs of Hsp90 arose as a result of a gene duplication event and that they diverged early in the evolution of vertebrates, before tetrapods separated from the teleost lineage. Among several differences distinguishing poikilothermic Hsp90?? sequences from their bird and mammal orthologs, the teleost versions specifically lack a characteristic QTQDQP phosphorylation site near the N-terminus. We used the cDNA to develop an RNA (Northern) blot to quantify cellular Hsp90 mRNA levels. Chinook salmon embryonic (CHSE-214) cells responded to heat shock with a rapid rise in Hsp90 mRNA through 4 h, followed by a gradual decline over the next 20 h. Hsp90 mRNA level may be useful as a stress indicator, especially in a laboratory setting or in response to acute heat stress.

  8. Involvement of yeast HSP90 isoforms in response to stress and cell death induced by acetic acid.

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    Alexandra Silva

    Full Text Available Acetic acid-induced apoptosis in yeast is accompanied by an impairment of the general protein synthesis machinery, yet paradoxically also by the up-regulation of the two isoforms of the heat shock protein 90 (HSP90 chaperone family, Hsc82p and Hsp82p. Herein, we show that impairment of cap-dependent translation initiation induced by acetic acid is caused by the phosphorylation and inactivation of eIF2α by Gcn2p kinase. A microarray analysis of polysome-associated mRNAs engaged in translation in acetic acid challenged cells further revealed that HSP90 mRNAs are over-represented in this polysome fraction suggesting preferential translation of HSP90 upon acetic acid treatment. The relevance of HSP90 isoform translation during programmed cell death (PCD was unveiled using genetic and pharmacological abrogation of HSP90, which suggests opposing roles for HSP90 isoforms in cell survival and death. Hsc82p appears to promote survival and its deletion leads to necrotic cell death, while Hsp82p is a pro-death molecule involved in acetic acid-induced apoptosis. Therefore, HSP90 isoforms have distinct roles in the control of cell fate during PCD and their selective translation regulates cellular response to acetic acid stress.

  9. Ammonia stress under high environmental ammonia induces Hsp70 and Hsp90 in the mud eel, Monopterus cuchia.

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    Hangzo, Hnunlalliani; Banerjee, Bodhisattwa; Saha, Shrabani; Saha, Nirmalendu

    2017-02-01

    The obligatory air-breathing mud eel (Monopterus cuchia) is frequently being challenged with high environmental ammonia (HEA) exposure in its natural habitats. The present study investigated the possible induction of heat shock protein 70 and 90 (hsp70, hsc70, hsp90α and hsp90β) genes and more expression of Hsp70 and Hsp90 proteins under ammonia stress in different tissues of the mud eel after exposure to HEA (50 mM NH 4 Cl) for 14 days. HEA resulted in significant accumulation of toxic ammonia in different body tissues and plasma, which was accompanied with the stimulation of oxidative stress in the mud eel as evidenced by more accumulation of malondialdehyde (MDA) and hydrogen peroxide (H 2 O 2 ) during exposure to HEA. Further, hyper-ammonia stress led to significant increase in the levels of mRNA transcripts for inducible hsp70 and hsp90α genes and also their translated proteins in different tissues probably as a consequence of induction of hsp70 and hsp90α genes in the mud eel. However, hyper-ammonia stress was neither associated with any significant alterations in the levels of mRNA transcripts for constitutive hsc70 and hsp90β genes nor their translated proteins in any of the tissues studied. More abundance of Hsp70 and Hsp90α proteins might be one of the strategies adopted by the mud eel to defend itself from the ammonia-induced cellular damages under ammonia stress. Further, this is the first report of ammonia-induced induction of hsp70 and hsp90α genes under hyper-ammonia stress in any freshwater air-breathing teleost.

  10. Cloning of three heat shock protein genes (HSP70, HSP90α and HSP90β) and their expressions in response to thermal stress in loach (Misgurnus anguillicaudatus) fed with different levels of vitamin C.

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    Yan, Jie; Liang, Xiao; Zhang, Yin; Li, Yang; Cao, Xiaojuan; Gao, Jian

    2017-07-01

    Heat shock protein 70 (HSP70) and 90 (HSP90) are the most broadly studied proteins in HSP families. They play key roles in cells as molecular chaperones, in response to stress conditions such as thermal stress. In this study, full-length cDNA sequences of HSP70, HSP90α and HSP90β from loach Misgurnus anguillicaudatus were cloned. The full-length cDNA of HSP70 in loach was 2332bp encoding 644 amino acids, while HSP90α and HSP90β were 2586bp and 2678bp in length, encoding 729 and 727 amino acids, respectively. The deduced amino acid sequences of HSP70 in loach shared the highest identity with those of Megalobrama amblycephala and Cyprinus carpio. The deduced amino acid sequences of HSP90α and HSP90β in loach both shared the highest identity with those of M. amblycephala. Their mRNA tissue expression results showed that the maximum expressions of HSP70, HSP90α and HSP90β were respectively present in the intestine, brain and kidney of loach. Quantitative real-time PCR was employed to analyze the temporal expressions of HSP70, HSP90α and HSP90β in livers of loaches fed with different levels of vitamin C under thermal stress. Expression levels of the three HSP genes in loach fed the diet without vitamin C supplemented at 0 h of thermal stress were significantly lower than those at 2 h, 6 h, 12 h and 24 h of thermal stress. It indicated that expressions of the three HSP genes were sensitive to thermal stress in loach. The three HSP genes in loaches fed with 1000 mg/kg vitamin C expressed significantly lower than other vitamin C groups at many time points of thermal stress, suggesting 1000 mg/kg dietary vitamin C might decrease the body damages caused by the thermal stress. This study will be of value for further studies into thermal stress tolerance in loach. Copyright © 2017 Elsevier Ltd. All rights reserved.

  11. A comparative evaluation of crowding stress on muscle HSP90 and myostatin expression in salmonids

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    Galt, Nicholas J.; Froehlich, Jacob Michael; McCormick, Stephen; Biga, Peggy R.

    2018-01-01

    Stress is a major factor that contributes to poor production and animal welfare concerns in aquaculture. As such, a thorough understanding of mechanisms involved in the stress response is imperative to developing strategies to mitigate the negative side effects of stressors, including the impact of high stocking densities on growth. The purpose of this study was to determine how the muscle growth inhibitor, myostatin, and the stress-responsive gene HSP90 are regulated in response to crowding stress in rainbow trout (Oncorhynchus mykiss), cutthroat trout (Oncorhynchus clarki), brook trout (Salvelinus fontinalis), and Atlantic salmon (Salmo salar). All species exhibited higher cortisol and glucose levels following the handling stress, indicating physiological response to the treatment. Additionally, all species, except rainbow trout, exhibited higher HSP90 levels in muscle after a 48 h crowding stress. Crowding stress resulted in a decrease of myostatin-1ain brook trout white muscle but not red muscle, while, myostatin-1a and -2a levels increased in white muscle and myostatin-1b levels increased in red muscle in Atlantic salmon. In rainbow trout, no significant changes were detected in either muscle type, but myostatin-1awas upregulated in both white and red skeletal muscle in the closely related cutthroat trout. The variation in response to crowding suggests a complex and species-specific interaction between stress and the muscle gene regulation in these salmonids. Only Atlantic salmon and cutthroat trout exhibited increased muscle myostatin transcription, and also exhibited the largest increase in circulating glucose in response to crowding. These results suggest that species-specific farming practices should be carefully examined in order to optimize low stress culture conditions.

  12. Molecular characterization of three Hsp90 from Pieris and expression patterns in response to cold and thermal stress in summer and winter diapause of Pieris melete.

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    Wu, Yue-Kun; Zou, Chao; Fu, Dao-Meng; Zhang, Wan-Na; Xiao, Hai-Jun

    2018-04-01

    Heat shock proteins (Hsps) have been linked to stresses and winter diapause in insects, but whether they are components of summer diapause is still unknown. In this study, complementary DNAs of Hsp90 from Pieris melete, Pieris rapae and Pieris canidia named PmHsp90, PrHsp90 and PcHsp90, respectively, were cloned and sequenced. The deduced amino acid sequence consisted of 718 amino acid residues with a putative molecular mass of 82.6, 82.6 and 82.7 kDa, respectively. The amino acid sequences contained all of the five conserved signature motifs in the Hsp90 family and a bHLH protein folding activity region. The differential expression pattern of PmHsp90 in response to summer diapause and winter diapause, which are related to heat/cold stress, was investigated. Cold stress induced Hsp90 up-regulation in summer and winter diapause pupae, but not in non-diapause individuals. Heat shock up-regulated PmHsp90 gradually with an increase in temperature in summer diapause, and PmHsp90 was rapidly up-regulated in winter diapause. After 30 min heat shock at 39°C, substantial up-regulation of PmHsp90 transcript levels were observed both in summer and winter diapause. However, in non-diapause a relatively stable expression was found under different durations of 39°C heat shock. Compared to the optimal treatment of 18°C for diapause development, a high temperature acclimation of 31°C induced PmHsp90 up-regulation in summer diapause, whereas a low temperature acclimation of 4°C induced up-regulation in winter diapause. The current results indicate that Hsp90 may play an important role in response to heat/cold stress both in summer and winter diapause. © 2016 Institute of Zoology, Chinese Academy of Sciences.

  13. Molecular mechanisms of canalization: Hsp90 and beyond

    Indian Academy of Sciences (India)

    2006-03-26

    Mar 26, 2006 ... ... and regulatory circuits, accounting for the important role Hsp90 plays in ... by environmental stress, genetic or pharmaceutical targeting of Hsp90. The ... Here, we discuss the role of Hsp90 in canalization and organismal ...

  14. Effect of electromagnetic fields at 2.45 GHz on the levels of cellular stress proteins HSP-90 and 70 in the rat thyroid

    International Nuclear Information System (INIS)

    Misa Agustino, M. J.; Alvarez-Folgueras, M.; Jorge-Mora, M. T.; Jorge Barreiro, F. J.; Ares Pena, F. J.; Lleiro, J.; Lopez Martin, M. E.

    2011-01-01

    In this study we analyzed the cellular stress levels achieved by heat shock proteins (HSP) 90 and 70 in rat thyroid tissue after exposure to radio waves in TWG experimental system. Parallel measurements of body stress in animals by rectal temperature probes allow us to determine whether there is any interaction between temperature increases and cellular stress.

  15. Expression dynamics of HSP90 and nitric oxide synthase (NOS) isoforms during heat stress acclimation in Tharparkar cattle

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    Bharati, Jaya; Dangi, S. S.; Bag, S.; Maurya, V. P.; Singh, G.; Kumar, P.; Sarkar, M.

    2017-08-01

    Six male Tharparkar cattle of 2-3 years old were selected for the study. After 15-day acclimation at thermoneutral zone (TNZ) in psychrometric chamber, animals were exposed at 42 °C for 6 h up to 23 days followed by 12 days of recovery period. Blood samples were collected during control period at TNZ (days 1, 5, and 12), after heat stress exposure (day 1, immediate heat stress acclimation (IHSA); days 2 to 10, short-term heat stress acclimation (STHSA); days 15 to 23, long-term heat stress acclimation (LTHSA); days 7 and 12, recovery period), and peripheral blood mononuclear cells (PBMCs) were isolated for RNA and protein extraction. The messenger RNA (mRNA) and protein expression in PBMCs were determined by qPCR and western blot, respectively. Samples at TNZ were taken as control. The mRNA expression of HSP90, iNOS, and eNOS was significantly upregulated ( P heat stress exposure in Tharparkar cattle.

  16. Hsp90 depletion goes wild

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    Siegal, Mark L; Masel, Joanna

    2012-01-01

    Abstract Hsp90 reveals phenotypic variation in the laboratory, but is Hsp90 depletion important in the wild? Recent work from Chen and Wagner in BMC Evolutionary Biology has discovered a naturally occurring Drosophila allele that downregulates Hsp90, creating sensitivity to cryptic genetic variation. Laboratory studies suggest that the exact magnitude of Hsp90 downregulation is important. Extreme Hsp90 depletion might reactivate transposable elements and/or induce aneuploidy, in addition to r...

  17. Effect of heat stress on the expression profile of Hsp90 among Sahiwal (Bos indicus) and Frieswal (Bos indicus × Bos taurus) breed of cattle: a comparative study.

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    Deb, Rajib; Sajjanar, Basavaraj; Singh, Umesh; Kumar, Sushil; Singh, Rani; Sengar, G; Sharma, Arjava

    2014-02-25

    We evaluated the effect of thermal challenge on the expression profile of heat shock protein 90 (Hsp90) among Sahiwal (Bos indicus) and Frieswal (Bos indicus × Bos taurus) breeds of cattle. The present investigation was focused on the comparative studies on Hsp90 expression among Frieswal and Sahiwal under in vitro and environmental heat stress. Measured immediately after the in vitro heat shock to the peripheral blood mononuclear cells (PBMCs), the relative expression of Hsp90 mRNA was significantly (Pcows consistently recorded higher rectal temperatures than the Sahiwal breed. Further during this peak summer stress, Sahiwal showed significantly higher levels of mRNA transcripts as well as protein concentration compared to the Frieswal breed. Our findings also interestingly showed that, the cell viability of PBMC are significantly higher among the Sahiwal than Frieswal. Taken together, the experiments of both induced in vitro and environmental stress conditions indicate that, Sahiwal may express higher levels of Hsp90 then Frieswal to regulate their body temperature and increase cell survivality under heat stressed conditions. Copyright © 2013 Elsevier B.V. All rights reserved.

  18. Effects of Thermal Stress on the mRNA Expression of SOD, HSP90, and HSP70 in the Spotted Sea Bass ( Lateolabrax maculatus)

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    Shin, Moon-Kyeong; Park, Ho-Ra; Yeo, Won-Jun; Han, Kyung-Nam

    2018-03-01

    The aim of this study was to elucidate the molecular mechanisms underlying the thermal stress response in the spotted sea bass ( Lateolabrax maculatus). Spotted sea basses were exposed to 4 different water temperatures (20, 22, 24, and 28°C) in increasing increments of 2°C/h from 18°C (control) for different time periods (0, 6, 12, 24, 48, 72, and 96 h). Subsequently, 3 tissues (liver, muscle, and gill) were isolated, and the levels of SOD, HSP90, and HSP70 mRNA were assessed. SOD mRNA expression was maintained at baseline levels of control fish at all water temperatures in the liver, while muscle and gill tissue showed an increase followed by a decrease over each certain time with higher water temperature. HSP90 mRNA expression increased in the liver at ≤ 24°C over time, but maintained baseline expression at 28°C. In muscle, HSP90 mRNA expression gradually increased at all water temperatures, but increased and then decreased at ≥ 24°C in gill tissue. HSP70 mRNA expression exhibited an increase and then a decrease in liver tissue at 28°C, but mainly showed similar expression patterns to HSP90 in all tissues. These results suggest the activity of a defense mechanism using SOD, HSP90, and HSP70 in the spotted sea bass upon rapid increases in water temperature, where the expression of these genes indicated differences between tissues in the extent of the defense mechanisms. Also, these results indicate that high water temperature and long-term thermal stress exposure can inhibit physiological defense mechanisms.

  19. Hsp90 depletion goes wild

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    Siegal Mark L

    2012-02-01

    Full Text Available Abstract Hsp90 reveals phenotypic variation in the laboratory, but is Hsp90 depletion important in the wild? Recent work from Chen and Wagner in BMC Evolutionary Biology has discovered a naturally occurring Drosophila allele that downregulates Hsp90, creating sensitivity to cryptic genetic variation. Laboratory studies suggest that the exact magnitude of Hsp90 downregulation is important. Extreme Hsp90 depletion might reactivate transposable elements and/or induce aneuploidy, in addition to revealing cryptic genetic variation. See research article http://wwww.biomedcentral.com/1471-2148/12/25

  20. Effect of electromagnetic fields at 2.45 GHz on the levels of cellular stress proteins HSP-90 and 70 in the rat thyroid; Efecto de los campos electromagneticos a 2,45 GHz sobre los niveles de proteinas de estres celular HSP-90 y 70 en el toroides de rata

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    Misa Agustino, M. J.; Alvarez-Folgueras, M.; Jorge-Mora, M. T.; Jorge Barreiro, F. J.; Ares Pena, F. J.; Lleiro, J.; Lopez Martin, M. E.

    2011-07-01

    In this study we analyzed the cellular stress levels achieved by heat shock proteins (HSP) 90 and 70 in rat thyroid tissue after exposure to radio waves in TWG experimental system. Parallel measurements of body stress in animals by rectal temperature probes allow us to determine whether there is any interaction between temperature increases and cellular stress.

  1. Synergistic role of HSP90α and HSP90β to promote myofibroblast persistence in lung fibrosis.

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    Bellaye, Pierre-Simon; Shimbori, Chiko; Yanagihara, Toyoshi; Carlson, David A; Hughes, Philip; Upagupta, Chandak; Sato, Seidai; Wheildon, Nolan; Haystead, Timothy; Ask, Kjetil; Kolb, Martin

    2018-02-01

    Idiopathic pulmonary fibrosis (IPF) is a progressive disease of the lung parenchyma, causing significant morbidity through worsening dyspnoea and overall functional decline. IPF is characterised by apoptosis-resistant myofibroblasts, which are a major source for the excessive production of extracellular matrix (ECM) overtaking normal lung tissue. We sought to study the role of heat shock protein (HSP) isoforms HSP90α and HSP90β, whose distinct roles in lung fibrogenesis remain elusive.We determined the level of circulating HSP90α in IPF patients (n=31) and age-matched healthy controls (n=9) by ELISA. The release of HSP90α and HSP90β was evaluated in vitro in primary IPF and control lung fibroblasts and ex vivo after mechanical stretch on fibrotic lung slices from rats receiving adenovector-mediated transforming growth factor-β1.We demonstrate that circulating HSP90α is upregulated in IPF patients in correlation with disease severity. The release of HSP90α is enhanced by the increase in mechanical stress of the fibrotic ECM. This increase in extracellular HSP90α signals through low-density lipoprotein receptor-related protein 1 (LRP1) to promote myofibroblast differentiation and persistence. In parallel, we demonstrate that the intracellular form of HSP90β stabilises LRP1, thus amplifying HSP90α extracellular action.We believe that the specific inhibition of extracellular HSP90α is a promising therapeutic strategy to reduce pro-fibrotic signalling in IPF. Copyright ©ERS 2018.

  2. A novel biomarker for marine environmental pollution of HSP90 from Mytilus coruscus

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    Liu, Huihui; Wu, Jiong; Xu, Mengshan; He, Jianyu

    2016-01-01

    Heat shock protein 90 (HSP90) is a conserved molecular chaperone contributing to cell cycle control, organism development and the proper regulation of cytosolic proteins. The full-length HSP90 cDNA of Mytilus coruscus (McHSP90, KT946644) was 2420 bp, including an ORF of 2169 bp encoding a polypeptide of 722 amino acids with predicted pI/MW 4.89/83.22 kDa. BLASTp analysis and phylogenetic relationship strongly suggested McHSP90 was a member of HSP90 family, and it was highly conserved with other known HSP90, especially in the HSP90 family signatures, ATP/GTP-Binding sites and ‘EEVD’ motif. The mRNA of McHSP90 in haemolymph was upregulated in all treatments including Vibrio alginolyticus and Vibrio harveyi challenge, metals stresses (copper and cadmium) and 180 CST fuel exposure. All the results implied the expression of McHSP90 could be affected by Vibrio challenge and environmental stress, which might help us gain more insight into the molecular mechanism of HSP against adverse stresses in mollusca. - Highlights: • A novel HSP90 (McHSP90) was identified from Mytilus coruscus. • McHSP90 significantly affected by Vibrio challenge for immune defense. • McHSP90 mRNA was obviously up-regulated under stress of heavy metals and 180CST fuel. • McHSP90 might be an ideal marine pollution indicator.

  3. Comprehensive identification and expression analysis of Hsp90s gene family in Solanum lycopersicum.

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    Zai, W S; Miao, L X; Xiong, Z L; Zhang, H L; Ma, Y R; Li, Y L; Chen, Y B; Ye, S G

    2015-07-14

    Heat shock protein 90 (Hsp90) is a protein produced by plants in response to adverse environmental stresses. In this study, we identified and analyzed Hsp90 gene family members using a bioinformatic method based on genomic data from tomato (Solanum lycopersicum L.). The results illustrated that tomato contains at least 7 Hsp90 genes distributed on 6 chromosomes; protein lengths ranged from 267-794 amino acids. Intron numbers ranged from 2-19 in the genes. The phylogenetic tree revealed that Hsp90 genes in tomato (Solanum lycopersicum L.), rice (Oryza sativa L.), and Arabidopsis (Arabidopsis thaliana L.) could be divided into 5 groups, which included 3 pairs of orthologous genes and 4 pairs of paralogous genes. Expression analysis of RNA-sequence data showed that the Hsp90-1 gene was specifically expressed in mature fruits, while Hsp90-5 and Hsp90-6 showed opposite expression patterns in various tissues of cultivated and wild tomatoes. The expression levels of the Hsp90-1, Hsp90-2, and Hsp90- 3 genes in various tissues of cultivated tomatoes were high, while both the expression levels of genes Hsp90-3 and Hsp90-4 were low. Additionally, quantitative real-time polymerase chain reaction showed that these genes were involved in the responses to yellow leaf curl virus in tomato plant leaves. Our results provide a foundation for identifying the function of the Hsp90 gene in tomato.

  4. Hsp90 selectively modulates phenotype in vertebrate development.

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    Patricia L Yeyati

    2007-03-01

    Full Text Available Compromised heat shock protein 90 (Hsp90 function reveals cryptic phenotypes in flies and plants. These observations were interpreted to suggest that this molecular stress-response chaperone has a capacity to buffer underlying genetic variation. Conversely, the protective role of Hsp90 could account for the variable penetrance or severity of some heritable developmental malformations in vertebrates. Using zebrafish as a model, we defined Hsp90 inhibitor levels that did not induce a heat shock response or perturb phenotype in wild-type strains. Under these conditions the severity of the recessive eye phenotype in sunrise, caused by a pax6b mutation, was increased, while in dreumes, caused by a sufu mutation, it was decreased. In another strain, a previously unobserved spectrum of severe structural eye malformations, reminiscent of anophthalmia, microphthalmia, and nanophthalmia complex in humans, was uncovered by this limited inhibition of Hsp90 function. Inbreeding of offspring from selected unaffected carrier parents led to significantly elevated malformation frequencies and revealed the oligogenic nature of this phenotype. Unlike in Drosophila, Hsp90 inhibition can decrease developmental stability in zebrafish, as indicated by increased asymmetric presentation of anophthalmia, microphthalmia, and nanophthalmia and sunrise phenotypes. Analysis of the sunrise pax6b mutation suggests a molecular mechanism for the buffering of mutations by Hsp90. The zebrafish studies imply that mild perturbation of Hsp90 function at critical developmental stages may underpin the variable penetrance and expressivity of many developmental anomalies where the interaction between genotype and environment plays a major role.

  5. Perbedaan Kadar HSP90 pada Preeklamsi Berat dengan Kehamilan Normal

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    Soetrisno

    2015-06-01

    Full Text Available Severe pre-eclampsia is the second highest cause of maternal mortality. Free radicals that stimulate heat shock protein 90 (HSP 90 are believed to determine severe pre-eclampsia. HSP90 is an important protein that helps the establishment and maintenance of other proteins. It also increases the life time of cells after various pathological conditions (chaperone function. The chaperone function is the adaptation key factor to endogenous stress in tissues. By recognizing HSP90 level in early detection of severe pre-eclampsia, prevention and management can be started early. This study aimed to prove that the HSP90 level in pregnancy with severe pre-eclampsia is higher than normal pregnancy. This was a quantitative study using cross sectional approach by testing the HSP90 level. The study was conducted during the period of September to November 2013, at the Obstetrics and Gynecological Unit, Moewardi Hospital Surakarta and Prodia Laboratory Jakarta. The number of subjects was 30 patients, consisting of 15 normal pregnant mothers and 15 pregnant mothers with pre-eclampsia . The calculation of serum HSP90 level was conducted using enzyme-linked immunosorbent assay (ELISA. Data were analyzed using t-test using SPSS for Windows version 17 for Windows. The mean of HSP90 in the severe pre-eclampsia group was 131.91±26.66 while the mean in the normal pregnancy group was 80.28±13.39 with p=0.00 (p<0.05. Level of HSP90 serum in severe pre-eclampsia is higher than in normal pregnancy, due to the occurrence of oxidative stress in severe pre-eclampsia

  6. Heat Shock Protein 90 (Hsp90 Expression and Breast Cancer

    Directory of Open Access Journals (Sweden)

    Christos A. Papadimitriou

    2012-09-01

    Full Text Available Hsp90 is an abundant protein in mammalian cells. It forms several discrete complexes, each containing distinct groups of co-chaperones that assist protein folding and refolding during stress, protein transport and degradation. It interacts with a variety of proteins that play key roles in breast neoplasia including estrogen receptors, tumor suppressor p53 protein, angiogenesis transcription factor HIF-1alpha, antiapoptotic kinase Akt, Raf-1 MAP kinase and a variety of receptor tyrosine kinases of the erbB family. Elevated Hsp90 expression has been documented in breast ductal carcinomas contributing to the proliferative activity of breast cancer cells; whilst a significantly decreased Hsp90 expression has been shown in infiltrative lobular carcinomas and lobular neoplasia. Hsp90 overexpression has been proposed as a component of a mechanism through which breast cancer cells become resistant to various stress stimuli. Therefore, pharmacological inhibition of HSPs can provide therapeutic opportunities in the field of cancer treatment. 17-allylamino,17-demethoxygeldanamycin is the first Hsp90 inhibitor that has clinically been investigated in phase II trial, yielding promising results in patients with HER2-overexpressing metastatic breast cancer, whilst other Hsp90 inhibitors (retaspimycin HCL, NVP-AUY922, NVP-BEP800, CNF2024/BIIB021, SNX-5422, STA-9090, etc. are currently under evaluation.

  7. Hsp90 governs echinocandin resistance in the pathogenic yeast Candida albicans via calcineurin.

    Directory of Open Access Journals (Sweden)

    Sheena D Singh

    2009-07-01

    Full Text Available Candida albicans is the leading fungal pathogen of humans, causing life-threatening disease in immunocompromised individuals. Treatment of candidiasis is hampered by the limited number of antifungal drugs whose efficacy is compromised by host toxicity, fungistatic activity, and the emergence of drug resistance. We previously established that the molecular chaperone Hsp90, which regulates the form and function of diverse client proteins, potentiates resistance to the azoles in C. albicans and in the model yeast Saccharomyces cerevisiae. Genetic studies in S. cerevisiae revealed that Hsp90's role in azole resistance is to enable crucial cellular responses to the membrane stress exerted by azoles via the client protein calcineurin. Here, we demonstrate that Hsp90 governs cellular circuitry required for resistance to the only new class of antifungals to reach the clinic in decades, the echinocandins, which inhibit biosynthesis of a critical component of the fungal cell wall. Pharmacological or genetic impairment of Hsp90 function reduced tolerance of C. albicans laboratory strains and resistance of clinical isolates to the echinocandins and created a fungicidal combination. Compromising calcineurin function phenocopied compromising Hsp90 function. We established that calcineurin is an Hsp90 client protein in C. albicans: reciprocal co-immunoprecipitation validated physical interaction; Hsp90 inhibition blocked calcineurin activation; and calcineurin levels were depleted upon genetic reduction of Hsp90. The downstream effector of calcineurin, Crz1, played a partial role in mediating calcineurin-dependent stress responses activated by echinocandins. Hsp90's role in echinocandin resistance has therapeutic potential given that genetic compromise of C. albicans HSP90 expression enhanced the efficacy of an echinocandin in a murine model of disseminated candidiasis. Our results identify the first Hsp90 client protein in C. albicans, establish an entirely

  8. Cloning and Expression of a Cytosolic HSP90 Gene in Chlorella vulgaris

    Directory of Open Access Journals (Sweden)

    Zhengyi Liu

    2014-01-01

    Full Text Available Heat shock protein 90 (HSP90, a highly conserved molecular chaperone, plays essential roles in folding, keeping structural integrity, and regulating the subset of cytosolic proteins. We cloned the cDNA of Chlorella vulgaris HSP90 (named CvHSP90 by combining homology cloning with rapid amplification of cDNA ends (RACE. Sequence analysis indicated that CvHSP90 is a cytosolic member of the HSP90 family. Quantitative RT-PCR was applied to determine the expression level of messenger RNA (mRNA in CvHSP90 under different stress conditions. C. vulgaris was kept in different temperatures (5–45°C for 1 h. The mRNA expression level of CvHSP90 increased with temperature from 5 to 10°C, went further from 35 to 40°C, and reached the maximum at 40°C. On the other hand, for C. vulgaris kept at 35°C for different durations, the mRNA expression level of CvHSP90 increased gradually and reached the peak at 7 h and then declined progressively. In addition, the expression level of CvHSP90 at 40 or 45 in salinity (‰ was almost fourfold of that at 25 in salinity (‰ for 2 h. Therefore, CvHSP90 may be a potential biomarker to monitor environment changes.

  9. The Double-Edged Sword: Conserved Functions of Extracellular Hsp90 in Wound Healing and Cancer

    Energy Technology Data Exchange (ETDEWEB)

    Hance, Michael W.; Nolan, Krystal D.; Isaacs, Jennifer S., E-mail: isaacsj@musc.edu [Department of Cell and Molecular Pharmacology, Medical University of South Carolina, Hollings Cancer Center, Charleston, SC 29412 (United States)

    2014-05-06

    Heat shock proteins (Hsps) represent a diverse group of chaperones that play a vital role in the protection of cells against numerous environmental stresses. Although our understanding of chaperone biology has deepened over the last decade, the “atypical” extracellular functions of Hsps have remained somewhat enigmatic and comparatively understudied. The heat shock protein 90 (Hsp90) chaperone is a prototypic model for an Hsp family member exhibiting a duality of intracellular and extracellular functions. Intracellular Hsp90 is best known as a master regulator of protein folding. Cancers are particularly adept at exploiting this function of Hsp90, providing the impetus for the robust clinical development of small molecule Hsp90 inhibitors. However, in addition to its maintenance of protein homeostasis, Hsp90 has also been identified as an extracellular protein. Although early reports ascribed immunoregulatory functions to extracellular Hsp90 (eHsp90), recent studies have illuminated expanded functions for eHsp90 in wound healing and cancer. While the intended physiological role of eHsp90 remains enigmatic, its evolutionarily conserved functions in wound healing are easily co-opted during malignancy, a pathology sharing many properties of wounded tissue. This review will highlight the emerging functions of eHsp90 and shed light on its seemingly dichotomous roles as a benevolent facilitator of wound healing and as a sinister effector of tumor progression.

  10. Hsp90: Friends, clients and natural foes.

    Science.gov (United States)

    Verma, Sharad; Goyal, Sukriti; Jamal, Salma; Singh, Aditi; Grover, Abhinav

    2016-08-01

    Hsp90, a homodimeric ATPase, is responsible for the correct folding of a number of newly synthesized polypeptides in addition to the correct folding of denatured/misfolded client proteins. It requires several co-chaperones and other partner proteins for chaperone activity. Due to the involvement of Hsp90-dependent client proteins in a variety of oncogenic signaling pathways, Hsp90 inhibition has emerged as one of the leading strategies for anticancer chemotherapeutics. Most of Hsp90 inhibitors blocks the N terminal ATP binding pocket and prevents the conformational changes which are essential for the loading of co-chaperones and client proteins. Several other inhibitors have also been reported which disrupt chaperone cycle in ways other than binding to N terminal ATP binding pocket. The Hsp90 inhibition is associated with heat shock response, mediated by HSF-1, to overcome the loss of Hsp90 and sustain cell survival. This review is an attempt to give an over view of all the important players of chaperone cycle. Copyright © 2016 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.

  11. Probing molecular mechanisms of the Hsp90 chaperone: biophysical modeling identifies key regulators of functional dynamics.

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    Anshuman Dixit

    Full Text Available Deciphering functional mechanisms of the Hsp90 chaperone machinery is an important objective in cancer biology aiming to facilitate discovery of targeted anti-cancer therapies. Despite significant advances in understanding structure and function of molecular chaperones, organizing molecular principles that control the relationship between conformational diversity and functional mechanisms of the Hsp90 activity lack a sufficient quantitative characterization. We combined molecular dynamics simulations, principal component analysis, the energy landscape model and structure-functional analysis of Hsp90 regulatory interactions to systematically investigate functional dynamics of the molecular chaperone. This approach has identified a network of conserved regions common to the Hsp90 chaperones that could play a universal role in coordinating functional dynamics, principal collective motions and allosteric signaling of Hsp90. We have found that these functional motifs may be utilized by the molecular chaperone machinery to act collectively as central regulators of Hsp90 dynamics and activity, including the inter-domain communications, control of ATP hydrolysis, and protein client binding. These findings have provided support to a long-standing assertion that allosteric regulation and catalysis may have emerged via common evolutionary routes. The interaction networks regulating functional motions of Hsp90 may be determined by the inherent structural architecture of the molecular chaperone. At the same time, the thermodynamics-based "conformational selection" of functional states is likely to be activated based on the nature of the binding partner. This mechanistic model of Hsp90 dynamics and function is consistent with the notion that allosteric networks orchestrating cooperative protein motions can be formed by evolutionary conserved and sparsely connected residue clusters. Hence, allosteric signaling through a small network of distantly connected

  12. Intracellular dynamics of the Hsp90 co-chaperone p23 is dictated by Hsp90

    International Nuclear Information System (INIS)

    Picard, Didier

    2006-01-01

    p23 is a component of the Hsp90 molecular chaperone machine. It binds and stabilizes the ATP-bound dimeric form of Hsp90. Since Hsp90 binds protein substrates in the ATP conformation, p23 has been proposed to stabilize Hsp90-substrate complexes. In addition, p23 can also function as a molecular chaperone by itself and even possesses an unrelated enzymatic activity. Whether it fulfills the latter functions in cells while bound to Hsp90 remains unknown and is difficult to extrapolate from cell-free biochemical experiments. Using the 'fluorescence recovery after photobleaching' (FRAP) technology, I have examined the dynamics of human p23, expressed as a fusion protein with the green fluorescent protein (GFP), in living human HeLa cells. GFP-p23 is distributed throughout the cell, and its mobility is identical in the cytoplasm and in the nucleus. When the Hsp90 interaction is disrupted either with the Hsp90 inhibitor geldanamycin or by introduction of point mutations into p23, the mobility of p23 is greatly accelerated. Under these conditions, its intracellular movement may be diffusion-controlled. In contrast, when wild-type p23 is able to bind Hsp90, a more complex FRAP behavior is observed, suggesting that it is quantitatively bound in Hsp90 complexes undergoing a multitude of other interactions

  13. Targeting Hsp90-Cdc37: A Promising Therapeutic Strategy by Inhibiting Hsp90 Chaperone Function.

    Science.gov (United States)

    Wang, Lei; Li, Li; Gu, Kai; Xu, Xiao-Li; Sun, Yuan; You, Qi-Dong

    2017-01-01

    The Hsp90 chaperone protein regulates the folding, maturation and stability of a wide variety of oncoproteins. In recent years, many Hsp90 inhibitors have entered into the clinical trials while all of them target ATPase showing similar binding capacity and kinds of side-effects so that none have reached to the market. During the regulation progress, numerous protein- protein interactions (PPI) such as Hsp90 and client proteins or cochaperones are involved. With the Hsp90-cochaperones PPI networks being more and more clear, many cancerous proteins have been reported to be tightly correlated to Hsp90-cochaperones PPI. Among them, Hsp90-Cdc37 PPI has been widely reported to associate with numerous protein kinases, making it a novel target for the treatment of cancers. In this paper, we briefly review the strategies and modulators targeting Hsp90-Cdc37 complex including direct and indirect regulation mechanism. Through these discussions we expect to present inspirations for new insights into an alternative way to inhibit Hsp90 chaperone function. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  14. Molecular mechanisms of canalization: Hsp90 and beyond

    Indian Academy of Sciences (India)

    Madhu Sudhan

    2007-03-26

    Mar 26, 2007 ... In plants, Hsp90 function remained little characterized until recent .... unknown if Hsp90 and chromatin remodeling factors act independently or in ..... protein 90 in plant disease resistance; EMBO J. 22 5690–5699. Maloof J N ...

  15. Hsp90α regulates ATM and NBN functions in sensing and repair of DNA double-strand breaks.

    Science.gov (United States)

    Pennisi, Rosa; Antoccia, Antonio; Leone, Stefano; Ascenzi, Paolo; di Masi, Alessandra

    2017-08-01

    The molecular chaperone heat shock protein 90 (Hsp90α) regulates cell proteostasis and mitigates the harmful effects of endogenous and exogenous stressors on the proteome. Indeed, the inhibition of Hsp90α ATPase activity affects the cellular response to ionizing radiation (IR). Although the interplay between Hsp90α and several DNA damage response (DDR) proteins has been reported, its role in the DDR is still unclear. Here, we show that ataxia-telangiectasia-mutated kinase (ATM) and nibrin (NBN), but not 53BP1, RAD50, and MRE11, are Hsp90α clients as the Hsp90α inhibitor 17-(allylamino)-17-demethoxygeldanamycin (17-AAG) induces ATM and NBN polyubiquitination and proteosomal degradation in normal fibroblasts and lymphoblastoid cell lines. Hsp90α-ATM and Hsp90α-NBN complexes are present in unstressed and irradiated cells, allowing the maintenance of ATM and NBN stability that is required for the MRE11/RAD50/NBN complex-dependent ATM activation and the ATM-dependent phosphorylation of both NBN and Hsp90α in response to IR-induced DNA double-strand breaks (DSBs). Hsp90α forms a complex also with ph-Ser1981-ATM following IR. Upon phosphorylation, NBN dissociates from Hsp90α and translocates at the DSBs, while phThr5/7-Hsp90α is not recruited at the damaged sites. The inhibition of Hsp90α affects nuclear localization of MRE11 and RAD50, impairs DDR signaling (e.g., BRCA1 and CHK2 phosphorylation), and slows down DSBs repair. Hsp90α inhibition does not affect DNA-dependent protein kinase (DNA-PK) activity, which possibly phosphorylates Hsp90α and H2AX after IR. Notably, Hsp90α inhibition causes H2AX phosphorylation in proliferating cells, this possibly indicating replication stress events. Overall, present data shed light on the regulatory role of Hsp90α on the DDR, controlling ATM and NBN stability and influencing the DSBs signaling and repair. © 2017 Federation of European Biochemical Societies.

  16. Heat shock protein Hsp90-2 expression in the Arabidopsis thaliana seedlings under clinorotation

    Science.gov (United States)

    Kozeko, Liudmyla

    Heat shock proteins 90 kDa (Hsp90) are abundant under normal conditions and induced by stress. This family is distinguished from other chaperones in that most of its substrates are signal transduction proteins. Previously, we determined some time-dependent increase in the Hsp90 level in pea seedlings in response to simulated microgravity that indicated a stress-reaction. However, expression of the individual members of the Hsp90 family have specific pattern. The purpose of this study was to investigate possible alterations in the gene expression pattern of cytosolic Hsp90-2 in Arabidopsis thaliana seedlings under 2D-clinorotation. To obtain detailed expression pattern of the HSP90-2 genes we used seeds that provides a resource of loss-of-function mutations gene expression patterns via translational fusions with the reporter gene, GUS (a line N 166718, NASC). There were two variants of the experiment: 1) seedlings grew under clinorotation for 10, 12, 14 d; 2) seedlings grew in the stationary conditions for 10 d followed by clinorotation for 3 h -at 22o C and 16h light cycle. The seedlings grown in the stationary conditions were used as a control. GUS staining showed that HSP90-2 expression was regulated during seedling development and affected by clinorotation in the heterozygous mutant plants. In the homozygous for the mutation plants, HSP90-2 expression was stable during seedling development and not affected by clinorotation. GUS staining was observed in cotyledons, leaves and hypocotyls of the seedlings (especially intense in vascular bundles), indicating intensive cellular processes with participation of this chaperone. Possible pathways of influence of clinorotation on HSP90-2 expression are discussed.

  17. Differences in the ovine HSP90AA1 gene expression rates caused by two linked polymorphisms at its promoter affect rams sperm DNA fragmentation under environmental heat stress conditions.

    Science.gov (United States)

    Salces-Ortiz, Judit; Ramón, Manuel; González, Carmen; Pérez-Guzmán, M Dolores; Garde, J Julián; García-Álvarez, Olga; Maroto-Morales, Alejandro; Calvo, Jorge H; Serrano, M Magdalena

    2015-01-01

    Heat shock (HS) is one of the best-studied exogenous cellular stresses. Almost all tissues, cell types, metabolic pathways and biochemical reactions are affected in greater or lesser extent by HS. However, there are some especially thermo sensible cellular types such as the mammalian male germ cells. The present study examined the role of three INDELs in conjunction with the -660G/C polymorphism located at the HSP90AA1 promoter region over the gene expression rate under HS. Specially, the -668insC INDEL, which is very close to the -660G/C transversion, is a good candidate to be implied in the transcriptional regulation of the gene by itself or in a cooperative way with this SNP. Animals carrying the genotype II-668 showed higher transcription rates than those with ID-668 (FC = 3.07) and DD-668 (FC = 3.40) genotypes for samples collected under HS. A linkage between gene expression and sperm DNA fragmentation was also found. When HS conditions were present along or in some stages of the spermatogenesis, alternative genotypes of the -668insC and -660G/C mutations are involved in the effect of HS over sperm DNA fragmentation. Thus, unfavorable genotypes in terms of gene expression induction (ID-668GC-660 and DD-668GG-660) do not produce enough mRNA (stored as messenger ribonucleoprotein particles) and Hsp90α protein to cope with future thermal stress which might occur in posterior stages when transcriptional activity is reduced and cell types and molecular processes are more sensible to heat (spermatocytes in pachytene and spermatids protamination). This would result in the impairment of DNA packaging and the consequent commitment of the events occurring shortly after fertilization and during embryonic development. In the short-term, the assessment of the relationship between sperm DNA fragmentation sensitivity and ram's fertility will be of interest to a better understanding of the mechanisms of response to HS and its consequences on animal production and

  18. NMNAT2:HSP90 Complex Mediates Proteostasis in Proteinopathies.

    Directory of Open Access Journals (Sweden)

    Yousuf O Ali

    2016-06-01

    Full Text Available Nicotinamide mononucleotide adenylyl transferase 2 (NMNAT2 is neuroprotective in numerous preclinical models of neurodegeneration. Here, we show that brain nmnat2 mRNA levels correlate positively with global cognitive function and negatively with AD pathology. In AD brains, NMNAT2 mRNA and protein levels are reduced. NMNAT2 shifts its solubility and colocalizes with aggregated Tau in AD brains, similar to chaperones, which aid in the clearance or refolding of misfolded proteins. Investigating the mechanism of this observation, we discover a novel chaperone function of NMNAT2, independent from its enzymatic activity. NMNAT2 complexes with heat shock protein 90 (HSP90 to refold aggregated protein substrates. NMNAT2's refoldase activity requires a unique C-terminal ATP site, activated in the presence of HSP90. Furthermore, deleting NMNAT2 function increases the vulnerability of cortical neurons to proteotoxic stress and excitotoxicity. Interestingly, NMNAT2 acts as a chaperone to reduce proteotoxic stress, while its enzymatic activity protects neurons from excitotoxicity. Taken together, our data indicate that NMNAT2 exerts its chaperone or enzymatic function in a context-dependent manner to maintain neuronal health.

  19. Downregulation of the Hsp90 system causes defects in muscle cells of Caenorhabditis elegans.

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    Andreas M Gaiser

    Full Text Available The ATP-dependent molecular chaperone Hsp90 is required for the activation of a variety of client proteins involved in various cellular processes. Despite the abundance of known client proteins, functions of Hsp90 in the organismal context are not fully explored. In Caenorhabditis elegans, Hsp90 (DAF-21 has been implicated in the regulation of the stress-resistant dauer state, in chemosensing and in gonad formation. In a C. elegans strain carrying a DAF-21 mutation with a lower ATP turnover, we observed motility defects. Similarly, a reduction of DAF-21 levels in wild type nematodes leads to reduced motility and induction of the muscular stress response. Furthermore, aggregates of the myosin MYO-3 are visible in muscle cells, if DAF-21 is depleted, implying a role of Hsp90 in the maintenance of muscle cell functionality. Similar defects can also be observed upon knockdown of the Hsp90-cochaperone UNC-45. In life nematodes YFP-DAF-21 localizes to the I-band and the M-line of the muscular ultrastructure, but the protein is not stably attached there. The Hsp90-cofactor UNC-45-CFP contrarily can be found in all bands of the nematode muscle ultrastructure and stably associates with the UNC-54 containing A-band. Thus, despite the physical interaction between DAF-21 and UNC-45, apparently the two proteins are not always localized to the same muscular structures. While UNC-45 can stably bind to myofilaments in the muscular ultrastructure, Hsp90 (DAF-21 appears to participate in the maintenance of muscle structures as a transiently associated diffusible factor.

  20. Exploring the Functional Complementation between Grp94 and Hsp90.

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    Kevin A Maharaj

    Full Text Available Grp94 and Hsp90 are the ER and cytoplasmic paralog members, respectively, of the hsp90 family of molecular chaperones. The structural and biochemical differences between Hsp90 and Grp94 that allow each paralog to efficiently chaperone its particular set of clients are poorly understood. The two paralogs exhibit a high degree of sequence similarity, yet also display significant differences in their quaternary conformations and ATPase activity. In order to identify the structural elements that distinguish Grp94 from Hsp90, we characterized the similarities and differences between the two proteins by testing the ability of Hsp90/Grp94 chimeras to functionally substitute for the wild-type chaperones in vivo. We show that the N-terminal domain or the combination of the second lobe of the Middle domain plus the C-terminal domain of Grp94 can functionally substitute for their yeast Hsp90 counterparts but that the equivalent Hsp90 domains cannot functionally replace their counterparts in Grp94. These results also identify the interface between the Middle and C-terminal domains as an important structural unit within the Hsp90 family.

  1. Identification of HSP90 gene from the Chinese oak silkworm ...

    African Journals Online (AJOL)

    ajl user 1

    2012-06-28

    Jun 28, 2012 ... College of Life Science, Anhui Agricultural University, 130 Changjiang West Road 230036, Peoples ... program consisted of 5 min at 94°C followed by 35 cycles of 94°C ... HSP90 and 73.1% identity with Bombyx mori HSP90.

  2. HSP90 Shapes the Consequences of Human Genetic Variation.

    Science.gov (United States)

    Karras, Georgios I; Yi, Song; Sahni, Nidhi; Fischer, Máté; Xie, Jenny; Vidal, Marc; D'Andrea, Alan D; Whitesell, Luke; Lindquist, Susan

    2017-02-23

    HSP90 acts as a protein-folding buffer that shapes the manifestations of genetic variation in model organisms. Whether HSP90 influences the consequences of mutations in humans, potentially modifying the clinical course of genetic diseases, remains unknown. By mining data for >1,500 disease-causing mutants, we found a strong correlation between reduced phenotypic severity and a dominant (HSP90 ≥ HSP70) increase in mutant engagement by HSP90. Examining the cancer predisposition syndrome Fanconi anemia in depth revealed that mutant FANCA proteins engaged predominantly by HSP70 had severely compromised function. In contrast, the function of less severe mutants was preserved by a dominant increase in HSP90 binding. Reducing HSP90's buffering capacity with inhibitors or febrile temperatures destabilized HSP90-buffered mutants, exacerbating FA-related chemosensitivities. Strikingly, a compensatory FANCA somatic mutation from an "experiment of nature" in monozygotic twins both prevented anemia and reduced HSP90 binding. These findings provide one plausible mechanism for the variable expressivity and environmental sensitivity of genetic diseases. Copyright © 2017 Elsevier Inc. All rights reserved.

  3. A C-terminal HSP90 inhibitor restores glucocorticoid sensitivity and relieves a mouse allograft model of Cushing disease.

    Science.gov (United States)

    Riebold, Mathias; Kozany, Christian; Freiburger, Lee; Sattler, Michael; Buchfelder, Michael; Hausch, Felix; Stalla, Günter K; Paez-Pereda, Marcelo

    2015-03-01

    One function of the glucocorticoid receptor (GR) in corticotroph cells is to suppress the transcription of the gene encoding proopiomelanocortin (POMC), the precursor of the stress hormone adrenocorticotropin (ACTH). Cushing disease is a neuroendocrine condition caused by partially glucocorticoid-resistant corticotroph adenomas that excessively secrete ACTH, which leads to hypercortisolism. Mutations that impair GR function explain glucocorticoid resistance only in sporadic cases. However, the proper folding of GR depends on direct interactions with the chaperone heat shock protein 90 (HSP90, refs. 7,8). We show here that corticotroph adenomas overexpress HSP90 compared to the normal pituitary. N- and C-terminal HSP90 inhibitors act at different steps of the HSP90 catalytic cycle to regulate corticotroph cell proliferation and GR transcriptional activity. C-terminal inhibitors cause the release of mature GR from HSP90, which promotes its exit from the chaperone cycle and potentiates its transcriptional activity in a corticotroph cell line and in primary cultures of human corticotroph adenomas. In an allograft mouse model, the C-terminal HSP90 inhibitor silibinin showed anti-tumorigenic effects, partially reverted hormonal alterations, and alleviated symptoms of Cushing disease. These results suggest that the pathogenesis of Cushing disease caused by overexpression of heat shock proteins and consequently misregulated GR sensitivity may be overcome pharmacologically with an appropriate HSP90 inhibitor.

  4. Structure, Function and Regulation of the Hsp90 Machinery

    Directory of Open Access Journals (Sweden)

    Jing Li

    2013-06-01

    Full Text Available Heat shock protein 90 (Hsp90 is an ATP-dependent molecular chaperone which is essential in eukaryotes. It is required for the activation and stabilization of a wide variety of client proteins and many of them are involved in important cellular pathways. Since Hsp90 affects numerous physiological processes such as signal transduction, intracellular transport, and protein degradation, it became an interesting target for cancer therapy. Structurally, Hsp90 is a flexible dimeric protein composed of three different domains which adopt structurally distinct conformations. ATP binding triggers directionality in these conformational changes and leads to a more compact state. To achieve its function, Hsp90 works together with a large group of cofactors, termed co-chaperones. Co-chaperones form defined binary or ternary complexes with Hsp90, which facilitate the maturation of client proteins. In addition, posttranslational modifications of Hsp90, such as phosphorylation and acetylation, provide another level of regulation. They influence the conformational cycle, co-chaperone interaction, and inter-domain communications. In this review, we discuss the recent progress made in understanding the Hsp90 machinery.

  5. Antiviral evaluation of an Hsp90 inhibitor, gedunin, against dengue ...

    African Journals Online (AJOL)

    Further, in silico molecular docking data revealed strong interaction of gedunin with the ATP/ADP ... Keywords: Dengue virus replication, Hsp90, Gedunin, Antiviral, Molecular docking ..... Conformational dynamics of the molecular chaperone.

  6. Review: The HSP90 molecular chaperone-an enigmatic ATPase.

    Science.gov (United States)

    Pearl, Laurence H

    2016-08-01

    The HSP90 molecular chaperone is involved in the activation and cellular stabilization of a range of 'client' proteins, of which oncogenic protein kinases and nuclear steroid hormone receptors are of particular biomedical significance. Work over the last two decades has revealed a conformational cycle critical to the biological function of HSP90, coupled to an inherent ATPase activity that is regulated and manipulated by many of the co-chaperones proteins with which it collaborates. Pharmacological inhibition of HSP90 ATPase activity results in degradation of client proteins in vivo, and is a promising target for development of new cancer therapeutics. Despite this, the actual function that HSP90s conformationally-coupled ATPase activity provides in its biological role as a molecular chaperone remains obscure. © 2016 Wiley Periodicals, Inc. Biopolymers 105: 594-607, 2016. © 2016 The Authors. Biopolymers Published by Wiley Periodicals, Inc.

  7. Molecular cloning, sequencing, and expression profiles of heat shock protein 90 (HSP90 in Hyriopsis cumingii exposed to different stressors: Temperature, cadmium and Aeromonas hydrophila

    Directory of Open Access Journals (Sweden)

    Qin Wang

    2017-03-01

    Full Text Available Heat shock protein 90 (HSP90 represents a suite of highly conserved and multi-functional molecular chaperone proteins that play an important role in cellular stress responses. In order to better understand the expression of HSP90 in mollusks, a full-length complementary DNA (cDNA of HSP90 (HcHSP90 was identified in Hyriopsis cumingii. HcHSP90 cDNA was 2659 bp in length, consisting of 3′ and 5′-untranslated regions and an open reading frame of 2187 bp, that encoded a 728 amino acid protein. Homology analyses showed that the HcHSP90 protein was highly conserved and had 5 well-conserved family signatures of HSP90 proteins. HcHSP90 mRNA expressed in various tissues of H. cumingii. The expression level of HcHSP90 was the highest in the digestive gland. In all tissues, with the exception of the digestive gland where it was down-regulated, HcHSP90 mRNA expression was significantly induced by temperature treatments (0, 5, 25, and 35 °C relative to the control (15 °C. Exposure of H. cumingii to different concentrations of cadmium (50, 100, and 200 μg/L, up-regulated HcHSP90 mRNA in the haemolymph and gill but without an obvious dose-dependent response. When H. cumingii were infected with Aeromonas hydrophila, HcHSP90 mRNA expression in the haemolymph was up-regulated and peaked 36 h post-infection, while in the gills it was significantly up-regulated 3 h post-infection in the gills, then remained constant until returning to pre-challenge expression levels at 36 h post-infection. The results show that HcHSP90 expression can be significantly regulated by changes in temperature, cadmium exposure and bacterial infection. We deduced that HSP90 may play an important role in helping H. cumingii to cope with environmental stress.

  8. The Hsp90 co-chaperones Sti1, Aha1, and P23 regulate adaptive responses to antifungal azoles

    Directory of Open Access Journals (Sweden)

    Xiaokui Gu

    2016-10-01

    Full Text Available Heat Shock Protein 90 (Hsp90 is essential for tumor progression in humans and drug resistance in fungi. However, the roles of its many co-chaperones in antifungal resistance are unknown. In this study, by susceptibility test of Neurospora crassa mutants lacking each of 18 Hsp90/Calcineurin system member genes (including 8 Hsp90 co-chaperone genes to antifungal drugs and other stresses, we demonstrate that the Hsp90 co-chaperones Sti1 (Hop1 in yeast, Aha1, and P23 (Sba1 in yeast were required for the basal resistance to antifungal azoles and heat stress. Deletion of any of them resulted in hypersensitivity to azoles and heat. Liquid chromatography–mass spectrometry (LC-MS analysis showed that the toxic sterols eburicol and 14α-methyl-3,6-diol were significantly accumulated in the sti1 and p23 deletion mutants after ketoconazole treatment, which has been shown before to led to cell membrane stress. At the transcriptional level, Aha1, Sti1, and P23 positively regulate responses to ketoconazole stress by erg11 and erg6, key genes in the ergosterol biosynthetic pathway. Aha1, Sti1, and P23 are highly conserved in fungi, and sti1 and p23 deletion also increased the susceptibility to azoles in Fusarium verticillioides. These results indicate that Hsp90-cochaperones Aha1, Sti1, and P23 are critical for the basal azole resistance and could be potential targets for developing new antifungal agents.

  9. Targeting HSP90 and monoclonal protein trafficking modulates the unfolded protein response, chaperone regulation and apoptosis in myeloma cells

    International Nuclear Information System (INIS)

    Born, E J; Hartman, S V; Holstein, S A

    2013-01-01

    Multiple myeloma is characterized by the production of substantial quantities of monoclonal protein. We have previously demonstrated that select inhibitors of the isoprenoid biosynthetic pathway (IBP) induce apoptosis of myeloma cells via inhibition of Rab geranylgeranylation, leading to disruption of monoclonal protein trafficking and induction of the unfolded protein response (UPR) pathway. Heat-shock protein 90 (HSP90) inhibitors disrupt protein folding and are currently under clinical investigation in myeloma. The effects of combining IBP and HSP90 inhibitors on cell death, monoclonal protein trafficking, the UPR and chaperone regulation were investigated in monoclonal protein-producing cells. An enhanced induction of cell death was observed following treatment with IBP and HSP90 inhibitors, which occurred through both ER stress and non-ER stress pathways. The HSP90 inhibitor 17-AAG abrogated the effects of the IBP inhibitors on intracellular monoclonal protein levels and localization as well as induction of the UPR in myeloma cells. Disparate effects on chaperone expression were observed in myeloma vs amyloid light chain cells. Here we demonstrate that the novel strategy of targeting MP trafficking in concert with HSP90 enhances myeloma cell death via a complex modulation of ER stress, UPR, and cell death pathways

  10. Hsp90 inhibitors reduce influenza virus replication in cell culture

    International Nuclear Information System (INIS)

    Chase, Geoffrey; Deng, Tao; Fodor, Ervin; Leung, B.W.; Mayer, Daniel; Schwemmle, Martin; Brownlee, George

    2008-01-01

    The viral RNA polymerase complex of influenza A virus consists of three subunits PB1, PB2 and PA. Recently, the cellular chaperone Hsp90 was shown to play a role in nuclear import and assembly of the trimeric polymerase complex by binding to PB1 and PB2. Here we show that Hsp90 inhibitors, geldanamycin or its derivative 17-AAG, delay the growth of influenza virus in cell culture resulting in a 1-2 log reduction in viral titre early in infection. We suggest that this is caused by the reduced half-life of PB1 and PB2 and inhibition of nuclear import of PB1 and PA which lead to reduction in viral RNP assembly. Hsp90 inhibitors may represent a new class of antiviral compounds against influenza viruses

  11. Hsp90 and hepatobiliary transformation during sea lamprey metamorphosis.

    Science.gov (United States)

    Chung-Davidson, Yu-Wen; Yeh, Chu-Yin; Bussy, Ugo; Li, Ke; Davidson, Peter J; Nanlohy, Kaben G; Brown, C Titus; Whyard, Steven; Li, Weiming

    2015-12-01

    Biliary atresia (BA) is a human infant disease with inflammatory fibrous obstructions in the bile ducts and is the most common cause for pediatric liver transplantation. In contrast, the sea lamprey undergoes developmental BA with transient cholestasis and fibrosis during metamorphosis, but emerges as a fecund adult. Therefore, sea lamprey liver metamorphosis may serve as an etiological model for human BA and provide pivotal information for hepatobiliary transformation and possible therapeutics. We hypothesized that liver metamorphosis in sea lamprey is due to transcriptional reprogramming that dictates cellular remodeling during metamorphosis. We determined global gene expressions in liver at several metamorphic landmark stages by integrating mRNA-Seq and gene ontology analyses, and validated the results with real-time quantitative PCR, histological and immunohistochemical staining. These analyses revealed that gene expressions of protein folding chaperones, membrane transporters and extracellular matrices were altered and shifted during liver metamorphosis. HSP90, important in protein folding and invertebrate metamorphosis, was identified as a candidate key factor during liver metamorphosis in sea lamprey. Blocking HSP90 with geldanamycin facilitated liver metamorphosis and decreased the gene expressions of the rate limiting enzyme for cholesterol biosynthesis, HMGCoA reductase (hmgcr), and bile acid biosynthesis, cyp7a1. Injection of hsp90 siRNA for 4 days altered gene expressions of met, hmgcr, cyp27a1, and slc10a1. Bile acid concentrations were increased while bile duct and gall bladder degeneration was facilitated and synchronized after hsp90 siRNA injection. HSP90 appears to play crucial roles in hepatobiliary transformation during sea lamprey metamorphosis. Sea lamprey is a useful animal model to study postembryonic development and mechanisms for hsp90-induced hepatobiliary transformation.

  12. The HSP90 inhibitor 17-AAG potentiates the antileishmanial activity of the ether lipid edelfosine.

    Science.gov (United States)

    Varela-M, Rubén E; Mollinedo-Gajate, Cristina; Muro, Antonio; Mollinedo, Faustino

    2014-03-01

    HSP90 is an abundant protein in Leishmania parasites that plays a major role in the parasite survival under stress conditions. Here we found that the HSP90 inhibitor 17-AAG (≥100nM 17-AAG) induced cell cycle arrest at G0/G1 in Leishmania infantum and Leishmania panamensis promastigotes, and highly potentiated the induction of cell death by an apoptotic-like process mediated by the ether phospholipid edelfosine (5-20μM). These data suggest that the combined treatment of 17-AAG and edelfosine might be a novel and effective approach of combination therapy in the treatment of leishmaniasis. Copyright © 2013 Elsevier B.V. All rights reserved.

  13. Antiviral evaluation of an Hsp90 inhibitor, gedunin, against dengue ...

    African Journals Online (AJOL)

    Purpose: To evaluate the antiviral potential of a tetranortriterpenoid, gedunin, against dengue virus (DENV) replication by targeting the host chaperone, Hsp90. Methods: The compound, gedunin, was tested against the replication of DENV in vitro using BHK-15 cells transfected with DENV-2 subgenomic replicon. Molecular ...

  14. Cyclophilin 40 facilitates HSP90-mediated RISC assembly in plants.

    Science.gov (United States)

    Iki, Taichiro; Yoshikawa, Manabu; Meshi, Tetsuo; Ishikawa, Masayuki

    2012-01-18

    Posttranscriptional gene silencing is mediated by RNA-induced silencing complexes (RISCs) that contain AGO proteins and single-stranded small RNAs. The assembly of plant AGO1-containing RISCs depends on the molecular chaperone HSP90. Here, we demonstrate that cyclophilin 40 (CYP40), protein phosphatase 5 (PP5), and several other proteins with the tetratricopeptide repeat (TPR) domain associates with AGO1 in an HSP90-dependent manner in extracts of evacuolated tobacco protoplasts (BYL). Intriguingly, CYP40, but not the other TPR proteins, could form a complex with small RNA duplex-bound AGO1. Moreover, CYP40 that was synthesized by in-vitro translation using BYL uniquely facilitated binding of small RNA duplexes to AGO1, and as a result, increased the amount of mature RISCs that could cleave target RNAs. CYP40 was not contained in mature RISCs, indicating that the association is transient. Addition of PP5 or cyclophilin-binding drug cyclosporine A prevented the association of endogenous CYP40 with HSP90-AGO1 complex and inhibited RISC assembly. These results suggest that a complex of AGO1, HSP90, CYP40, and a small RNA duplex is a key intermediate of RISC assembly in plants.

  15. Hsp90: A New Player in DNA Repair?

    Directory of Open Access Journals (Sweden)

    Rosa Pennisi

    2015-10-01

    Full Text Available Heat shock protein 90 (Hsp90 is an evolutionary conserved molecular chaperone that, together with Hsp70 and co-chaperones makes up the Hsp90 chaperone machinery, stabilizing and activating more than 200 proteins, involved in protein homeostasis (i.e., proteostasis, transcriptional regulation, chromatin remodeling, and DNA repair. Cells respond to DNA damage by activating complex DNA damage response (DDR pathways that include: (i cell cycle arrest; (ii transcriptional and post-translational activation of a subset of genes, including those associated with DNA repair; and (iii triggering of programmed cell death. The efficacy of the DDR pathways is influenced by the nuclear levels of DNA repair proteins, which are regulated by balancing between protein synthesis and degradation as well as by nuclear import and export. The inability to respond properly to either DNA damage or to DNA repair leads to genetic instability, which in turn may enhance the rate of cancer development. Multiple components of the DNA double strand breaks repair machinery, including BRCA1, BRCA2, CHK1, DNA-PKcs, FANCA, and the MRE11/RAD50/NBN complex, have been described to be client proteins of Hsp90, which acts as a regulator of the diverse DDR pathways. Inhibition of Hsp90 actions leads to the altered localization and stabilization of DDR proteins after DNA damage and may represent a cell-specific and tumor-selective radiosensibilizer. Here, the role of Hsp90-dependent molecular mechanisms involved in cancer onset and in the maintenance of the genome integrity is discussed and highlighted.

  16. Hsp70/Hsp90 organising protein (hop): beyond interactions with chaperones and prion proteins.

    Science.gov (United States)

    Baindur-Hudson, Swati; Edkins, Adrienne L; Blatch, Gregory L

    2015-01-01

    The Hsp70/Hsp90 organising protein (Hop), also known as stress-inducible protein 1 (STI1), has received considerable attention for diverse cellular functions in both healthy and diseased states. There is extensive evidence that intracellular Hop is a co-chaperone of the major chaperones Hsp70 and Hsp90, playing an important role in the productive folding of Hsp90 client proteins. Consequently, Hop is implicated in a number of key signalling pathways, including aberrant pathways leading to cancer. However, Hop is also secreted and it is now well established that Hop also serves as a receptor for the prion protein, PrP(C). The intracellular and extracellular forms of Hop most likely represent two different isoforms, although the molecular determinants of these divergent functions are yet to be identified. There is also a growing body of research that reports the involvement of Hop in cellular activities that appear independent of either chaperones or PrP(C). While Hop has been shown to have various cellular functions, its biological function remains elusive. However, recent knockout studies in mammals suggest that Hop has an important role in embryonic development. This review provides a critical overview of the latest molecular, cellular and biological research on Hop, critically evaluating its function in healthy systems and how this function is adapted in diseases states.

  17. Jasmonate signalling in Arabidopsis involves SGT1b-HSP70-HSP90 chaperone complexes.

    Science.gov (United States)

    Zhang, Xue-Cheng; Millet, Yves A; Cheng, Zhenyu; Bush, Jenifer; Ausubel, Frederick M

    Plant hormones play pivotal roles in growth, development and stress responses. Although it is essential to our understanding of hormone signalling, how plants maintain a steady state level of hormone receptors is poorly understood. We show that mutation of the Arabidopsis thaliana co-chaperone SGT1b impairs responses to the plant hormones jasmonate, auxin and gibberellic acid, but not brassinolide and abscisic acid, and that SGT1b and its homologue SGT1a are involved in maintaining the steady state levels of the F-box proteins COI1 and TIR1, receptors for jasmonate and auxin, respectively. The association of SGT1b with COI1 is direct and is independent of the Arabidopsis SKP1 protein, ASK1. We further show that COI1 is a client protein of SGT1b-HSP70-HSP90 chaperone complexes and that the complexes function in hormone signalling by stabilizing the COI1 protein. This study extends the SGT1b-HSP90 client protein list and broadens the functional scope of SGT1b-HSP70-HSP90 chaperone complexes.

  18. Stability of the Human Hsp90-p50Cdc37 Chaperone Complex against Nucleotides and Hsp90 Inhibitors, and the Influence of Phosphorylation by Casein Kinase 2

    Directory of Open Access Journals (Sweden)

    Sanne H. Olesen

    2015-01-01

    Full Text Available The molecular chaperone Hsp90 is regulated by co-chaperones such as p50Cdc37, which recruits a wide selection of client protein kinases. Targeted disruption of the Hsp90-p50Cdc37 complex by protein–protein interaction (PPI inhibitors has emerged as an alternative strategy to treat diseases characterized by aberrant Hsp90 activity. Using isothermal microcalorimetry, ELISA and GST-pull down assays we evaluated reported Hsp90 inhibitors and nucleotides for their ability to inhibit formation of the human Hsp90β-p50Cdc37 complex, reconstituted in vitro from full-length proteins. Hsp90 inhibitors, including the proposed PPI inhibitors gedunin and H2-gamendazole, did not affect the interaction of Hsp90 with p50Cdc37 in vitro. Phosphorylation of Hsp90 and p50Cdc37 by casein kinase 2 (CK2 did not alter the thermodynamic signature of complex formation. However, the phosphorylated complex was vulnerable to disruption by ADP (IC50 = 32 µM, while ATP, AMPPNP and Hsp90 inhibitors remained largely ineffective. The differential inhibitory activity of ADP suggests that phosphorylation by CK2 primes the complex for dissociation in response to a drop in ATP/ADP levels. The approach applied herein provides robust assays for a comprehensive biochemical evaluation of potential effectors of the Hsp90-p50Cdc37 complex, such as phosphorylation by a kinase or the interaction with small molecule ligands.

  19. Systemic and mucosal immunization with Candida albicans hsp90 elicits hsp90-specific humoral response in vaginal mucosa which is further enhanced during experimental vaginal candidiasis.

    Science.gov (United States)

    Raska, Milan; Belakova, Jana; Horynova, Milada; Krupka, Michal; Novotny, Jiri; Sebestova, Martina; Weigl, Evzen

    2008-08-01

    The Candida albicans heat shock protein 90 kDa (hsp90-CA) is an important target for protective antibodies in disseminated candidiasis of experimental mice and humans. Hsp90-CA is present in the cell wall of Candida pseudohyphae or hyphae--typical pathogenic morphotypes in both mucosal and systemic Candida infections. However, the potential protective effects of hsp90-CA-specific antibodies in vaginal candidiasis has not yet been reported. In the present study we used various vaccine formulations (recombinant hsp90-CA protein and hsp90-CA-encoding DNA vaccine) and routes of administration (intradermal, intranasal, and intravenous) to induce both hsp90-CA-specific systemic and vaginal mucosa immune responses in experimental BALB/c mice. The results showed that intradermal recombinant hsp90-CA protein priming, followed by intranasal or intradermal recombinant hsp90-CA protein boosting induced significant increases in both serum and vaginal hsp90-CA-specific IgG and IgA antibodies compared to the control group, as well as enhanced hsp90-CA-specific splenocyte responses in vitro. In the intradermally boosted group, subsequent experimental vaginal Candida infection induced additional increases in the hsp90-CA specific IgG isotype, suggesting that Candida has the ability to induce a local hsp90-specific antibody (IgG) response during vulvovaginal candidiasis. Further work is required to elucidate the importance of immunity to highly conserved antigens during infection of the human female reproductive tract where a balance between immunity to and tolerance for commonly antigens such as hsp90 is necessary for the maintenance of fertility.

  20. The HSP90 Inhibitor Ganetespib Radiosensitizes Human Lung Adenocarcinoma Cells

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    Roberto Gomez-Casal

    2015-05-01

    Full Text Available The molecular chaperone HSP90 is involved in stabilization and function of multiple client proteins, many of which represent important oncogenic drivers in NSCLC. Utilization of HSP90 inhibitors as radiosensitizing agents is a promising approach. The antitumor activity of ganetespib, HSP90 inhibitor, was evaluated in human lung adenocarcinoma (AC cells for its ability to potentiate the effects of IR treatment in both in vitro and in vivo. The cytotoxic effects of ganetespib included; G2/M cell cycle arrest, inhibition of DNA repair, apoptosis induction, and promotion of senescence. All of these antitumor effects were both concentration- and time-dependent. Both pretreatment and post-radiation treatment with ganetespib at low nanomolar concentrations induced radiosensitization in lung AC cells in vitro. Ganetespib may impart radiosensitization through multiple mechanisms: such as down regulation of the PI3K/Akt pathway; diminished DNA repair capacity and promotion of cellular senescence. In vivo, ganetespib reduced growth of T2821 tumor xenografts in mice and sensitized tumors to IR. Tumor irradiation led to dramatic upregulation of β-catenin expression in tumor tissues, an effect that was mitigated in T2821 xenografts when ganetespib was combined with IR treatments. These data highlight the promise of combining ganetespib with IR therapies in the treatment of AC lung tumors.

  1. Whey protein hydrolysate enhances HSP90 but does not alter HSP60 and HSP25 in skeletal muscle of rats.

    Directory of Open Access Journals (Sweden)

    Carolina Soares Moura

    Full Text Available Whey protein hydrolysate (WPH intake has shown to increase HSP70 expression. The aim of the present study was to investigate whether WPH intake would also influences HSP90, HSP60 and HSP25 expression, as well as associated parameters. Forty-eight male Wistar rats were divided into sedentary (unstressed and exercised (stressed groups, and were fed with three different sources of protein: whey protein (WP, whey protein hydrolysate (WPH and casein (CAS as a control, based on the AIN93G diet for 3 weeks. WPH intake increased HSP90 expression in both sedentary and exercised animals compared to WP or CAS, however no alteration was found from exercise or diet to HSP60 or HSP25. Co-chaperone Aha1 and p-HSF1 were also increased in the exercised animals fed with WPH in comparison with WP or CAS, consistent with enhanced HSP90 expression. VEGF and p-AKT were increased in the WPH exercised group. No alteration was found in BCKDH, PI3-Kinase (p85, GFAT, OGT or PGC for diet or exercise. The antioxidant system GPx, catalase and SOD showed different responses to diet and exercise. The data indicate that WPH intake enhanced factors related to cell survival, such as HSP90 and VEGF, but does not alter HSP60 or HSP25 in rat skeletal muscle.

  2. Targeting Plasmodium falciparum Hsp90: Towards Reversing Antimalarial Resistance

    Directory of Open Access Journals (Sweden)

    Dea Shahinas

    2013-02-01

    Full Text Available Malaria continues to exact a great human toll in tropical settings. Antimalarial resistance is rife and the parasite inexorably develops mechanisms to outwit our best drugs, including the now first-line choice, artesunate. Novel strategies to circumvent resistance are needed. Here we detail drug development focusing on heat shock protein 90 and its central role as a chaperone. A growing body of evidence supports the role for Hsp90 inhibitors as adjunctive drugs able to restore susceptibility to traditionally efficacious compounds like chloroquine.

  3. The association of SNPs in Hsp90β gene 5' flanking region with thermo tolerance traits and tissue mRNA expression in two chicken breeds.

    Science.gov (United States)

    Chen, Zhuo-Yu; Gan, Jian-Kang; Xiao, Xiong; Jiang, Li-Yan; Zhang, Xi-Quan; Luo, Qing-Bin

    2013-09-01

    Thermo stress induces heat shock proteins (HSPs) expression and HSP90 family is one of them that has been reported to involve in cellular protection against heat stress. But whether there is any association of genetic variation in the Hsp90β gene in chicken with thermo tolerance is still unknown. Direct sequencing was used to detect possible SNPs in Hsp90β gene 5' flanking region in 3 chicken breeds (n = 663). Six mutations, among which 2 SNPs were chosen and genotypes were analyzed with PCR-RFLP method, were found in Hsp90β gene in these 3 chicken breeds. Association analysis indicated that SNP of C.-141G>A in the 5' flanking region of the Hsp90β gene in chicken had some effect on thermo tolerance traits, which may be a potential molecular marker of thermo tolerance, and the genotype GG was the thermo tolerance genotype. Hsp90β gene mRNA expression in different tissues detected by quantitative real-time PCR assay were demonstrated to be tissue dependent, implying that different tissues have distinct sensibilities to thermo stress. Besides, it was shown time specific and varieties differences. The expression of Hsp90β mRNA in Lingshan chickens in some tissues including heart, liver, brain and spleen were significantly higher or lower than that of White Recessive Rock (WRR). In this study, we presume that these mutations could be used in marker assisted selection for anti-heat stress chickens in our breeding program, and WRR were vulnerable to tropical thermo stress whereas Lingshan chickens were well adapted.

  4. Spatio-temporal regulation of Hsp90-ligand complex leads to immune activation.

    Directory of Open Access Journals (Sweden)

    Yasuaki eTamura

    2016-05-01

    Full Text Available Hsp90 is the most abundant cytosolic HSP and is known to act as a molecular chaperone. We found that an Hsp90-cancer antigen peptide complex was efficiently cross-presented by human monocyte-derived dendritic cells and induced peptide-specific cytotoxic T lymphocytes. Furthermore, we observed that the internalized Hsp90-peptide complex was strictly sorted to the Rab5+, EEA1+ static early endosome and the Hsp90-chaperoned peptide was processed and bound to MHC class I molecules through a endosome-recycling pathway. We also found that extracellular Hsp90 complexed with CpG-A or self-DNA stimulates production of a large amount of IFN-α from pDCs via static early endosome targeting. Thus, extracellular Hsp90 can target the antigen or nucleic acid to a static early endosome by spatio-temporal regulation. Moreover, we showed that Hsp90 associates with and delivers TLR7/9 from the ER to early endosomes for ligand recognition. Hsp90 inhibitor, geldanamycin derivative inhibited the Hsp90 association with TLR7/9, resulting in inhibition IFN-α production, leading to improvement of SLE symptoms. Interstingly, we observed that serum Hsp90 is clearly increased in patients with active SLE compared with that in patients with inactive disease. Serum Hsp90 detected in SLE patients binds to self-DNA and/or anti-DNA Ab, thus leading to stimulation of pDCs to produce IFN-α. Thus, Hsp90 plays a crucial role in the pathogenesis of SLE and that an Hsp90 inhibitor will therefore provide a new therapeutic approach to SLE and other nucleic acid-related autoimmune diseases. We will discuss how spatio-temporal regulation of Hsp90-ligand complexes within antigen-presenting cells affects the innate immunity and adaptive immunity.

  5. Novel Hsp90 inhibitor NVP-AUY922 radiosensitizes prostate cancer cells.

    Science.gov (United States)

    Gandhi, Nishant; Wild, Aaron T; Chettiar, Sivarajan T; Aziz, Khaled; Kato, Yoshinori; Gajula, Rajendra P; Williams, Russell D; Cades, Jessica A; Annadanam, Anvesh; Song, Danny; Zhang, Yonggang; Hales, Russell K; Herman, Joseph M; Armour, Elwood; DeWeese, Theodore L; Schaeffer, Edward M; Tran, Phuoc T

    2013-04-01

    Outcomes for poor-risk localized prostate cancers treated with radiation are still insufficient. Targeting the "non-oncogene" addiction or stress response machinery is an appealing strategy for cancer therapeutics. Heat-shock-protein-90 (Hsp90), an integral member of this machinery, is a molecular chaperone required for energy-driven stabilization and selective degradation of misfolded "client" proteins, that is commonly overexpressed in tumor cells. Hsp90 client proteins include critical components of pathways implicated in prostate cancer cell survival and radioresistance, such as androgen receptor signaling and the PI3K-Akt-mTOR pathway. We examined the effects of a novel non-geldanamycin Hsp90 inhibitor, AUY922, combined with radiation (RT) on two prostate cancer cell lines, Myc-CaP and PC3, using in vitro assays for clonogenic survival, apoptosis, cell cycle distribution, γ-H2AX foci kinetics and client protein expression in pathways important for prostate cancer survival and radioresistance. We then evaluated tumor growth delay and effects of the combined treatment (RT-AUY922) on the PI3K-Akt-mTOR and AR pathways in a hind-flank tumor graft model. We observed that AUY922 caused supra-additive radiosensitization in both cell lines at low nanomolar doses with enhancement ratios between 1.4-1.7 (p < 0.01). RT-AUY922 increased apoptotic cell death compared with either therapy alone, induced G 2-M arrest and produced marked changes in client protein expression. These results were confirmed in vivo, where RT-AUY922 combination therapy produced supra-additive tumor growth delay compared with either therapy by itself in Myc-CaP and PC3 tumor grafts (both p < 0.0001). Our data suggest that combined RT-AUY922 therapy exhibits promising activity against prostate cancer cells, which should be investigated in clinical studies.

  6. HSP90 inhibitor 17-AAG selectively eradicates lymphoma stem cells.

    Science.gov (United States)

    Newman, Bryan; Liu, Yan; Lee, Hsiu-Fang; Sun, Duxin; Wang, Yin

    2012-09-01

    Cancer stem cells (CSC; also called tumor-initiating cells) comprise tumor cell subpopulations that preserve the properties of quiescence, self-renewal, and differentiation of normal stem cells. In addition, CSCs are therapeutically important because of their key contributions toward drug resistance. The hypoxia-inducible transcription factor HIF1α is critical for CSC maintenance in mouse lymphoma. In this study, we showed that low concentrations of the HSP90 inhibitor 17-AAG eliminate lymphoma CSCs in vitro and in vivo by disrupting the transcriptional function of HIF1α, a client protein of HSP90. 17-AAG preferentially induced apoptosis and eliminated the colony formation capacity of mouse lymphoma CSCs and human acute myeloid leukemia (AML) CSCs. However, low concentrations of 17-AAG failed to eliminate highly proliferative lymphoma and AML cells (non-CSCs), in which the AKT-GSK3 signaling pathway is constitutively active. The heat shock transcription factor HSF1 is highly expressed in non-CSCs, but it was weakly expressed in lymphoma CSCs. However, siRNA-mediated attenuation of HSF1 abrogated the colony formation ability of both lymphoma and AML CSCs. This study supports the use of 17-AAG as a CSC targeting agent and, in addition, shows that HSF1 is an important target for elimination of both CSCs and non-CSCs in cancer. ©2012 AACR.

  7. Hsp90 inhibitor 17-allylamino-17-demethoxygeldanamycin inhibits the proliferation of ARPE-19 cells

    Directory of Open Access Journals (Sweden)

    Wang Lin

    2010-04-01

    Full Text Available Abstract Background The antiproliferative effect of the Hsp90 inhibitor 17-AAG (17-allylamino-17-demethoxygeldanamycin on human retinal pigment epithelial cells is investigated. Methods MTT and flow cytometry were used to study the antiproliferative effects of the 17-AAG treatment of ARPE-19 cells. 2D gel electrophoresis (2-DE and mass spectrometry were applied to detect the altered expression of proteins, which was verified by real-time PCR. Gene Ontology analysis and Ingenuity Pathway Analysis (IPA were utilized to analyze the signaling pathways, cellular location, function, and network connections of the identified proteins. And SOD assay was employed to confirm the analysis. Results 17-AAG suppressed the proliferation of ARPE-19 cells by inducing cell cycle arrest and apoptosis. Proteomic analysis revealed that the expression of 94 proteins was altered by a factor of more than 1.5 following exposure to 17-AAG. Of these 94, 87 proteins were identified. Real-time PCR results indicated that Hsp90 and Hsp70, which were not identified by proteomic analysis, were both upregulated upon 17-AAG treatment. IPA revealed that most of the proteins have functions that are related to oxidative stress, as verified by SOD assay, while canonical pathway analysis revealed glycolysis/gluconeogenesis. Conclusions 17-AAG suppressed the proliferation of ARPE-19 cells by inducing cell cycle arrest and apoptosis, and possibly by oxidative stress.

  8. 真蛸热休克蛋白90基因(HSP90)的克隆及表达%Molecular cloning and feature analysis of heat shock protein 90 ( HSP90 ) from Octopus vulgaris

    Institute of Scientific and Technical Information of China (English)

    孙田田; 苏永全; 洪婧妮; 邬阳; 张曼; 毛勇

    2012-01-01

    为深入了解真蛸热应激响应机制,实验以真蛸的应激蛋白为研究对象,借助同源扩增和cDNA末端快速扩增(rapid amplification of cDNA ends,RACE)技术获得真蛸热休克蛋白90( HSP90)的cDNA全序列(2 709 bp),它包含119 bp的5′非编码区(untranslated regions,UTR)、454 bp的3′UTR和2 136 bp的开放阅读框(opening reading frame,ORF),ORF共编码71 1个氨基酸,推算其分子量为81.5 ku,理论等电点为5.09.同源分析显示,所推导的氨基酸序列高度保守,并且含有HSP90家族特有的5个保守信号序列区域.实时荧光定量PCR分析表明,该基因在所有选取的组织中均有表达,肝组织中表达量最高.%Octopus vulgaris,which owns a number of unique stress protection mechanisms is considered to be the most intelligent species of all invertebrates. A 2 709 bp full-length cDNA sequence of heat shock protein 90 ( HSP90 ) gene from O. vulgaris was obtained by homologous amplification and rapid amplification of cDNA ends (RACE). It consisted of a 119 bp 5'untranslated region (UTR) ,a2 136 bp open reading frame(ORF)and a 454 bp 3'UTR. The inferred amino acids sequence was composed of 711 amino acids, whose molecular weight and isoelectric point were 81. 5 ku and 5. 09, respectively. Homologous analysis showed the inferred amino acids sequence was highly conserved and contained five classical HSP90 signature sequences. Real-time quantitative PCR ( RT-PCR) results displayed HSP90 expressed in every tissues chosen, with the most in liver. This study which was the first time to obtain HSP90 gene from Cephalopod played an important role in studying stress mechanisms on the most intelligent invertebrate.

  9. Cryptic variation in morphological evolution: HSP90 as a capacitor for loss of eyes in cavefish.

    Science.gov (United States)

    Rohner, Nicolas; Jarosz, Dan F; Kowalko, Johanna E; Yoshizawa, Masato; Jeffery, William R; Borowsky, Richard L; Lindquist, Susan; Tabin, Clifford J

    2013-12-13

    In the process of morphological evolution, the extent to which cryptic, preexisting variation provides a substrate for natural selection has been controversial. We provide evidence that heat shock protein 90 (HSP90) phenotypically masks standing eye-size variation in surface populations of the cavefish Astyanax mexicanus. This variation is exposed by HSP90 inhibition and can be selected for, ultimately yielding a reduced-eye phenotype even in the presence of full HSP90 activity. Raising surface fish under conditions found in caves taxes the HSP90 system, unmasking the same phenotypic variation as does direct inhibition of HSP90. These results suggest that cryptic variation played a role in the evolution of eye loss in cavefish and provide the first evidence for HSP90 as a capacitor for morphological evolution in a natural setting.

  10. Selection Transforms the Landscape of Genetic Variation Interacting with Hsp90.

    Science.gov (United States)

    Geiler-Samerotte, Kerry A; Zhu, Yuan O; Goulet, Benjamin E; Hall, David W; Siegal, Mark L

    2016-10-01

    The protein-folding chaperone Hsp90 has been proposed to buffer the phenotypic effects of mutations. The potential for Hsp90 and other putative buffers to increase robustness to mutation has had major impact on disease models, quantitative genetics, and evolutionary theory. But Hsp90 sometimes contradicts expectations for a buffer by potentiating rapid phenotypic changes that would otherwise not occur. Here, we quantify Hsp90's ability to buffer or potentiate (i.e., diminish or enhance) the effects of genetic variation on single-cell morphological features in budding yeast. We corroborate reports that Hsp90 tends to buffer the effects of standing genetic variation in natural populations. However, we demonstrate that Hsp90 tends to have the opposite effect on genetic variation that has experienced reduced selection pressure. Specifically, Hsp90 tends to enhance, rather than diminish, the effects of spontaneous mutations and recombinations. This result implies that Hsp90 does not make phenotypes more robust to the effects of genetic perturbation. Instead, natural selection preferentially allows buffered alleles to persist and thereby creates the false impression that Hsp90 confers greater robustness.

  11. The HSP90 inhibitor 17-AAG exhibits potent antitumor activity for pheochromocytoma in a xenograft model.

    Science.gov (United States)

    Xu, Yunze; Zhu, Qi; Chen, Dongning; Shen, Zhoujun; Wang, Weiqing; Ning, Guang; Zhu, Yu

    2015-07-01

    This study aims to investigate the effect of heat shock protein 90 (HSP90) inhibitor 17-allylamino-17-demethoxygeldanamycin (17-AAG) in the malignant pheochromocytoma using a xenograft mouse model. Treatment with 17-AAG induced a marked reduction in the volume and weight of PC12 pheochromocytoma cell tumor xenografts in mice. Furthermore, 17-AAG also significantly inhibited the expression of HSP90 and its client proteins. Our results validated HSP90 as an important target in pheochromocytoma and provided rationale for the testing of HSP90 inhibitors as a promising therapeutic agent in the antitumor therapy of pheochromocytoma.

  12. Plant Hsp90 Proteins Interact with B-Cells and Stimulate Their Proliferation

    Science.gov (United States)

    Corigliano, Mariana G.; Maglioco, Andrea; Laguía Becher, Melina; Goldman, Alejandra; Martín, Valentina; Angel, Sergio O.; Clemente, Marina

    2011-01-01

    Background The molecular chaperone heat shock protein 90 (Hsp90) plays an important role in folding stabilization and activation of client proteins. Besides, Hsp90 of mammals and mammalian pathogens displays immunostimulatory properties. Here, we investigated the role of plant-derived Hsp90s as B-cell mitogens by measuring their proliferative responses in vitro. Methodology Plant cytosolic Hsp90 isoforms from Arabidopsis thaliana (AtHsp81.2) and Nicotiana benthamiana (NbHsp90.3) were expressed in E. coli. Over-expression of recombinant plant Hsp90s (rpHsp90s) was confirmed by SDS-PAGE and western blot using and anti-AtHsp81.2 polyclonal anti-body. Both recombinant proteins were purified by Ni-NTA affinity chromatography and their identity confirmed by MALDI-TOF-TOF. Recombinant AtHsp81.2 and NbHsp90.3 proteins induced prominent proliferative responses in spleen cells form BALB/c mice. Polymyxin-B, a potent inhibitor of lipopolysaccharide (LPS), did not eliminate the rpHsp90-induced proliferation. In addition, in vitro incubation of spleen cells with rpHsp90 led to the expansion of CD19-bearing populations, suggesting a direct effect of these proteins on B lymphocytes. This effect was confirmed by immunofluorescence analysis, where a direct binding of rpHsp90 to B- but not to T-cells was observed in cells from BALB/c and C3H/HeN mice. Finally, we examined the involvement of Toll Like Receptor 4 (TLR4) molecules in the rpHsp90s induction of B-cell proliferation. Spleen cells from C3H/HeJ mice, which carry a point mutation in the cytoplasmic region of TLR4, responded poorly to prAtHsp90. However, the interaction between rpHsp90 and B-cells from C3H/HeJ mice was not altered, suggesting that the mutation on TLR4 would be affecting the signal cascade but not the rpHsp90-TLR4 receptor interaction. Conclusions Our results show for the first time that spleen cell proliferation can be stimulated by a non-pathogen-derived Hsp90. Furthermore, our data provide a new example of

  13. Epigallocatechin-3-gallate suppresses the expression of HSP70 and HSP90 and exhibits anti-tumor activity in vitro and in vivo

    International Nuclear Information System (INIS)

    Tran, Phan LCHB; Kim, Soo-A; Choi, Hong Seok; Yoon, Jung-Hoon; Ahn, Sang-Gun

    2010-01-01

    Epigallocatechin-3-gallate (EGCG), one of the major catechins in green tea, is a potential chemopreventive agent for various cancers. The aim of this study was to examine the effect of EGCG on the expression of heat shock proteins (HSPs) and tumor suppression. Cell colony formation was evaluated by a soft agar assay. Transcriptional activity of HSP70 and HSP90 was determined by luciferase reporter assay. An EGCG-HSPs complex was prepared using EGCG attached to the cyanogen bromide (CNBr)-activated Sepharose 4B. In vivo effect of EGCG on tumor growth was examined in a xenograft model. Treatment with EGCG decreased cell proliferation and colony formation of MCF-7 human breast cancer cells. EGCG specifically inhibited the expression of HSP70 and HSP90 by inhibiting the promoter activity of HSP70 and HSP90. Pretreatment with EGCG increased the stress sensitivity of MCF-7 cells upon heat shock (44°C for 1 h) or oxidative stress (H 2 O 2 , 500 μM for 24 h). Moreover, treatment with EGCG (10 mg/kg) in a xenograft model resulted in delayed tumor incidence and reduced tumor size, as well as the inhibition of HSP70 and HSP90 expression. Overall, these findings demonstrate that HSP70 and HSP90 are potent molecular targets of EGCG and suggest EGCG as a drug candidate for the treatment of human cancer

  14. Decreased Hsp90 expression in infiltrative lobular carcinoma: an immunohistochemical study

    International Nuclear Information System (INIS)

    Zagouri, Flora; Patsouris, Effstratios; Zografos, George; Sergentanis, Theodoros; Nonni, Afrodite; Papadimitriou, Christos; Pazaiti, Anastasia; Michalopoulos, Nikolaos V; Safioleas, Panagiotis; Lazaris, Andreas; Theodoropoulos, George

    2010-01-01

    Elevated Hsp90 expression has been documented in breast ductal carcinomas, whereas decreased Hsp90 expression has been reported in precursor lobular lesions. This study aims to assess Hsp90 expression in infiltrative lobular carcinomas of the breast. Tissue specimens were taken from 32 patients with infiltrative lobular carcinoma. Immunohistochemical assessment of Hsp90 was performed both in the lesion and the adjacent normal breast ducts and lobules; the latter serving as control. Concerning Hsp90 assessment: i) the percentage of positive cells and ii) the intensity were separately analyzed. Subsequently, the Allred score was adopted and calculated. The intensity was treated as an ordinal variable-score (0: negative, low: 1, moderate: 2, high: 3). Statistical analysis followed. All infiltrative lobular carcinoma foci mainly presented with a positive cytoplasmic immunoreaction for Hsp90. Compared to the adjacent normal ducts and lobules, infiltrative lobular carcinoma exhibited a statistically significant decrease in Hsp90 expression, both in terms of Hsp90 positive cells (%) and Allred score (74.2 ± 11.2 vs. 59.1 ± 14.2 p = 0.0001; 7.00 ± 0.95 vs. 6.22 ± 1.01, p = 0.007, Wilcoxon matched-pairs signed-ranks test). Concerning the intensity of Hsp90 immunostaining only a marginal decrease was noted (2.16 ± 0.68 vs. 1.84 ± 0.63, p = 0.087, Wilcoxon matched-pairs signed-ranks test). ILC lesions seem to exhibit decreased Hsp90 expression, a finding contrary to what might have been expected, given that high Hsp90 expression is a trait of invasive ductal carcinomas

  15. Decreased Hsp90 expression in infiltrative lobular carcinoma: an immunohistochemical study

    Directory of Open Access Journals (Sweden)

    Zagouri Flora

    2010-08-01

    Full Text Available Abstract Background Elevated Hsp90 expression has been documented in breast ductal carcinomas, whereas decreased Hsp90 expression has been reported in precursor lobular lesions. This study aims to assess Hsp90 expression in infiltrative lobular carcinomas of the breast. Methods Tissue specimens were taken from 32 patients with infiltrative lobular carcinoma. Immunohistochemical assessment of Hsp90 was performed both in the lesion and the adjacent normal breast ducts and lobules; the latter serving as control. Concerning Hsp90 assessment: i the percentage of positive cells and ii the intensity were separately analyzed. Subsequently, the Allred score was adopted and calculated. The intensity was treated as an ordinal variable-score (0: negative, low: 1, moderate: 2, high: 3. Statistical analysis followed. Results All infiltrative lobular carcinoma foci mainly presented with a positive cytoplasmic immunoreaction for Hsp90. Compared to the adjacent normal ducts and lobules, infiltrative lobular carcinoma exhibited a statistically significant decrease in Hsp90 expression, both in terms of Hsp90 positive cells (% and Allred score (74.2 ± 11.2 vs. 59.1 ± 14.2 p = 0.0001; 7.00 ± 0.95 vs. 6.22 ± 1.01, p = 0.007, Wilcoxon matched-pairs signed-ranks test. Concerning the intensity of Hsp90 immunostaining only a marginal decrease was noted (2.16 ± 0.68 vs. 1.84 ± 0.63, p = 0.087, Wilcoxon matched-pairs signed-ranks test. Conclusion ILC lesions seem to exhibit decreased Hsp90 expression, a finding contrary to what might have been expected, given that high Hsp90 expression is a trait of invasive ductal carcinomas.

  16. HDAC6 inhibition enhances 17-AAG--mediated abrogation of hsp90 chaperone function in human leukemia cells.

    Science.gov (United States)

    Rao, Rekha; Fiskus, Warren; Yang, Yonghua; Lee, Pearl; Joshi, Rajeshree; Fernandez, Pravina; Mandawat, Aditya; Atadja, Peter; Bradner, James E; Bhalla, Kapil

    2008-09-01

    Histone deacetylase 6 (HDAC6) is a heat shock protein 90 (hsp90) deacetylase. Treatment with pan-HDAC inhibitors or depletion of HDAC6 by siRNA induces hyperacetylation and inhibits ATP binding and chaperone function of hsp90. Treatment with 17-allylamino-demothoxy geldanamycin (17-AAG) also inhibits ATP binding and chaperone function of hsp90, resulting in polyubiquitylation and proteasomal degradation of hsp90 client proteins. In this study, we determined the effect of hsp90 hyperacetylation on the anti-hsp90 and antileukemia activity of 17-AAG. Hyperacetylation of hsp90 increased its binding to 17-AAG, as well as enhanced 17-AAG-mediated attenuation of ATP and the cochaperone p23 binding to hsp90. Notably, treatment with 17-AAG alone also reduced HDAC6 binding to hsp90 and induced hyperacetylation of hsp90. This promoted the proteasomal degradation of HDAC6. Cotreatment with 17-AAG and siRNA to HDAC6 induced more inhibition of hsp90 chaperone function and depletion of BCR-ABL and c-Raf than treatment with either agent alone. In addition, cotreatment with 17-AAG and tubacin augmented the loss of survival of K562 cells and viability of primary acute myeloid leukemia (AML) and chronic myeloid leukemia (CML) samples. These findings demonstrate that HDAC6 is an hsp90 client protein and hyperacetylation of hsp90 augments the anti-hsp90 and antileukemia effects of 17-AAG.

  17. Effect of tributyltin chloride (TBT-Cl) exposure on expression of HSP90β1 in the river pufferfish (Takifugu obscurus): Evidences for its immunologic function involving in exploring process.

    Science.gov (United States)

    Dong-Po, Xu; Di-An, Fang; Chang-Sheng, Zhao; Shu-Lun, Jiang; Hao-Yuan, Hu

    2018-08-05

    HSP90β1 (known as glyco-protein 96, GP96) is a vital endoplasmic reticulum (ER) depended chaperonin among the HSPs (heat shock proteins) family. Furthermore, it always processes and presents antigen of the tumor and keeps balance for the intracellular environment. In the present study, we explored the effect of tributyltin chloride (TBT-Cl) exposure on HSP90β1 expression in river pufferfish, Takifugu obscurus. The full length of To-HSP90β1 was gained with 2775 bp in length, with an ORF (open reading frame) encoding an 803 aa polypeptide. A phylogenetic tree was constructed and showed the close relationship to other fish species. The HSP90β1 mRNA transcript was expressed in all tissues investigated with higher level in the gill and liver. After the acute and chronic exposure of TBT-Cl, the To-HSP90β1 mRNA transcript significantly was up-regulated in gills. Moreover, the histology study indicated the different injury degree of TBT-Cl in liver and gill. Immunohistochemistry (IHC) staining results implied the cytoplasm reorganization after TBT-Cl stress and the function of immunoregulation for To-HSP90β1 to TBT-Cl exposure. All the results indicated that HSP90β1 may be involved in the resistance to the invasion of TBT-Cl for keeping autoimmune homeostasis. Copyright © 2018. Published by Elsevier B.V.

  18. The Effects of Hsp90α1 Mutations on Myosin Thick Filament Organization.

    Science.gov (United States)

    He, Qiuxia; Liu, Kechun; Tian, Zhenjun; Du, Shao Jun

    2015-01-01

    Heat shock protein 90α plays a key role in myosin folding and thick filament assembly in muscle cells. To assess the structure and function of Hsp90α and its potential regulation by post-translational modification, we developed a combined knockdown and rescue assay in zebrafish embryos to systematically analyze the effects of various mutations on Hsp90α function in myosin thick filament organization. DNA constructs expressing the Hsp90α1 mutants with altered putative ATP binding, phosphorylation, acetylation or methylation sites were co-injected with Hsp90α1 specific morpholino into zebrafish embryos. Myosin thick filament organization was analyzed in skeletal muscles of the injected embryos by immunostaining. The results showed that mutating the conserved D90 residue in the Hsp90α1 ATP binding domain abolished its function in thick filament organization. In addition, phosphorylation mimicking mutations of T33D, T33E and T87E compromised Hsp90α1 function in myosin thick filament organization. Similarly, K287Q acetylation mimicking mutation repressed Hsp90α1 function in myosin thick filament organization. In contrast, K206R and K608R hypomethylation mimicking mutations had not effect on Hsp90α1 function in thick filament organization. Given that T33 and T87 are highly conserved residues involved post-translational modification (PTM) in yeast, mouse and human Hsp90 proteins, data from this study could indicate that Hsp90α1 function in myosin thick filament organization is potentially regulated by PTMs involving phosphorylation and acetylation.

  19. A comparative examination of cortisol effects on muscle myostatin and HSP90 gene expression in salmonids.

    Science.gov (United States)

    Galt, Nicholas J; McCormick, Stephen D; Froehlich, Jacob Michael; Biga, Peggy R

    2016-10-01

    Cortisol, the primary corticosteroid in teleost fishes, is released in response to stressors to elicit local functions, however little is understood regarding muscle-specific responses to cortisol in these fishes. In mammals, glucocorticoids strongly regulate the muscle growth inhibitor, myostatin, via glucocorticoid response elements (GREs) leading to muscle atrophy. Bioinformatics methods suggest that this regulatory mechanism is conserved among vertebrates, however recent evidence suggests some fishes exhibit divergent regulation. Therefore, the aim of this study was to evaluate the conserved actions of cortisol on myostatin and hsp90 expression to determine if variations in cortisol interactions have emerged in salmonid species. Representative salmonids; Chinook salmon (Oncorhynchus tshawytscha), cutthroat trout (Oncorhynchus clarki), brook trout (Salvelinus fontinalis), and Atlantic salmon (Salmo salar); were injected intraperitoneally with a cortisol implant (50μg/g body weight) and muscle gene expression was quantified after 48h. Plasma glucose and cortisol levels were significantly elevated by cortisol in all species, demonstrating physiological effectiveness of the treatment. HSP90 mRNA levels were elevated by cortisol in brook trout, Chinook salmon, and Atlantic salmon, but were decreased in cutthroat trout. Myostatin mRNA levels were affected in a species, tissue (muscle type), and paralog specific manner. Cortisol treatment increased myostatin expression in brook trout (Salvelinus) and Atlantic salmon (Salmo), but not in Chinook salmon (Oncorhynchus) or cutthroat trout (Oncorhynchus). Interestingly, the VC alone increased myostatin mRNA expression in Chinook and Atlantic salmon, while the addition of cortisol blocked the response. Taken together, these results suggest that cortisol affects muscle-specific gene expression in species-specific manners, with unique Oncorhynchus-specific divergence observed, that are not predictive solely based upon

  20. Synthesis and Evaluation of the Tumor Cell Growth Inhibitory Potential of New Putative HSP90 Inhibitors.

    Science.gov (United States)

    Bizarro, Ana; Sousa, Diana; Lima, Raquel T; Musso, Loana; Cincinelli, Raffaella; Zuco, Vantina; De Cesare, Michelandrea; Dallavalle, Sabrina; Vasconcelos, M Helena

    2018-02-13

    Heat shock protein 90 (HSP90) is a well-known target for cancer therapy. In a previous work, some of us have reported a series of 3-aryl-naphtho[2,3- d ]isoxazole-4,9-diones as inhibitors of HSP90. In the present work, various compounds with new chromenopyridinone and thiochromenopyridinone scaffolds were synthesized as potential HSP90 inhibitors. Their binding affinity to HSP90 was studied in vitro. Selected compounds ( 5 and 8 ) were further studied in various tumor cell lines regarding their potential to cause cell growth inhibition, alter the cell cycle profile, inhibit proliferation, and induce apoptosis. Their effect on HSP90 client protein levels was also confirmed in two cell lines. Finally, the antitumor activity of compound 8 was studied in A431 squamous cell carcinoma xenografts in nude mice. Our results indicated that treatment with compounds 5 and 8 decreased the proliferation of tumor cell lines and compound 8 induced apoptosis. In addition, these two compounds were able to downregulate selected proteins known as "clients" of HSP90. Finally, treatment of xenografted mice with compound 5 resulted in a considerable dose-dependent inhibition of tumor growth. Our results show that two new compounds with a chromenopyridinone and thiochromenopyridinone scaffold are promising putative HSP90 inhibitors causing tumor cell growth inhibition.

  1. Molecular mechanism of bacterial Hsp90 pH-dependent ATPase activity.

    Science.gov (United States)

    Jin, Yi; Hoxie, Reyal S; Street, Timothy O

    2017-06-01

    Hsp90 is a dimeric molecular chaperone that undergoes an essential and highly regulated open-to-closed-to-open conformational cycle upon ATP binding and hydrolysis. Although it has been established that a large energy barrier to closure is responsible for Hsp90's low ATP hydrolysis rate, the specific molecular contacts that create this energy barrier are not known. Here we discover that bacterial Hsp90 (HtpG) has a pH-dependent ATPase activity that is unique among other Hsp90 homologs. The underlying mechanism is a conformation-specific electrostatic interaction between a single histidine, H255, and bound ATP. H255 stabilizes ATP only while HtpG adopts a catalytically inactive open configuration, resulting in a striking anti-correlation between nucleotide binding affinity and chaperone activity over a wide range of pH. Linkage analysis reveals that the H255-ATP salt bridge contributes 1.5 kcal/mol to the energy barrier of closure. This energetic contribution is structurally asymmetric, whereby only one H255-ATP salt-bridge per dimer of HtpG controls ATPase activation. We find that a similar electrostatic mechanism regulates the ATPase of the endoplasmic reticulum Hsp90, and that pH-dependent activity can be engineered into eukaryotic cytosolic Hsp90. These results reveal site-specific energetic information about an evolutionarily conserved conformational landscape that controls Hsp90 ATPase activity. © 2017 The Protein Society.

  2. A primate specific extra domain in the molecular chaperone Hsp90.

    Directory of Open Access Journals (Sweden)

    Vishwadeepak Tripathi

    Full Text Available Hsp90 (heat shock protein 90 is an essential molecular chaperone that mediates folding and quality control of client proteins. Many of them such as protein kinases, steroid receptors and transcription factors are involved in cellular signaling processes. Hsp90 undergoes an ATP hydrolysis dependent conformational cycle to assist folding of the client protein. The canonical Hsp90 shows a typical composition of three distinct domains and interacts with individual cochaperone partners such as Hop, Cdc37 and Aha1 (activator of Hsp90 ATPase that regulate the reaction cycle of the molecular chaperone. A bioinformatic survey identified an additional domain of 122 amino acids in front of the canonical Hsp90 sequence. This extra domain (E domain is specific to the Catarrhini or drooping nose monkeys, a subdivision of the higher primates that includes man, the great apes and the old world monkeys but is absent from all other species. Our biochemical analysis reveals that Hsp103 associates with cochaperone proteins such as Hop, Cdc37 and Aha1 similar to Hsp90. However, the extra domain reduces the ATP hydrolysis rate to about half when compared to Hsp90 thereby acting as a negative regulator of the molecular chaperonés intrinsic ATPase activity.

  3. Heat Shock Protein 90 (HSP90 and Her2 in Adenocarcinomas of the Esophagus

    Directory of Open Access Journals (Sweden)

    Julia Slotta-Huspenina

    2014-06-01

    Full Text Available Her2 overexpression and amplification can be found in a significant subset of esophageal adenocarcinomas. The activity of Her2 has been shown to be modulated by molecular chaperones such as HSP90. We analyzed expression/amplification data for HSP90 and Her2 on 127 primary resected esophageal adenocarcinomas in order to evaluate a possible relationship between these two molecules. HSP90 expression determined by immunohistochemistry was observed in various levels. Thirty nine (39 tumors (30.7% were classified as Her2-positive according to their immunoreactivity and amplification status. There was a significant correlation between HSP90 expression and Her2-status (p = 0.008. This could also be demonstrated by quantitative protein expression analysis with reverse phase protein arrays (r = 0.9; p < 0.001. Her2-status was associated withpT-category (p = 0.041, lymph node metastases (p = 0.049 and tumor differentiation (p = 0.036 with a higher percentage of cases with negative Her2 status in lower tumor stagesA negative Her2-status was also associated with better survival in univariate and multivariate analysis (p = 0.001 and p = 0.014. For HSP90, no associations between clinical and pathological parameters were found. The observed association between HSP90 expression and Her2 suggests a co-regulation of these molecules in at least a subset of esophageal adenocarcinomas. Anti-HSP90 drugs, which recently have been introduced in cancer treatment, may also be an option for these tumors by targeting HSP90 alone or in combination with Her2.

  4. Tah1 helix-swap dimerization prevents mixed Hsp90 co-chaperone complexes

    International Nuclear Information System (INIS)

    Morgan, Rhodri M. L.; Pal, Mohinder; Roe, S. Mark; Pearl, Laurence H.; Prodromou, Chrisostomos

    2015-01-01

    A helix swap involving the fifth helix between two adjacently bound Tah1 molecules restores the normal binding environment of the conserved MEEVD peptide of Hsp90. Dimerization also explains how other monomeric TPR-domain proteins are excluded from forming inappropriate mixed co-chaperone complexes with Hsp90 and Tah1. Specific co-chaperone adaptors facilitate the recruitment of client proteins to the Hsp90 system. Tah1 binds the C-terminal conserved MEEVD motif of Hsp90, thus linking an eclectic set of client proteins to the R2TP complex for their assembly and regulation by Hsp90. Rather than the normal complement of seven α-helices seen in other tetratricopeptide repeat (TPR) domains, Tah1 unusually consists of the first five only. Consequently, the methionine of the MEEVD peptide remains exposed to solvent when bound by Tah1. In solution Tah1 appears to be predominantly monomeric, and recent structures have failed to explain how Tah1 appears to prevent the formation of mixed TPR domain-containing complexes such as Cpr6–(Hsp90) 2 –Tah1. To understand this further, the crystal structure of Tah1 in complex with the MEEVD peptide of Hsp90 was determined, which shows a helix swap involving the fifth α-helix between two adjacently bound Tah1 molecules. Dimerization of Tah1 restores the normal binding environment of the bound Hsp90 methionine residue by reconstituting a TPR binding site similar to that in seven-helix-containing TPR domain proteins. Dimerization also explains how other monomeric TPR-domain proteins are excluded from forming inappropriate mixed co-chaperone complexes

  5. Tah1 helix-swap dimerization prevents mixed Hsp90 co-chaperone complexes

    Energy Technology Data Exchange (ETDEWEB)

    Morgan, Rhodri M. L.; Pal, Mohinder; Roe, S. Mark; Pearl, Laurence H., E-mail: laurence.pearl@sussex.ac.uk; Prodromou, Chrisostomos, E-mail: laurence.pearl@sussex.ac.uk [University of Sussex, Falmer, Brighton BN1 9RQ (United Kingdom)

    2015-05-01

    A helix swap involving the fifth helix between two adjacently bound Tah1 molecules restores the normal binding environment of the conserved MEEVD peptide of Hsp90. Dimerization also explains how other monomeric TPR-domain proteins are excluded from forming inappropriate mixed co-chaperone complexes with Hsp90 and Tah1. Specific co-chaperone adaptors facilitate the recruitment of client proteins to the Hsp90 system. Tah1 binds the C-terminal conserved MEEVD motif of Hsp90, thus linking an eclectic set of client proteins to the R2TP complex for their assembly and regulation by Hsp90. Rather than the normal complement of seven α-helices seen in other tetratricopeptide repeat (TPR) domains, Tah1 unusually consists of the first five only. Consequently, the methionine of the MEEVD peptide remains exposed to solvent when bound by Tah1. In solution Tah1 appears to be predominantly monomeric, and recent structures have failed to explain how Tah1 appears to prevent the formation of mixed TPR domain-containing complexes such as Cpr6–(Hsp90){sub 2}–Tah1. To understand this further, the crystal structure of Tah1 in complex with the MEEVD peptide of Hsp90 was determined, which shows a helix swap involving the fifth α-helix between two adjacently bound Tah1 molecules. Dimerization of Tah1 restores the normal binding environment of the bound Hsp90 methionine residue by reconstituting a TPR binding site similar to that in seven-helix-containing TPR domain proteins. Dimerization also explains how other monomeric TPR-domain proteins are excluded from forming inappropriate mixed co-chaperone complexes.

  6. Heat Shock Protein 90 (HSP90) and Her2 in Adenocarcinomas of the Esophagus

    Energy Technology Data Exchange (ETDEWEB)

    Slotta-Huspenina, Julia; Becker, Karl-Friedrich [Institute of Pathology, Technische Universität München, München 81765 (Germany); Feith, Marcus [Department of Surgery, Klinikum Rechts der Isar der Technischen Universität München, München 81622 (Germany); Walch, Axel [Institute of Pathology, Helmholtz Zentrum München, Deutsches Forschungszentrum für Gesundheit und Umwelt (GmbH), Neuherberg 85764 (Germany); Langer, Rupert, E-mail: rupert.langer@pathology.unibe.ch [Institute of Pathology, University of Bern, Bern 3010 (Switzerland)

    2014-06-27

    Her2 overexpression and amplification can be found in a significant subset of esophageal adenocarcinomas. The activity of Her2 has been shown to be modulated by molecular chaperones such as HSP90. We analyzed expression/amplification data for HSP90 and Her2 on 127 primary resected esophageal adenocarcinomas in order to evaluate a possible relationship between these two molecules. HSP90 expression determined by immunohistochemistry was observed in various levels. Thirty nine (39) tumors (30.7%) were classified as Her2-positive according to their immunoreactivity and amplification status. There was a significant correlation between HSP90 expression and Her2-status (p = 0.008). This could also be demonstrated by quantitative protein expression analysis with reverse phase protein arrays (r = 0.9; p < 0.001). Her2-status was associated withpT-category (p = 0.041), lymph node metastases (p = 0.049) and tumor differentiation (p = 0.036) with a higher percentage of cases with negative Her2 status in lower tumor stagesA negative Her2-status was also associated with better survival in univariate and multivariate analysis (p = 0.001 and p = 0.014). For HSP90, no associations between clinical and pathological parameters were found. The observed association between HSP90 expression and Her2 suggests a co-regulation of these molecules in at least a subset of esophageal adenocarcinomas. Anti-HSP90 drugs, which recently have been introduced in cancer treatment, may also be an option for these tumors by targeting HSP90 alone or in combination with Her2.

  7. Little effect of HSP90 inhibition on the quantitative wing traits variation in Drosophila melanogaster.

    Science.gov (United States)

    Takahashi, Kazuo H

    2017-02-01

    Drosophila wings have been a model system to study the effect of HSP90 on quantitative trait variation. The effect of HSP90 inhibition on environmental buffering of wing morphology varies among studies while the genetic buffering effect of it was examined in only one study and was not detected. Variable results so far might show that the genetic background influences the environmental and genetic buffering effect of HSP90. In the previous studies, the number of the genetic backgrounds used is limited. To examine the effect of HSP90 inhibition with a larger number of genetic backgrounds than the previous studies, 20 wild-type strains of Drosophila melanogaster were used in this study. Here I investigated the effect of HSP90 inhibition on the environmental buffering of wing shape and size by assessing within-individual and among-individual variations, and as a result, I found little or very weak effects on environmental and genetic buffering. The current results suggest that the role of HSP90 as a global regulator of environmental and genetic buffering is limited at least in quantitative traits.

  8. HSP90 is essential for Jak-STAT signaling in classical Hodgkin lymphoma cells

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    Kube Dieter

    2009-07-01

    Full Text Available Abstract In classical Hodgkin lymphoma (cHL chemotherapeutic regimens are associated with stagnant rates of secondary malignancies requiring the development of new therapeutic strategies. We and others have shown that permanently activated Signal Transducer and Activator of Transcription (STAT molecules are essential for cHL cells. Recently an overexpression of heat-shock protein 90 (HSP90 in cHL cells has been shown and inhibition of HSP90 seems to affect cHL cell survival. Here we analysed the effects of HSP90 inhibition by geldanamycin derivative 17-AAG or RNA interference (RNAi on aberrant Jak-STAT signaling in cHL cells. Treatment of cHL cell lines with 17-AAG led to reduced cell proliferation and a complete inhibition of STAT1, -3, -5 and -6 tyrosine phosphorylation probably as a result of reduced protein expression of Janus kinases (Jaks. RNAi-mediated inhibition of HSP90 showed similar effects on Jak-STAT signaling in L428 cHL cells. These results suggest a central role of HSP90 in permanently activated Jak-STAT signaling in cHL cells. Therapeutics targeting HSP90 may be a promising strategy in cHL and other cancer entities associated with deregulated Jak-STAT pathway activation.

  9. Identification of the plant compound geraniin as a novel Hsp90 inhibitor.

    Directory of Open Access Journals (Sweden)

    Antonio Vassallo

    Full Text Available Besides its function in normal cellular growth, the molecular chaperone heat shock protein 90 (Hsp90 binds to a large number of client proteins required for promoting cancer cell growth and/or survival. In an effort to discover new small molecules able to inhibit the Hsp90 ATPase and chaperoning activities, we screened, by a surface plasmon resonance assay, a small library including different plant polyphenols. The ellagitannin geraniin, was identified as the most promising molecule, showing a binding affinity to Hsp90α similar to that of 17-(allylamino-17-demethoxygeldanamycin (17AGG. Geraniin was able to inhibit in vitro the Hsp90α ATPase activity in a dose-dependent manner, with an inhibitory efficiency comparable to that measured for 17-AAG. In addition, this compound compromised the chaperone activity of Hsp90α, monitored by the citrate synthase thermal induced aggregation assay. Geraniin decreased the viability of HeLa and Jurkat cell lines and caused an arrest in G2/M phase. We also proved that following exposure to different concentrations of geraniin, the level of expression of the client proteins c-Raf, pAkt, and EGFR was strongly down-regulated in both the cell lines. These results, along with the finding that geraniin did not exert any appreciable cytotoxicity on normal cells, encourage further studies on this compound as a promising chemical scaffold for the design of new Hsp90 inhibitors.

  10. The evolutionary capacitor HSP90 buffers the regulatory effects of mammalian endogenous retroviruses.

    Science.gov (United States)

    Hummel, Barbara; Hansen, Erik C; Yoveva, Aneliya; Aprile-Garcia, Fernando; Hussong, Rebecca; Sawarkar, Ritwick

    2017-03-01

    Understanding how genotypes are linked to phenotypes is important in biomedical and evolutionary studies. The chaperone heat-shock protein 90 (HSP90) buffers genetic variation by stabilizing proteins with variant sequences, thereby uncoupling phenotypes from genotypes. Here we report an unexpected role of HSP90 in buffering cis-regulatory variation affecting gene expression. By using the tripartite-motif-containing 28 (TRIM28; also known as KAP1)-mediated epigenetic pathway, HSP90 represses the regulatory influence of endogenous retroviruses (ERVs) on neighboring genes that are critical for mouse development. Our data based on natural variations in the mouse genome show that genes respond to HSP90 inhibition in a manner dependent on their genomic location with regard to strain-specific ERV-insertion sites. The evolutionary-capacitor function of HSP90 may thus have facilitated the exaptation of ERVs as key modifiers of gene expression and morphological diversification. Our findings add a new regulatory layer through which HSP90 uncouples phenotypic outcomes from individual genotypes.

  11. Activation of Hsp90 Enzymatic Activity and Conformational Dynamics through Rationally Designed Allosteric Ligands.

    Science.gov (United States)

    Sattin, Sara; Tao, Jiahui; Vettoretti, Gerolamo; Moroni, Elisabetta; Pennati, Marzia; Lopergolo, Alessia; Morelli, Laura; Bugatti, Antonella; Zuehlke, Abbey; Moses, Mike; Prince, Thomas; Kijima, Toshiki; Beebe, Kristin; Rusnati, Marco; Neckers, Len; Zaffaroni, Nadia; Agard, David A; Bernardi, Anna; Colombo, Giorgio

    2015-09-21

    Hsp90 is a molecular chaperone of pivotal importance for multiple cell pathways. ATP-regulated internal dynamics are critical for its function and current pharmacological approaches block the chaperone with ATP-competitive inhibitors. Herein, a general approach to perturb Hsp90 through design of new allosteric ligands aimed at modulating its functional dynamics is proposed. Based on the characterization of a first set of 2-phenylbenzofurans showing stimulatory effects on Hsp90 ATPase and conformational dynamics, new ligands were developed that activate Hsp90 by targeting an allosteric site, located 65 Å from the active site. Specifically, analysis of protein responses to first-generation activators was exploited to guide the design of novel derivatives with improved ability to stimulate ATP hydrolysis. The molecules' effects on Hsp90 enzymatic, conformational, co-chaperone and client-binding properties were characterized through biochemical, biophysical and cellular approaches. These designed probes act as allosteric activators of the chaperone and affect the viability of cancer cell lines for which proper functioning of Hsp90 is necessary. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  12. Essential oil from Cymbopogon flexuosus as the potential inhibitor for HSP90.

    Science.gov (United States)

    Gaonkar, Roopa; Shiralgi, Yallappa; Lakkappa, Dhananjaya B; Hegde, Gurumurthy

    2018-01-01

    The essential oil of Cymbopogon flexuosus or lemongrass oil (LO) is reported to have antibacterial, antifungal and anticancerous effects. HSP90 is one of the major chaperones responsible for the proper folding of cancer proteins. In this paper we show that the essential oil of C. flexuosus significantly suppresses the HSP90 gene expression. The cytotoxicity of the compounds was tested by MTT assay and the gene expression studies were carried out using HEK-293 and MCF-7 cells. Also we tested the efficacy of the major component of this essential oil viz. citral and geraniol in inhibiting the HSP90 expression. The oil was found to be more cytotoxic to MCF-7 cells with different IC 50 values for the oil (69.33 μg/mL), citral (140.7 μg/mL) and geraniol (117 μg/mL). The fold change of expression was calculated by RT-qPCR using ΔΔCt (2 ^-ΔΔCt ) method and it was 0.1 and 0.03 in MCF-7 cells at 80 μg/mL and 160 μg/mL of LO. Western blot results showed suppression of HSP90 protein expression and HSP90 - ATPase activity inhibition was also observed using LO. This study shows the anticancer mechanism exhibited by the essential oil of C. flexuosus is by the inhibition of the important chaperone protein HSP90.

  13. Combined x-ray crystallography and computational modeling approach to investigate the Hsp90 C-terminal peptide binding to FKBP51.

    Science.gov (United States)

    Kumar, Rajnish; Moche, Martin; Winblad, Bengt; Pavlov, Pavel F

    2017-10-27

    FK506 binding protein of 51 kDa (FKBP51) is a heat shock protein 90 (Hsp90) co-chaperone involved in the regulation of steroid hormone receptors activity. It is known for its role in various regulatory pathways implicated in mood and stress-related disorders, cancer, obesity, Alzheimer's disease and corticosteroid resistant asthma. It consists of two FKBP12 like active peptidyl prolyl isomerase (PPIase) domains (an active FK1 and inactive FK2 domain) and one tetratricopeptide repeat (TPR) domain that mediates interaction with Hsp90 via its C-terminal MEEVD peptide. Here, we report a combined x-ray crystallography and molecular dynamics study to reveal the binding mechanism of Hsp90 MEEVD peptide to the TPR domain of FKBP51. The results demonstrated that the Hsp90 C-terminal peptide binds to the TPR domain of FKBP51 with the help of di-carboxylate clamp involving Lys272, Glu273, Lys352, Asn322, and Lys329 which are conserved throughout several di-carboxylate clamp TPR proteins. Interestingly, the results from molecular dynamics study are also in agreement to the complex structure where all the contacts between these two partners were consistent throughout the simulation period. In a nutshell, our findings provide new opportunity to engage this important protein-protein interaction target by small molecules designed by structure based drug design strategy.

  14. The regulatory mechanism of Hsp90α secretion from endothelial cells and its role in angiogenesis during wound healing

    International Nuclear Information System (INIS)

    Song, Xiaomin; Luo, Yongzhang

    2010-01-01

    Research highlights: → Growth factors such as bFGF, VEGF, PDGF and SDF-1 stimulate Hsp90α secretion from endothelial cells. → Secreted Hsp90α localizes on the leading edge of activated endothelial cells. → Secreted Hsp90α promotes angiogenesis in wound healing. -- Abstract: Heat shock protein 90α (Hsp90α) is a ubiquitously expressed molecular chaperone, which is essential for the maintenance of eukaryote homeostasis. Hsp90α can also be secreted extracellularly and is associated with several physiological and pathological processes including wound healing, cancer, infectious diseases and diabetes. Angiogenesis, defined as the sprouting of new blood vessels from pre-existing capillaries via endothelial cell proliferation and migration, commonly occurs in and contributes to the above mentioned processes. However, the secretion of Hsp90α from endothelial cells and also its function in angiogenesis are still unclear. Here we investigated the role of extracellular Hsp90α in angiogenesis using dermal endothelial cells in vitro and a wound healing model in vivo. We find that the secretion of Hsp90α but not Hsp90β is increased in activated endothelial cells with the induction of angiogenic factors and matrix proteins. Secreted Hsp90α localizes on the leading edge of endothelial cells and promotes their angiogenic activities, whereas Hsp90α neutralizing antibodies reverse the effect. Furthermore, using a mouse skin wound healing model in vivo, we demonstrate that extracellular Hsp90α localizes on blood vessels in granulation tissues of wounded skin and promotes angiogenesis during wound healing. Taken together, our study reveals that Hsp90α can be secreted by activated endothelial cells and is a positive regulator of angiogenesis, suggesting the potential application of Hsp90α as a stimulator for wound repair.

  15. The regulatory mechanism of Hsp90{alpha} secretion from endothelial cells and its role in angiogenesis during wound healing

    Energy Technology Data Exchange (ETDEWEB)

    Song, Xiaomin [National Engineering Laboratory for Anti-tumor Protein Therapeutics, Tsinghua University, Beijing 100084 (China); Beijing Key Laboratory for Protein Therapeutics, Tsinghua University, Beijing 100084 (China); Cancer Biology Laboratory, School of Life Sciences, Tsinghua University, Beijing 100084 (China); Luo, Yongzhang, E-mail: yluo@tsinghua.edu.cn [National Engineering Laboratory for Anti-tumor Protein Therapeutics, Tsinghua University, Beijing 100084 (China); Beijing Key Laboratory for Protein Therapeutics, Tsinghua University, Beijing 100084 (China); Cancer Biology Laboratory, School of Life Sciences, Tsinghua University, Beijing 100084 (China)

    2010-07-16

    Research highlights: {yields} Growth factors such as bFGF, VEGF, PDGF and SDF-1 stimulate Hsp90{alpha} secretion from endothelial cells. {yields} Secreted Hsp90{alpha} localizes on the leading edge of activated endothelial cells. {yields} Secreted Hsp90{alpha} promotes angiogenesis in wound healing. -- Abstract: Heat shock protein 90{alpha} (Hsp90{alpha}) is a ubiquitously expressed molecular chaperone, which is essential for the maintenance of eukaryote homeostasis. Hsp90{alpha} can also be secreted extracellularly and is associated with several physiological and pathological processes including wound healing, cancer, infectious diseases and diabetes. Angiogenesis, defined as the sprouting of new blood vessels from pre-existing capillaries via endothelial cell proliferation and migration, commonly occurs in and contributes to the above mentioned processes. However, the secretion of Hsp90{alpha} from endothelial cells and also its function in angiogenesis are still unclear. Here we investigated the role of extracellular Hsp90{alpha} in angiogenesis using dermal endothelial cells in vitro and a wound healing model in vivo. We find that the secretion of Hsp90{alpha} but not Hsp90{beta} is increased in activated endothelial cells with the induction of angiogenic factors and matrix proteins. Secreted Hsp90{alpha} localizes on the leading edge of endothelial cells and promotes their angiogenic activities, whereas Hsp90{alpha} neutralizing antibodies reverse the effect. Furthermore, using a mouse skin wound healing model in vivo, we demonstrate that extracellular Hsp90{alpha} localizes on blood vessels in granulation tissues of wounded skin and promotes angiogenesis during wound healing. Taken together, our study reveals that Hsp90{alpha} can be secreted by activated endothelial cells and is a positive regulator of angiogenesis, suggesting the potential application of Hsp90{alpha} as a stimulator for wound repair.

  16. In Vivo Detection of HSP90 Identifies Breast Cancers with Aggressive Behavior.

    Science.gov (United States)

    Osada, Takuya; Kaneko, Kensuke; Gwin, William R; Morse, Michael A; Hobeika, Amy; Pogue, Brian W; Hartman, Zachary C; Hughes, Philip F; Haystead, Timothy; Lyerly, H Kim

    2017-12-15

    Purpose: Hsp90, a chaperone to numerous molecular pathways in malignant cells, is elevated in aggressive breast cancers. We hypothesized that identifying breast cells with elevated Hsp90 activity in situ could result in early detection of aggressive breast cancers. Experimental Design: We exploited the uptake of an Hsp90 inhibitor by malignant cells to create an imaging probe (HS131) of Hsp90 activity by linking it to a near-infrared (nIR) dye. HS131 uptake into cells correlated with cell membrane expression of Hsp90 and was used to image molecular subtypes of murine and human breast cancers in vitro and in murine models. Results: HS131 imaging was both sensitive and specific in detecting the murine 4T1 breast cancer cell line, as well as subclones with differing metastatic potential. Highly metastatic subclones (4T07) had high HS131 uptake, but subclones with lower metastatic potential (67NR, 168FARN) had low HS131 uptake. We generated isogenic cell lines to demonstrate that overexpression of a variety of specific oncogenes resulted in high HS131 uptake and retention. Finally, we demonstrated that HS131 could be used to detect spontaneous tumors in MMTV-neu mice, as well as primary and metastatic human breast cancer xenografts. HS131 could image invasive lobular breast cancer, a histologic subtype of breast cancer which is often undetectable by mammography. Conclusions: An HSP90-targeting nIR probe is sensitive and specific in imaging all molecular subtypes of murine and human breast cancer, with higher uptake in aggressive and highly metastatic clones. Clinical studies with Hsp90-targeting nIR probes will be initiated shortly. Clin Cancer Res; 23(24); 7531-42. ©2017 AACR . ©2017 American Association for Cancer Research.

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

    International Nuclear Information System (INIS)

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

    2010-01-01

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

  18. Gene expression of Hsp70, Hsp90 and Hsp110 families in normal palate and cleft palate during mouse embryogenesis.

    Science.gov (United States)

    Zhu, Yongfei; Ren, Chuanlu; Wan, Xuying; Zhu, Yuping; Zhu, Jiangbo; Zhou, Hongyuan; Zhang, Tianbao

    2013-11-01

    Most previous studies focused on a small number of heat shock proteins (Hsps) and their relationships with embryogenesis, and the actual roles of these Hsps in normal and abnormal embryonic development remain unclear. It was found in the present systemic study that except for Grp170, whose expression was not detectable at GD18, all 19 Hsps of Hsp70, Hsp90 and Hsp110 families were expressed in the normal development of embryonic palate tissue in mice, but their expression patterns varied with different Hsps, presenting as a correlation with the developmental phases. In the treatment group by all-trans retinoic acid (atRA), the messenger RNA (mRNA) abundance of HspA1A, HspA1L, HspA8, HspA9, HspA12A, HspA12B, HspA13, HspA14, Hsp90AA1, Hsp90AB1, Grp94, Trap1, Hsp105, Hsp110 and Grp170 was higher in the palates at GD11 (the beginning of palate development), the mRNA abundance of HspA1A, HspA12A and HspA12B was higher at GD18 (before birth) and an mRNA expression peak of HspA1L, HspA8, HspA9, Hsp90AA1, Grp94, Hsp110 and Grp170 was observed at GD17. The mRNA abundance of most genes in atRA-induced cleft palates of the treatment group was different from that of the control group. Grp78, HspA14 and Hsp105 were closely associated with the normal palate development and cleft palate in mouse embryo, possibly as palate development-related genes. Except Grp170, the other genes may be closely associated with the development of mouse palates through participating in the stress response process and/or the antiapoptosis process.

  19. Hsp90-downregulation influences the heat-shock response, innate immune response and onset of oocyte development in nematodes.

    Directory of Open Access Journals (Sweden)

    Julia Eckl

    Full Text Available Hsp90 is a molecular chaperone involved in the regulation and maturation of kinases and transcription factors. In Caenorhabditis elegans, it contributes to the development of fertility, maintenance of muscle structure, the regulation of heat-shock response and dauer state. To understand the consequences of Hsp90-depletion, we studied Hsp90 RNAi-treated nematodes by DNA microarrays and mass spectrometry. We find that upon development of phenotypes the levels of chaperones and Hsp90 cofactors are increased, while specific proteins related to the innate immune response are depleted. In microarrays, we further find many differentially expressed genes related to gonad and larval development. These genes form an expression cluster that is regulated independently from the immune response implying separate pathways of Hsp90-involvement. Using fluorescent reporter strains for the differentially expressed immune response genes skr-5, dod-24 and clec-60 we observe that their activity in intestinal tissues is influenced by Hsp90-depletion. Instead, effects on the development are evident in both gonad arms. After Hsp90-depletion, changes can be observed in early embryos and adults containing fluorescence-tagged versions of SEPA-1, CAV-1 or PUD-1, all of which are downregulated after Hsp90-depletion. Our observations identify molecular events for Hsp90-RNAi induced phenotypes during development and immune responses, which may help to separately investigate independent Hsp90-influenced processes that are relevant during the nematode's life and development.

  20. A rat retinal damage model predicts for potential clinical visual disturbances induced by Hsp90 inhibitors

    International Nuclear Information System (INIS)

    Zhou, Dan; Liu, Yuan; Ye, Josephine; Ying, Weiwen; Ogawa, Luisa Shin; Inoue, Takayo; Tatsuta, Noriaki; Wada, Yumiko; Koya, Keizo; Huang, Qin; Bates, Richard C.; Sonderfan, Andrew J.

    2013-01-01

    In human trials certain heat shock protein 90 (Hsp90) inhibitors, including 17-DMAG and NVP-AUY922, have caused visual disorders indicative of retinal dysfunction; others such as 17-AAG and ganetespib have not. To understand these safety profile differences we evaluated histopathological changes and exposure profiles of four Hsp90 inhibitors, with or without clinical reports of adverse ocular effects, using a rat retinal model. Retinal morphology, Hsp70 expression (a surrogate marker of Hsp90 inhibition), apoptotic induction and pharmacokinetic drug exposure analysis were examined in rats treated with the ansamycins 17-DMAG and 17-AAG, or with the second-generation compounds NVP-AUY922 and ganetespib. Both 17-DMAG and NVP-AUY922 induced strong yet restricted retinal Hsp70 up-regulation and promoted marked photoreceptor cell death 24 h after the final dose. In contrast, neither 17-AAG nor ganetespib elicited photoreceptor injury. When the relationship between drug distribution and photoreceptor degeneration was examined, 17-DMAG and NVP-AUY922 showed substantial retinal accumulation, with high retina/plasma (R/P) ratios and slow elimination rates, such that 51% of 17-DMAG and 65% of NVP-AUY922 present at 30 min post-injection were retained in the retina 6 h post-dose. For 17-AAG and ganetespib, retinal elimination was rapid (90% and 70% of drugs eliminated from the retina at 6 h, respectively) which correlated with lower R/P ratios. These findings indicate that prolonged inhibition of Hsp90 activity in the eye results in photoreceptor cell death. Moreover, the results suggest that the retina/plasma exposure ratio and retinal elimination rate profiles of Hsp90 inhibitors, irrespective of their chemical class, may predict for ocular toxicity potential. - Highlights: • In human trials some Hsp90 inhibitors cause visual disorders, others do not. • Prolonged inhibition of Hsp90 in the rat eye results in photoreceptor cell death. • Retina/plasma ratio and retinal

  1. An Impermeant Ganetespib Analog Inhibits Extracellular Hsp90-Mediated Cancer Cell Migration that Involves Lysyl Oxidase 2-like Protein

    Energy Technology Data Exchange (ETDEWEB)

    McCready, Jessica [Department of Natural Sciences, Assumption College, Worcester, MA 01609 (United States); Wong, Daniel S. [Department of Developmental Molecular and Chemical Biology, Tufts University School of Medicine, Boston, MA 02111 (United States); Cell and Molecular Physiology Program, Sackler School of Graduate Biomedical Sciences, Tufts University, Boston, MA 02111 (United States); Burlison, Joseph A.; Ying, Weiwen [Synta Pharmaceuticals, Lexington, MA 02421 (United States); Jay, Daniel G., E-mail: daniel.jay@tufts.edu [Department of Developmental Molecular and Chemical Biology, Tufts University School of Medicine, Boston, MA 02111 (United States); Cell and Molecular Physiology Program, Sackler School of Graduate Biomedical Sciences, Tufts University, Boston, MA 02111 (United States)

    2014-04-30

    Extracellular Hsp90 (eHsp90) activates a number of client proteins outside of cancer cells required for migration and invasion. Therefore, eHsp90 may serve as a novel target for anti-metastatic drugs as its inhibition using impermeant Hsp90 inhibitors would not affect the numerous vital intracellular Hsp90 functions in normal cells. While some eHsp90 clients are known, it is important to establish other proteins that act outside the cell to validate eHsp90 as a drug target to limit cancer spread. Using mass spectrometry we identified two precursor proteins Galectin 3 binding protein (G3BP) and Lysyl oxidase 2-like protein (LOXL2) that associate with eHsp90 in MDA-MB231 breast cancer cell conditioned media and confirmed that LOXL2 binds to eHsp90 in immunoprecipitates. We introduce a novel impermeant Hsp90 inhibitor STA-12-7191 derived from ganetespib and show that it is markedly less toxic to cells and can inhibit cancer cell migration in a dose dependent manner. We used STA-12-7191 to test if LOXL2 and G3BP are potential eHsp90 clients. We showed that while LOXL2 can increase wound healing and compensate for STA-12-7191-mediated inhibition of wound closure, addition of G3BP had no affect on this assay. These findings support of role for LOXL2 in eHsp90 stimulated cancer cell migration and provide preliminary evidence for the use of STA-12-7191 to inhibit eHsp90 to limit cancer invasion.

  2. An Impermeant Ganetespib Analog Inhibits Extracellular Hsp90-Mediated Cancer Cell Migration that Involves Lysyl Oxidase 2-like Protein

    International Nuclear Information System (INIS)

    McCready, Jessica; Wong, Daniel S.; Burlison, Joseph A.; Ying, Weiwen; Jay, Daniel G.

    2014-01-01

    Extracellular Hsp90 (eHsp90) activates a number of client proteins outside of cancer cells required for migration and invasion. Therefore, eHsp90 may serve as a novel target for anti-metastatic drugs as its inhibition using impermeant Hsp90 inhibitors would not affect the numerous vital intracellular Hsp90 functions in normal cells. While some eHsp90 clients are known, it is important to establish other proteins that act outside the cell to validate eHsp90 as a drug target to limit cancer spread. Using mass spectrometry we identified two precursor proteins Galectin 3 binding protein (G3BP) and Lysyl oxidase 2-like protein (LOXL2) that associate with eHsp90 in MDA-MB231 breast cancer cell conditioned media and confirmed that LOXL2 binds to eHsp90 in immunoprecipitates. We introduce a novel impermeant Hsp90 inhibitor STA-12-7191 derived from ganetespib and show that it is markedly less toxic to cells and can inhibit cancer cell migration in a dose dependent manner. We used STA-12-7191 to test if LOXL2 and G3BP are potential eHsp90 clients. We showed that while LOXL2 can increase wound healing and compensate for STA-12-7191-mediated inhibition of wound closure, addition of G3BP had no affect on this assay. These findings support of role for LOXL2 in eHsp90 stimulated cancer cell migration and provide preliminary evidence for the use of STA-12-7191 to inhibit eHsp90 to limit cancer invasion

  3. Localization of MRP-1 to the outer mitochondrial membrane by the chaperone protein HSP90β.

    Science.gov (United States)

    Roundhill, Elizabeth; Turnbull, Doug; Burchill, Susan

    2016-05-01

    Overexpression of plasma membrane multidrug resistance-associated protein 1 (MRP-1) in Ewing's sarcoma (ES) predicts poor outcome. MRP-1 is also expressed in mitochondria, and we have examined the submitochondrial localization of MRP-1 and investigated the mechanism of MRP-1 transport and role of this organelle in the response to doxorubicin. The mitochondrial localization of MRP-1 was examined in ES cell lines by differential centrifugation and membrane solubilization by digitonin. Whether MRP-1 is chaperoned by heat shock proteins (HSPs) was investigated by immunoprecipitation, immunofluorescence microscopy, and HSP knockout using small hairpin RNA and inhibitors (apoptozole, 17-AAG, and NVPAUY). The effect of disrupting mitochondrial MRP-1-dependent efflux activity on the cytotoxic effect of doxorubicin was investigated by counting viable cell number. Mitochondrial MRP-1 is glycosylated and localized to the outer mitochondrial membrane, where it is coexpressed with HSP90. MRP-1 binds to both HSP90 and HSP70, although only inhibition of HSP90β decreases expression of MRP-1 in the mitochondria. Disruption of mitochondrial MRP-1-dependent efflux significantly increases the cytotoxic effect of doxorubicin (combination index, MRP-1 is expressed in the outer mitochondrial membrane and is a client protein of HSP90β, where it may play a role in the doxorubicin-induced resistance of ES.-Roundhill, E., Turnbull, D., Burchill, S. Localization of MRP-1 to the outer mitochondrial membrane by the chaperone protein HSP90β. © FASEB.

  4. SGT, a Hsp90β binding partner, is accumulated in the nucleus during cell apoptosis

    International Nuclear Information System (INIS)

    Yin Hongyan; Wang Hanzhou; Zong Hongliang; Chen Xiaoning; Wang Yanlin; Yun Xiaojing; Wu Yihong; Wang Jiadong; Gu Jianxin

    2006-01-01

    In this study, we reported that small glutamine-rich TPR-containing protein (SGT) interacted with not only Hsp90α but also Hsp90β. Confocal analysis showed that treatment of cells with Hsp90-specific inhibitor geldanamycin (GA) disrupted the interaction of SGT with Hsp90β and this contributed to the increase of nuclear localization of SGT in HeLa cells. The increased nuclear localization of SGT was further confirmed by the Western blotting in GA-treated HeLa cells and H1299 cells. In our previous study, SGT was found to be a new pro-apoptotic factor, so we wondered whether the sub-cellular localization of SGT was related with cell apoptosis. By confocal analysis we found that the nuclear import of SGT was significantly increased in STS-induced apoptotic HeLa cells, which implied that the sub-cellular localization of SGT was closely associated with Hsp90β and apoptosis

  5. HECTD3 Mediates an HSP90-Dependent Degradation Pathway for Protein Kinase Clients

    Directory of Open Access Journals (Sweden)

    Zhaobo Li

    2017-06-01

    Full Text Available Inhibition of the ATPase cycle of the HSP90 chaperone promotes ubiquitylation and proteasomal degradation of its client proteins, which include many oncogenic protein kinases. This provides the rationale for HSP90 inhibitors as cancer therapeutics. However, the mechanism by which HSP90 ATPase inhibition triggers ubiquitylation is not understood, and the E3 ubiquitin ligases involved are largely unknown. Using a siRNA screen, we have identified components of two independent degradation pathways for the HSP90 client kinase CRAF. The first requires CUL5, Elongin B, and Elongin C, while the second requires the E3 ligase HECTD3, which is also involved in the degradation of MASTL and LKB1. HECTD3 associates with HSP90 and CRAF in cells via its N-terminal DOC domain, which is mutationally disrupted in tumor cells with activated MAP kinase signaling. Our data implicate HECTD3 as a tumor suppressor modulating the activity of this important oncogenic signaling pathway.

  6. Hsp90 as a Gatekeeper of Tumor Angiogenesis: Clinical Promise and Potential Pitfalls

    Directory of Open Access Journals (Sweden)

    J. E. Bohonowych

    2010-01-01

    Full Text Available Tumor vascularization is an essential modulator of early tumor growth, progression, and therapeutic outcome. Although antiangiogenic treatments appear promising, intrinsic and acquired tumor resistance contributes to treatment failure. Clinical inhibition of the molecular chaperone heat shock protein 90 (Hsp90 provides an opportunity to target multiple aspects of this signaling resiliency, which may elicit more robust and enduring tumor repression relative to effects elicited by specifically targeted agents. This review highlights several primary effectors of angiogenesis modulated by Hsp90 and describes the clinical challenges posed by the redundant circuitry of these pathways. The four main topics addressed include (1 Hsp90-mediated regulation of HIF/VEGF signaling, (2 chaperone-dependent regulation of HIF-independent VEGF-mediated angiogenesis, (3 Hsp90-dependent targeting of key proangiogenic receptor tyrosine kinases and modulation of drug resistance, and (4 consideration of factors such as tumor microenvironment that pose several challenges for the clinical efficacy of anti-angiogenic therapy and Hsp90-targeted strategies.

  7. Spontaneous assembly of HSP90 inhibitors at water/octanol interface: A molecular dynamics simulation study

    Science.gov (United States)

    Zolghadr, Amin Reza; Boroomand, Samaneh

    2017-02-01

    Drug absorption at an acceptable dose depends on the pair of solubility and permeability. There are many potent therapeutics that are not active in vivo, presumably due to the lack of capability to cross the cell membrane. Molecular dynamics simulation of radicicol, diol-radicicol, cyclopropane-radicicol and 17-DMAG were performed at water/octanol interface to suggest interfacial activity as a physico-chemical characteristic of these heat shock protein 90 (HSP90) inhibitors. We have observed that orally active HSP90 inhibitors form aggregates at the water/octanol and DPPC-lipid/water interfaces by starting from an initial configuration with HSP90 inhibitors embedded in the water matrix.

  8. Hsp90 molecular chaperone: structure, functions and participation in cardio-vascular pathologies

    Directory of Open Access Journals (Sweden)

    Kroupskaya I. V.

    2009-10-01

    Full Text Available The review is devoted to the analysis of structural and functional properties of molecular chaperon Hsp90. Hsp90 is a representative of highly widespread family of heat shock proteins. The protein is found in eubacteria and all branches of eukarya, but it is apparently absent in archaea. It is one of key regulators of numerous signalling pathways, cell growth and development, apoptosis, induction of autoimmunity, and progression of heart failure. The full functional activity of Hsp90 shows up in a complex with other molecular chaperones and co-chaperones. Molecular interactions between chaperones, different signalling proteins and protein-partners are highly crucial for the normal functioning of signalling pathways and their destruction causes an alteration in the cell physiology up to its death.

  9. A Hyperactive Signalosome in Acute Myeloid Leukemia Drives Addiction to a Tumor-Specific Hsp90 Species

    Directory of Open Access Journals (Sweden)

    Hongliang Zong

    2015-12-01

    Full Text Available Acute myeloid leukemia (AML is a heterogeneous and fatal disease with an urgent need for improved therapeutic regimens given that most patients die from relapsed disease. Irrespective of mutation status, the development of aggressive leukemias is enabled by increasing dependence on signaling networks. We demonstrate that a hyperactive signalosome drives addiction of AML cells to a tumor-specific Hsp90 species (teHsp90. Through genetic, environmental, and pharmacologic perturbations, we demonstrate a direct and quantitative link between hyperactivated signaling pathways and apoptotic sensitivity of AML to teHsp90 inhibition. Specifically, we find that hyperactive JAK-STAT and PI3K-AKT signaling networks are maintained by teHsp90 and, in fact, gradual activation of these networks drives tumors increasingly dependent on teHsp90. Thus, although clinically aggressive AML survives via signalosome activation, this addiction creates a vulnerability that can be exploited with Hsp90-directed therapy.

  10. PKC signaling regulates drug resistance of the fungal pathogen Candida albicans via circuitry comprised of Mkc1, calcineurin, and Hsp90.

    Directory of Open Access Journals (Sweden)

    Shantelle L LaFayette

    2010-08-01

    Full Text Available Fungal pathogens exploit diverse mechanisms to survive exposure to antifungal drugs. This poses concern given the limited number of clinically useful antifungals and the growing population of immunocompromised individuals vulnerable to life-threatening fungal infection. To identify molecules that abrogate resistance to the most widely deployed class of antifungals, the azoles, we conducted a screen of 1,280 pharmacologically active compounds. Three out of seven hits that abolished azole resistance of a resistant mutant of the model yeast Saccharomyces cerevisiae and a clinical isolate of the leading human fungal pathogen Candida albicans were inhibitors of protein kinase C (PKC, which regulates cell wall integrity during growth, morphogenesis, and response to cell wall stress. Pharmacological or genetic impairment of Pkc1 conferred hypersensitivity to multiple drugs that target synthesis of the key cell membrane sterol ergosterol, including azoles, allylamines, and morpholines. Pkc1 enabled survival of cell membrane stress at least in part via the mitogen activated protein kinase (MAPK cascade in both species, though through distinct downstream effectors. Strikingly, inhibition of Pkc1 phenocopied inhibition of the molecular chaperone Hsp90 or its client protein calcineurin. PKC signaling was required for calcineurin activation in response to drug exposure in S. cerevisiae. In contrast, Pkc1 and calcineurin independently regulate drug resistance via a common target in C. albicans. We identified an additional level of regulatory control in the C. albicans circuitry linking PKC signaling, Hsp90, and calcineurin as genetic reduction of Hsp90 led to depletion of the terminal MAPK, Mkc1. Deletion of C. albicans PKC1 rendered fungistatic ergosterol biosynthesis inhibitors fungicidal and attenuated virulence in a murine model of systemic candidiasis. This work establishes a new role for PKC signaling in drug resistance, novel circuitry through which

  11. Hsp90 chaperone inhibitor 17-AAG attenuates Aβ-induced synaptic toxicity and memory impairment.

    Science.gov (United States)

    Chen, Yaomin; Wang, Bin; Liu, Dan; Li, Jing Jing; Xue, Yueqiang; Sakata, Kazuko; Zhu, Ling-qiang; Heldt, Scott A; Xu, Huaxi; Liao, Francesca-Fang

    2014-02-12

    The excessive accumulation of soluble amyloid peptides (Aβ) plays a crucial role in the pathogenesis of Alzheimer's disease (AD), particularly in synaptic dysfunction. The role of the two major chaperone proteins, Hsp70 and Hsp90, in clearing misfolded protein aggregates has been established. Despite their abundant presence in synapses, the role of these chaperones in synapses remains elusive. Here, we report that Hsp90 inhibition by 17-AAG elicited not only a heat shock-like response but also upregulated presynaptic and postsynaptic proteins, such as synapsin I, synaptophysin, and PSD95 in neurons. 17-AAG treatment enhanced high-frequency stimulation-evoked LTP and protected neurons from synaptic damage induced by soluble Aβ. In AD transgenic mice, the daily administration of 17-AAG over 7 d resulted in a marked increase in PSD95 expression in hippocampi. 17-AAG treatments in wild-type C57BL/6 mice challenged by soluble Aβ significantly improved contextual fear memory. Further, we demonstrate that 17-AAG activated synaptic protein expression via transcriptional mechanisms through the heat shock transcription factor HSF1. Together, our findings identify a novel function of Hsp90 inhibition in regulating synaptic plasticity, in addition to the known neuroprotective effects of the chaperones against Aβ and tau toxicity, thus further supporting the potential of Hsp90 inhibitors in treating neurodegenerative diseases.

  12. Extracellular Hsp90 as a Novel Epigenetic of EMT and Metastatic Risk in Prostate Cancer

    Science.gov (United States)

    2015-12-01

    effects of eHsp90. As expected, treatment with NPGA or UO126 restored E-cadherin expression in tan - dem with diminished P-ERK and EZH2 (Fig. 1D...co-occupy a cohort of EZH2 target sites and function as a repressive cofac- tor (43, 44). Therefore, additional studies are warranted to fur - ther

  13. Effect of Resuscitation Fluids on the Expression of hsp90α in ...

    African Journals Online (AJOL)

    muscles in the early phase of shock. In addition, the protective effect of hsp90α protein on cardiac muscles is mainly based on its effect as a molecular chaperone by combining with the. bHLH-PΑS domain of hypoxia inducible factor 1α. (HIF-1α) [11]. Table 1: Survival rate of the animals. Significant differences were observed ...

  14. Optimizing fluorescently tethered Hsp90 inhibitor dose for maximal specific uptake by breast tumors

    Science.gov (United States)

    Crouch, Brian T.; Duer, Joy; Wang, Roujia; Gallagher, Jennifer; Hall, Allison; Soo, Mary Scott; Hughes, Philip; Haystead, Timothy A. J.; Ramanujam, Nirmala

    2018-03-01

    Despite improvements in surgical resection, 20-40% of patients undergoing breast conserving surgery require at least one additional re-excision. Leveraging the unique surface expression of heat shock protein 90 (Hsp90), a chaperone protein involved in several key hallmarks of cancer, in breast cancer provides an exciting opportunity to identify residual disease during surgery. We developed a completely non-destructive strategy using HS-27, a fluorescently-tethered Hsp90 inhibitor, to assay surface Hsp90 expression on intact tissue specimens using a fluorescence microendoscope with a field of view of 750 μm and subcellular resolution of 4 μm. HS-27 consists of an FDA approved Hsp90 inhibitor tethered to fluorescein isothiocyanate (EX 488nm, EM 525nm). Here, we optimized ex vivo HS-27 administration in pre-clinical breast cancer models and validated our approach on 21 patients undergoing standard of care ultrasound guided core needle biopsy. HS-27 administration time was fixed at 1- minute to minimize imaging impact on clinical workflow. HS-27 and HS-217 (non-specific control) doses were modulated from 1 μM up to 100 μM to identify the dose maximizing the ratio of specific uptake (HS-27 fluorescence) to non-specific uptake (HS-217 fluorescence). The specificity ratio was maximized at 100 μM and was significantly greater than all other doses (pcancer makes this technology attractive for assessing tumor margins, as one agent can be used for all subtypes.

  15. Phosphorylation and oligomerization states of native pig brain HSP90 studied by mass spectrometry

    DEFF Research Database (Denmark)

    Garnier, C.; Lafitte, D.; Jorgensen, T.J.

    2001-01-01

    HSP90 purified from pig brain. The two protein isoforms were clearly distinguished by ESI-MS, the alpha isoform being approximately six times more abundant than the beta isoform. ESI-MS in combination with lambda phosphatase treatment provided direct evidence of the existence of four phosphorylated...

  16. Cyclophilin40 isomerase activity is regulated by a temperature-dependent allosteric interaction with Hsp90.

    Science.gov (United States)

    Blackburn, Elizabeth A; Wear, Martin A; Landré, Vivian; Narayan, Vikram; Ning, Jia; Erman, Burak; Ball, Kathryn L; Walkinshaw, Malcolm D

    2015-09-01

    Cyclophilin 40 (Cyp40) comprises an N-terminal cyclophilin domain with peptidyl-prolyl isomerase (PPIase) activity and a C-terminal tetratricopeptide repeat (TPR) domain that binds to the C-terminal-EEVD sequence common to both heat shock protein 70 (Hsp70) and Hsp90. We show in the present study that binding of peptides containing the MEEVD motif reduces the PPIase activity by ∼30%. CD and fluorescence assays show that the TPR domain is less stable than the cyclophilin domain and is stabilized by peptide binding. Isothermal titration calorimetry (ITC) shows that the affinity for the-MEEVD peptide is temperature sensitive in the physiological temperature range. Results from these biophysical studies fit with the MD simulations of the apo and holo (peptide-bound) structures which show a significant reduction in root mean square (RMS) fluctuation in both TPR and cyclophilin domains when-MEEVD is bound. The MD simulations of the apo-protein also highlight strong anti-correlated motions between residues around the PPIase-active site and a band of residues running across four of the seven helices in the TPR domain. Peptide binding leads to a distortion in the shape of the active site and a significant reduction in these strongly anti-correlated motions, providing an explanation for the allosteric effect of ligand binding and loss of PPIase activity. Together the experimental and MD results suggest that on heat shock, dissociation of Cyp40 from complexes mediated by the TPR domain leads to an increased pool of free Cyp40 capable of acting as an isomerase/chaperone in conditions of cellular stress. © 2015 Authors.

  17. Heat shock protein 90 (Hsp90) chaperone complex. A molecular target for enhancement of thermosensitivity and radiosensitivity

    International Nuclear Information System (INIS)

    Akimoto, Tetsuo; Nonaka, Tetsuo; Kitamoto, Yoshizumi; Sakurai, Hideyuki

    2002-01-01

    Heat shock protein 90 (Hsp90) is a highly conserved heat shock protein in animal and plants, and exists abundantly in the cytoplasm in unstressed condition, accounting for 1-2% in cytoplasmic proteins. Main difference of Hsp90 from other Hsps are its substrate that Hsp90 binds to. These substrates include various signal transduction proteins, kinase, steroid receptors and transcription factors, therefore, Hsp90 plays a key role in maintaining cellular signal transduction networks. Many chaperoned proteins (client proteins) of Hsp90 are associated with cellular proliferation or malignant transformation, thus Hsp90 chaperone complex has been focused as targets for cancer therapy. Among the client proteins, there are several molecules that have been defined as targets or factors for determination or enhancement of radiosensitivity or thermosensitivity. Thus, it is easily speculated that Hsp90 chaperone complex inhibitors that disrupt association of Hsp90 and client protein in combination with radiation or/and heat has potential effect on enhancement of radiosensitivity or thermosensitivity. In this paper, possible mechanisms in enhancing radiosensitivity or thermosensitivity according to the client proteins will be summarized. (author)

  18. Selection and Characterization of Single Chain Antibody Fragments Specific for Hsp90 as a Potential Cancer Targeting Molecule

    Directory of Open Access Journals (Sweden)

    Edyta Petters

    2015-08-01

    Full Text Available Heat shock proteins play an essential role in facilitating malignant transformation and they have been recognized as important factors in human cancers. One of the key elements of the molecular chaperones machinery is Hsp90 and it has recently become a target for anticancer therapeutic approaches. The potential and importance of Hsp90-directed agents becomes apparent when one realizes that disruption of Hsp90 function may influence over 200 oncogenic client proteins. Here, we described the selection and characterization of Hsp90-specific antibody fragments from commercially available Tomlinson I and J phage display libraries. The affinities of Hsp90-binding scFv variants were measured using SPR method. Then, based on the best clone selected, we performed the affinity maturation procedure and obtained valuable Hsp90-specific clones. The selected binders were expressed and applied for immunostaining, ELISA and SPR analysis using model cancer cell lines. All performed experiments confirmed the ability of selected antibodies to interact with the Hsp90. Therefore, the presented Hsp90-specific scFv, might be a starting point for the development of a novel antibody-based strategy targeting cancer.

  19. Active participation of Hsp90 in the biogenesis of the trimeric reovirus cell attachment protein sigma1.

    Science.gov (United States)

    Gilmore, R; Coffey, M C; Lee, P W

    1998-06-12

    The reovirus cell attachment protein, sigma1, is a lollipop-shaped homotrimer with an N-terminal fibrous tail and a C-terminal globular head. Biogenesis of this protein involves two trimerization events: N-terminal trimerization, which occurs cotranslationally and is Hsp70/ATP-independent, and C-terminal trimerization, which occurs posttranslationally and is Hsp70/ATP-dependent. To determine if Hsp90 also plays a role in sigma1 biogenesis, we analyzed sigma1 synthesized in rabbit reticulocyte lysate. Coprecipitation experiments using anti-Hsp90 antibodies revealed that Hsp90 was associated with immature sigma1 trimers (hydra-like intermediates with assembled N termini and unassembled C termini) but not with mature trimers. The use of truncated sigma1 further demonstrated that only the C-terminal half of sigma1 associated with Hsp90. In the presence of the Hsp90 binding drug geldanamycin, N-terminal trimerization proceeded normally, but C-terminal trimerization was blocked. Geldanamycin did not inhibit the association of Hsp90 with sigma 1 but prevented the subsequent release of Hsp90 from the immature sigma1 complex. We also examined the status of p23, an Hsp90-associated cochaperone. Like Hsp90, p23 only associated with immature sigma1 trimers, and this association was mapped to the C-terminal half of sigma1. However, unlike Hsp90, p23 was released from the sigma1 complex upon the addition of geldanamycin. These results highlight an all-or-none concept of chaperone involvement in different oligomerization domains within a single protein and suggest a possible common usage of chaperones in the regulation of general protein folding and of steroid receptor activation.

  20. Discovery of an L-alanine ester prodrug of the Hsp90 inhibitor, MPC-3100.

    Science.gov (United States)

    Kim, Se-Ho; Tangallapally, Rajendra; Kim, In Chul; Trovato, Richard; Parker, Daniel; Patton, J Scott; Reeves, Leslie; Bradford, Chad; Wettstein, Daniel; Baichwal, Vijay; Papac, Damon; Bajji, Ashok; Carlson, Robert; Yager, Kraig M

    2015-11-15

    Various types of Hsp90 inhibitors have been and continue to undergo clinical investigation. One development candidate is the purine-based, synthetic Hsp90 inhibitor 1 (MPC-3100), which successfully completed a phase I clinical study. However, further clinical development of 1 was hindered by poor solubility and consequent formulation issues and promoted development of a more water soluble prodrug. Towards this end, numerous pro-moieties were explored in vitro and in vivo. These studies resulted in identification of L-alanine ester mesylate, 2i (MPC-0767), which exhibited improved aqueous solubility, adequate chemical stability, and rapid bioconversion without the need for solubilizing excipients. Based on improved physical characteristics and favorable PK and PD profiles, 2i mesylate was selected for further development. A convergent, scalable, chromatography-free synthesis for 2i mesylate was developed to support further clinical evaluation. Copyright © 2015 Elsevier Ltd. All rights reserved.

  1. Mitochondrial carrier protein biogenesis: role of the chaperones Hsc70 and Hsp90.

    Science.gov (United States)

    Zara, Vincenzo; Ferramosca, Alessandra; Robitaille-Foucher, Philippe; Palmieri, Ferdinando; Young, Jason C

    2009-04-15

    Metabolite carrier proteins of the mitochondrial inner membrane share homology in their transmembrane domains, which also carries their targeting information. In addition, some carriers have cleavable presequences which are not essential for targeting, but have some other function before import. The cytosolic chaperones Hsc70 (heat-shock cognate 70) and Hsp90 (heat-shock protein 90) complex with carrier precursors and interact specifically with the Tom (translocase of the mitochondrial outer membrane) 70 import receptor to promote import. We analysed how the presequences of the PiC (phosphate carrier) and CIC (citrate carrier) relate to the mechanisms of chaperone-mediated import. Deletion of the PiC presequence reduced the efficiency of import but, notably, not by causing aggregation. Instead, binding of the protein to Hsc70 was reduced, as well as the dependence on Hsc70 for import. Hsp90 binding and function in import was not greatly affected, but it could not entirely compensate for the lack of Hsc70 interaction. Deletion of the presequence from CIC was shown to cause its aggregation, but had little effect on the contribution to import of either Hsc70 or Hsp90. The presequence of PiC, but not that of CIC, conferred Hsc70 binding to dihydrofolate reductase fusion proteins. In comparison, OGC (oxoglutarate carrier) lacks a presequence and was more soluble, though it is still dependent on both Hsc70 and Hsp90. We propose that carrier presequences evolved to improve targeting competence by different mechanisms, depending on physical properties of the precursors in the cytosolic targeting environment.

  2. Plasma Hsp90 Level as a Marker of Early Acute Lymphoblastic Leukemia Engraftment and Progression in Mice.

    Directory of Open Access Journals (Sweden)

    Mateus Milani

    Full Text Available Current monitoring of acute lymphoblastic leukemia (ALL in living mice is based on FACS analysis of blood hCD45+ cells. In this work, we evaluated the use of human IGFBP2, B2M or Hsp90 as soluble markers of leukemia. ELISA for B2M and IGFBP2 resulted in high background levels in healthy animals, precluding its use. Conversely, plasma levels of Hsp90 showed low background and linear correlation to FACS results. In another experiment, we compared Hsp90 levels with percentage of hCD45+ cells in blood, bone marrow, liver and spleen of animals weekly sacrificed. Hsp90 levels proved to be a superior method for the earlier detection of ALL engraftment and correlated linearly to ALL burden and progression in all compartments, even at minimal residual disease levels. Importantly, the Hsp90/hCD45+ ratio was not altered when animals were treated with dexamethasone or a PI3K inhibitor, indicating that chemotherapy does not directly interfere with leukemia production of Hsp90. In conclusion, plasma Hsp90 was validated as a soluble biomarker of ALL, useful for earlier detection of leukemia engraftment, monitoring leukemia kinetics at residual disease levels, and pre-clinical or mouse avatar evaluations of anti-leukemic drugs.

  3. Plasma Hsp90 Level as a Marker of Early Acute Lymphoblastic Leukemia Engraftment and Progression in Mice

    Science.gov (United States)

    de Vasconcellos, Jaíra Ferreira; Brandalise, Silvia Regina; Nowill, Alexandre Eduardo; Yunes, José Andrés

    2015-01-01

    Current monitoring of acute lymphoblastic leukemia (ALL) in living mice is based on FACS analysis of blood hCD45+ cells. In this work, we evaluated the use of human IGFBP2, B2M or Hsp90 as soluble markers of leukemia. ELISA for B2M and IGFBP2 resulted in high background levels in healthy animals, precluding its use. Conversely, plasma levels of Hsp90 showed low background and linear correlation to FACS results. In another experiment, we compared Hsp90 levels with percentage of hCD45+ cells in blood, bone marrow, liver and spleen of animals weekly sacrificed. Hsp90 levels proved to be a superior method for the earlier detection of ALL engraftment and correlated linearly to ALL burden and progression in all compartments, even at minimal residual disease levels. Importantly, the Hsp90/hCD45+ ratio was not altered when animals were treated with dexamethasone or a PI3K inhibitor, indicating that chemotherapy does not directly interfere with leukemia production of Hsp90. In conclusion, plasma Hsp90 was validated as a soluble biomarker of ALL, useful for earlier detection of leukemia engraftment, monitoring leukemia kinetics at residual disease levels, and pre-clinical or mouse avatar evaluations of anti-leukemic drugs. PMID:26068922

  4. Sensitization of multidrug-resistant human cancer cells to Hsp90 inhibitors by down-regulation of SIRT1

    Science.gov (United States)

    Kim, Hak-Bong; Lee, Su-Hoon; Um, Jee-Hyun; Oh, Won Keun; Kim, Dong-Wan; Kang, Chi-Dug; Kim, Sun-Hee

    2015-01-01

    The effectiveness of Hsp90 inhibitors as anticancer agents was limited in multidrug-resistant (MDR) human cancer cells due to induction of heat shock proteins (Hsps) such as Hsp70/Hsp27 and P-glycoprotein (P-gp)-mediated efflux. In the present study, we showed that resistance to Hsp90 inhibitors of MDR human cancer cells could be overcome with SIRT1 inhibition. SIRT1 knock-down or SIRT1 inhibitors (amurensin G and EX527) effectively suppressed the resistance to Hsp90 inhibitors (17-AAG and AUY922) in several MDR variants of human lymphoblastic leukemia and human breast cancer cell lines. SIRT1 inhibition down-regulated the expression of heat shock factor 1 (HSF1) and subsequently Hsps and facilitated Hsp90 multichaperone complex disruption via hyperacetylation of Hsp90/Hsp70. These findings were followed by acceleration of ubiquitin ligase CHIP-mediated mutant p53 (mut p53) degradation and subsequent down-regulation of P-gp in 17-AAG-treated MDR cancer cells expressing P-gp and mut p53 after inhibition of SIRT1. Therefore, combined treatment with Hsp90 inhibitor and SIRT1 inhibitor could be a more effective therapeutic approach for Hsp90 inhibitor-resistant MDR cells via down-regulation of HSF1/Hsps, mut p53 and P-gp. PMID:26416354

  5. Resolving Hot Spots in the C-Terminal Dimerization Domain that Determine the Stability of the Molecular Chaperone Hsp90

    Science.gov (United States)

    Reimann, Sven; Smits, Sander H. J.; Schmitt, Lutz; Groth, Georg; Gohlke, Holger

    2014-01-01

    Human heat shock protein of 90 kDa (hHsp90) is a homodimer that has an essential role in facilitating malignant transformation at the molecular level. Inhibiting hHsp90 function is a validated approach for treating different types of tumors. Inhibiting the dimerization of hHsp90 via its C-terminal domain (CTD) should provide a novel way to therapeutically interfere with hHsp90 function. Here, we predicted hot spot residues that cluster in the CTD dimerization interface by a structural decomposition of the effective energy of binding computed by the MM-GBSA approach and confirmed these predictions using in silico alanine scanning with DrugScorePPI. Mutation of these residues to alanine caused a significant decrease in the melting temperature according to differential scanning fluorimetry experiments, indicating a reduced stability of the mutant hHsp90 complexes. Size exclusion chromatography and multi-angle light scattering studies demonstrate that the reduced stability of the mutant hHsp90 correlates with a lower complex stoichiometry due to the disruption of the dimerization interface. These results suggest that the identified hot spot residues can be used as a pharmacophoric template for identifying and designing small-molecule inhibitors of hHsp90 dimerization. PMID:24760083

  6. MEK inhibition potentiates the activity of Hsp90 inhibitor 17-AAG against pancreatic cancer cells.

    Science.gov (United States)

    Zhang, Tao; Li, Yanyan; Zhu, Zhenkun; Gu, Mancang; Newman, Bryan; Sun, Duxin

    2010-10-04

    The Ras/Raf/MEK/ERK signaling has been implicated in uncontrolled cell proliferation and tumor progression in pancreatic cancer. The purpose of this study is to evaluate the antitumor activity of MEK inhibitor U0126 in combination with Hsp90 inhibitor 17-allylamino-17-demethoxygeldanamycin (17-AAG) in pancreatic cancer cells. Western blotting showed that 17-AAG caused a 2- to 3-fold transient activation of MEK/ERK signaling in pancreatic cancer cells. The activation sustained for 6 h before phospho-ERK (p-ERK) destabilization. The selective MEK inhibitor U0126 completely abolished 17-AAG induced ERK1/2 activation and resulted in more than 80% of phospho-ERK degradation after only 15 min treatment. Moreover, U0126 had complementary effect on 17-AAG regulated oncogenic and cell cycle related proteins. Although 17-AAG downregulated cyclin D1, cyclin E, CDK4 and CDK6, it led to cyclin A and CDK2 accumulation, which was reversed by the addition of U0126. Antiproliferation assay showed that combination of U0126 and 17-AAG resulted in synergistic cytotoxic effect. More importantly, 17-AAG alone only exhibited moderate inhibition of cell migration in vitro, while addition of U0126 dramatically enhanced the inhibitory effect by 2- to 5-fold. Taken together, these data demonstrate that MEK inhibitor U0126 potentiates the activity of Hsp90 inhibitor 17-AAG against pancreatic cancer cells. The combination of Hsp90 and MEK inhibition could provide a promising avenue for the treatment of pancreatic cancer.

  7. Discovery of new molecular entities able to strongly interfere with Hsp90 C-terminal domain.

    Science.gov (United States)

    Terracciano, Stefania; Russo, Alessandra; Chini, Maria G; Vaccaro, Maria C; Potenza, Marianna; Vassallo, Antonio; Riccio, Raffaele; Bifulco, Giuseppe; Bruno, Ines

    2018-01-26

    Heat shock protein 90 (Hsp90) is an ATP dependent molecular chaperone deeply involved in the complex network of cellular signaling governing some key functions, such as cell proliferation and survival, invasion and angiogenesis. Over the past years the N-terminal protein domain has been fully investigated as attractive strategy against cancer, but despite the many efforts lavished in the field, none of the N-terminal binders (termed "classical inhibitors"), currently in clinical trials, have yet successfully reached the market, because of the detrimental heat shock response (HSR) that showed to induce; thus, recently, the selective inhibition of Hsp90 C-terminal domain has powerfully emerged as a more promising alternative strategy for anti-cancer therapy, not eliciting this cell rescue cascade. However, the structural complexity of the target protein and, mostly, the lack of a co-crystal structure of C-terminal domain-ligand, essential to drive the identification of new hits, represent the largest hurdles in the development of new selective C-terminal inhibitors. Continuing our investigations on the identification of new anticancer drug candidates, by using an orthogonal screening approach, here we describe two new potent C-terminal inhibitors able to induce cancer cell death and a considerable down-regulation of Hsp90 client oncoproteins, without triggering the undesired heat shock response.

  8. Hsp90 inhibitors, part 1: definition of 3-D QSAutogrid/R models as a tool for virtual screening.

    Science.gov (United States)

    Ballante, Flavio; Caroli, Antonia; Wickersham, Richard B; Ragno, Rino

    2014-03-24

    The multichaperone heat shock protein (Hsp) 90 complex mediates the maturation and stability of a variety of oncogenic signaling proteins. For this reason, Hsp90 has emerged as a promising target for anticancer drug development. Herein, we describe a complete computational procedure for building several 3-D QSAR models used as a ligand-based (LB) component of a comprehensive ligand-based (LB) and structure-based (SB) virtual screening (VS) protocol to identify novel molecular scaffolds of Hsp90 inhibitors. By the application of the 3-D QSAutogrid/R method, eight SB PLS 3-D QSAR models were generated, leading to a final multiprobe (MP) 3-D QSAR pharmacophoric model capable of recognizing the most significant chemical features for Hsp90 inhibition. Both the monoprobe and multiprobe models were optimized, cross-validated, and tested against an external test set. The obtained statistical results confirmed the models as robust and predictive to be used in a subsequent VS.

  9. Repercussion of mitochondria deformity induced by anti-Hsp90 drug 17AAG in human tumor cells

    KAUST Repository

    Vishal, Chaturvedi; Kumar, Jonnala Ujwal; Veera Brahmendra Swamy, Cherukuvada; Nandini, Rangaraj; Srinivas, Gunda; Kumaresan, Rathinam; Shashi, Singh; Sreedhar, Amere Subbarao

    2011-01-01

    Inhibiting Hsp90 chaperone roles using 17AAG induces cytostasis or apoptosis in tumor cells through destabilization of several mutated cancer promoting proteins. Although mitochondria are central in deciding the fate of cells, 17AAG induced effects

  10. A Novel Therapeutic Strategy for the Treatment of Glioma, Combining Chemical and Molecular Targeting of Hsp90α

    International Nuclear Information System (INIS)

    Mehta, Adi; Shervington, Leroy; Munje, Chinmay; Shervington, Amal

    2011-01-01

    Hsp90α's vital role in tumour survival and progression, together with its highly inducible expression profile in gliomas and its absence in normal tissue and cell lines validates it as a therapeutic target for glioma. Hsp90α was downregulated using the post-transcriptional RNAi strategy (sihsp90α) and a post-translational inhibitor, the benzoquinone antibiotic 17-AAG. Glioblastoma U87-MG and normal human astrocyte SVGp12 were treated with sihsp90α, 17-AAG and concurrent sihsp90α/17-AAG (combined treatment). Both Hsp90α gene silencing and the protein inhibitor approaches resulted in a dramatic reduction in cell viability. Results showed that sihsp90α, 17-AAG and a combination of sihsp90α/17-AAG, reduced cell viability by 27%, 75% and 88% (p < 0.001), respectively, after 72 h. hsp90α mRNA copy numbers were downregulated by 65%, 90% and 99% after 72 h treatment with sihsp90α, 17-AAG and sihsp90α/17-AAG, respectively. The relationship between Hsp90α protein expression and its client Akt kinase activity levels were monitored following treatment with sihsp90α, 17-AAG and sihsp90α/17-AAG. Akt kinase activity was downregulated as a direct consequence of Hsp90α inhibition. Both Hsp90α and Akt kinase levels were significantly downregulated after 72 h. Although, 17-AAG when used as a single agent reduces the Hsp90α protein and the Akt kinase levels, the efficacy demonstrated by combinatorial treatment was found to be far more effective. Combination treatment reduced the Hsp90α protein and Akt kinase levels to 4.3% and 43%, respectively, after 72 h. hsp90α mRNA expression detected in SVGp12 was negligible compared to U87-MG, also, the combination treatment did not compromise the normal cell viability. Taking into account the role of Hsp90α in tumour progression and the involvement of Akt kinase in cell signalling and the anti-apoptotic pathways in tumours, this double targets treatment infers a novel therapeutic strategy

  11. C-terminal sequences of hsp70 and hsp90 as non-specific anchors for tetratricopeptide repeat (TPR) proteins.

    Science.gov (United States)

    Ramsey, Andrew J; Russell, Lance C; Chinkers, Michael

    2009-10-12

    Steroid-hormone-receptor maturation is a multi-step process that involves several TPR (tetratricopeptide repeat) proteins that bind to the maturation complex via the C-termini of hsp70 (heat-shock protein 70) and hsp90 (heat-shock protein 90). We produced a random T7 peptide library to investigate the roles played by the C-termini of the two heat-shock proteins in the TPR-hsp interactions. Surprisingly, phages with the MEEVD sequence, found at the C-terminus of hsp90, were not recovered from our biopanning experiments. However, two groups of phages were isolated that bound relatively tightly to HsPP5 (Homo sapiens protein phosphatase 5) TPR. Multiple copies of phages with a C-terminal sequence of LFG were isolated. These phages bound specifically to the TPR domain of HsPP5, although mutation studies produced no evidence that they bound to the domain's hsp90-binding groove. However, the most abundant family obtained in the initial screen had an aspartate residue at the C-terminus. Two members of this family with a C-terminal sequence of VD appeared to bind with approximately the same affinity as the hsp90 C-12 control. A second generation pseudo-random phage library produced a large number of phages with an LD C-terminus. These sequences acted as hsp70 analogues and had relatively low affinities for hsp90-specific TPR domains. Unfortunately, we failed to identify residues near hsp90's C-terminus that impart binding specificity to individual hsp90-TPR interactions. The results suggest that the C-terminal sequences of hsp70 and hsp90 act primarily as non-specific anchors for TPR proteins.

  12. Targeting GRP75 improves HSP90 inhibitor efficacy by enhancing p53-mediated apoptosis in hepatocellular carcinoma.

    Directory of Open Access Journals (Sweden)

    Weiwei Guo

    Full Text Available Heat shock protein 90 (HSP90 inhibitors are potential drugs for cancer therapy. The inhibition of HSP90 on cancer cell growth largely through degrading client proteins, like Akt and p53, therefore, triggering cancer cell apoptosis. Here, we show that the HSP90 inhibitor 17-AAG can induce the expression of GRP75, a member of heat shock protein 70 (HSP70 family, which, in turn, attenuates the anti-growth effect of HSP90 inhibition on cancer cells. Additionally, 17-AAG enhanced binding of GRP75 and p53, resulting in the retention of p53 in the cytoplasm. Blocking GRP75 with its inhibitor MKT-077 potentiated the anti-tumor effects of 17-AAG by disrupting the formation of GRP75-p53 complexes, thereby facilitating translocation of p53 into the nuclei and leading to the induction of apoptosis-related genes. Finally, dual inhibition of HSP90 and GRP75 was found to significantly inhibit tumor growth in a liver cancer xenograft model. In conclusion, the GRP75 inhibitor MKT-077 enhances 17-AAG-induced apoptosis in HCCs and increases p53-mediated inhibition of tumor growth in vivo. Dual targeting of GRP75 and HSP90 may be a useful strategy for the treatment of HCCs.

  13. SNX-25a, a novel Hsp90 inhibitor, inhibited human cancer growth more potently than 17-AAG.

    Science.gov (United States)

    Wang, Shaoxiang; Wang, Xiao; Du, Zhan; Liu, Yuting; Huang, Dane; Zheng, Kai; Liu, Kaisheng; Zhang, Yi; Zhong, Xueyun; Wang, Yifei

    2014-07-18

    17-Allylamino-17-demethoxygeldanamycin (17-AAG), a typical Hsp90 inhibitor derived from geldanamycin (GA), has entered Phase III clinical trials for cancer therapy. However, it has several significant limitations such as poor solubility, limited bioavailability and unacceptable hepatotoxicity. In this study, the anticancer activity and mechanism of SNX-25a, a novel Hsp90 inhibitor, was investigated comparing with that of 17-AAG. We showed that SNX-25a triggered growth inhibition more sensitively than 17-AAG against many human cancer cells, including K562, SW-620, A375, Hep-2, MCF-7, HepG2, HeLa, and A549 cell lines, especially at low concentrations (AAG, SNX-25a was more potent in arresting the cell cycle at G2 phase, and displayed more potent effects on human cancer cell apoptosis and Hsp90 client proteins. It also exhibited a stronger binding affinity to Hsp90 than 17-AAG using molecular docking. Considering the superiority effects on Hsp90 affinity, cell growth, cell cycle, apoptosis, and Hsp90 client proteins, SNX-25a is supposed as a potential anticancer agent that needs to be explored in detail. Copyright © 2014 Elsevier Inc. All rights reserved.

  14. Comparison of inhibitory effects of 17-AAG nanoparticles and free 17-AAG on HSP90 gene expression in breast cancer.

    Science.gov (United States)

    Ghalhar, Masoud Gandomkar; Akbarzadeh, Abolfazl; Rahmati, Mohammad; Mellatyar, Hassan; Dariushnejad, Hassan; Zarghami, Nosratallah; Barkhordari, Amin

    2014-01-01

    HSP90 may be overexpressed in cancer cells which are greatly dependent on Hsp90 function. Geldanamycin derivative 17 allylamino-17-demethoxygeldanamycin (17-AAG) inhibits the function and expression of HSP90. 17-AAG has poor water-solubility which is a potential problem for clinical practice. In this study for improving the stability and solubility of molecules in drug delivery systems we used a β-cyclodextrin- 17AAG complex. To assess cytotoxic effects of β-cyclodextrin-17AAG complexes and free 17AAG, colorimetric cell viability (MTT) assays were performed. Cells were treated with equal concentrations of β-cyclodextrin- 17AAG complex and free 17AAG and Hsp90 gene expression levels in the two groups was compared by real-time PCR. MTT assay confirmed that β-cyclodextrin- 17AAG complex enhanced 17AAG cytotoxicity and drug delivery in T47D breast cancer cells. The level of Hsp90 gene expression in cells treated with β-cyclodextrin- 17AAG complex was lower than that of cells treated with free 17AAG (P=0.001). The results demonstrated that β-cyclodextrin- 17AAG complexes are more effective than free 17AAG in down-regulating HSP90 expression due to enhanced β-cyclodextrin-17AAG uptake by cells. Therefore, β-cyclodextrin could be superior carrier for this kind of hydrophobic agent.

  15. Conformational Activation of Argonaute by Distinct yet Coordinated Actions of the Hsp70 and Hsp90 Chaperone Systems.

    Science.gov (United States)

    Tsuboyama, Kotaro; Tadakuma, Hisashi; Tomari, Yukihide

    2018-05-17

    Loading of small RNAs into Argonaute, the core protein in RNA silencing, requires the Hsp70/Hsp90 chaperone machinery. This machinery also activates many other clients, including steroid hormone receptors and kinases, but how their structures change during chaperone-dependent activation remains unclear. Here, we utilized single-molecule Förster resonance energy transfer (smFRET) to probe the conformational changes of Drosophila Ago2 mediated by the chaperone machinery. We found that empty Ago2 exists in various closed conformations. The Hsp70 system (Hsp40 and Hsp70) and the Hsp90 system (Hop, Hsp90, and p23) together render Ago2 into an open, active form. The Hsp70 system, but not the Hsp90 system alone, is sufficient for Ago2 to partially populate the open form. Instead, the Hsp90 system is required to extend the dwell time of Ago2 in the open state, which must be transiently primed by the Hsp70 system. Our data uncover distinct and coordinated actions of the chaperone machinery, where the Hsp70 system expands the structural ensembles of Ago2 and the Hsp90 system captures and stabilizes the active form. Copyright © 2018 Elsevier Inc. All rights reserved.

  16. A Novel Pulse-Chase SILAC Strategy Measures Changes in Protein Decay and Synthesis Rates Induced by Perturbation of Proteostasis with an Hsp90 Inhibitor

    Science.gov (United States)

    Fierro-Monti, Ivo; Racle, Julien; Hernandez, Celine; Waridel, Patrice; Hatzimanikatis, Vassily; Quadroni, Manfredo

    2013-01-01

    Standard proteomics methods allow the relative quantitation of levels of thousands of proteins in two or more samples. While such methods are invaluable for defining the variations in protein concentrations which follow the perturbation of a biological system, they do not offer information on the mechanisms underlying such changes. Expanding on previous work [1], we developed a pulse-chase (pc) variant of SILAC (stable isotope labeling by amino acids in cell culture). pcSILAC can quantitate in one experiment and for two conditions the relative levels of proteins newly synthesized in a given time as well as the relative levels of remaining preexisting proteins. We validated the method studying the drug-mediated inhibition of the Hsp90 molecular chaperone, which is known to lead to increased synthesis of stress response proteins as well as the increased decay of Hsp90 “clients”. We showed that pcSILAC can give information on changes in global cellular proteostasis induced by treatment with the inhibitor, which are normally not captured by standard relative quantitation techniques. Furthermore, we have developed a mathematical model and computational framework that uses pcSILAC data to determine degradation constants kd and synthesis rates Vs for proteins in both control and drug-treated cells. The results show that Hsp90 inhibition induced a generalized slowdown of protein synthesis and an increase in protein decay. Treatment with the inhibitor also resulted in widespread protein-specific changes in relative synthesis rates, together with variations in protein decay rates. The latter were more restricted to individual proteins or protein families than the variations in synthesis. Our results establish pcSILAC as a viable workflow for the mechanistic dissection of changes in the proteome which follow perturbations. Data are available via ProteomeXchange with identifier PXD000538. PMID:24312217

  17. A novel pulse-chase SILAC strategy measures changes in protein decay and synthesis rates induced by perturbation of proteostasis with an Hsp90 inhibitor.

    Directory of Open Access Journals (Sweden)

    Ivo Fierro-Monti

    Full Text Available Standard proteomics methods allow the relative quantitation of levels of thousands of proteins in two or more samples. While such methods are invaluable for defining the variations in protein concentrations which follow the perturbation of a biological system, they do not offer information on the mechanisms underlying such changes. Expanding on previous work [1], we developed a pulse-chase (pc variant of SILAC (stable isotope labeling by amino acids in cell culture. pcSILAC can quantitate in one experiment and for two conditions the relative levels of proteins newly synthesized in a given time as well as the relative levels of remaining preexisting proteins. We validated the method studying the drug-mediated inhibition of the Hsp90 molecular chaperone, which is known to lead to increased synthesis of stress response proteins as well as the increased decay of Hsp90 "clients". We showed that pcSILAC can give information on changes in global cellular proteostasis induced by treatment with the inhibitor, which are normally not captured by standard relative quantitation techniques. Furthermore, we have developed a mathematical model and computational framework that uses pcSILAC data to determine degradation constants kd and synthesis rates Vs for proteins in both control and drug-treated cells. The results show that Hsp90 inhibition induced a generalized slowdown of protein synthesis and an increase in protein decay. Treatment with the inhibitor also resulted in widespread protein-specific changes in relative synthesis rates, together with variations in protein decay rates. The latter were more restricted to individual proteins or protein families than the variations in synthesis. Our results establish pcSILAC as a viable workflow for the mechanistic dissection of changes in the proteome which follow perturbations. Data are available via ProteomeXchange with identifier PXD000538.

  18. Small molecule inhibitor screening identifified HSP90 inhibitor 17-AAG as potential therapeutic agent for gallbladder cancer.

    Science.gov (United States)

    Weber, Helga; Valbuena, José R; Barbhuiya, Mustafa A; Stein, Stefan; Kunkel, Hana; García, Patricia; Bizama, Carolina; Riquelme, Ismael; Espinoza, Jaime A; Kurtz, Stephen E; Tyner, Jeffrey W; Calderon, Juan Francisco; Corvalán, Alejandro H; Grez, Manuel; Pandey, Akhilesh; Leal-Rojas, Pamela; Roa, Juan C

    2017-04-18

    Gallbladder cancer (GBC) is a lethal cancer with poor prognosis associated with high invasiveness and poor response to chemotherapy and radiotherapy. New therapeutic approaches are urgently needed in order to improve survival and response rates of GBC patients. We screened 130 small molecule inhibitors on a panel of seven GBC cell lines and identified the HSP90 inhibitor 17-AAG as one of the most potent inhibitory drugs across the different lines. We tested the antitumor efficacy of 17-AAG and geldanamycin (GA) in vitro and in a subcutaneous preclinical tumor model NOD-SCID mice. We also evaluated the expression of HSP90 by immunohistochemistry in human GBC tumors.In vitro assays showed that 17-AAG and GA significantly reduced the expression of HSP90 target proteins, including EGFR, AKT, phospho-AKT, Cyclin B1, phospho-ERK and Cyclin D1. These molecular changes were consistent with reduced cell viability and cell migration and promotion of G2/M cell cycle arrest and apoptosis observed in our in vitro studies.In vivo, 17-AAG showed efficacy in reducing subcutaneous tumors size, exhibiting a 69.6% reduction in tumor size in the treatment group compared to control mice (p < 0.05).The HSP90 immunohistochemical staining was seen in 182/209 cases of GBC (87%) and it was strongly expressed in 70 cases (33%), moderately in 58 cases (28%), and weakly in 54 cases (26%).Our pre-clinical observations strongly suggest that the inhibition of HSP90 function by HSP90 inhibitors is a promising therapeutic strategy for gallbladder cancer that may benefit from new HSP90 inhibitors currently in development.

  19. Can RNAi-mediated hsp90α knockdown in combination with 17-AAG be a therapy for glioma?

    Science.gov (United States)

    Mehta, Adi; Shervington, Amal; Howl, John; Jones, Sarah; Shervington, Leroy

    2013-01-01

    Heat shock protein 90 promotes tumor progression and survival and has emerged as a vital therapeutic target. Previously we reported that the combinatorial treatment of 17AAG/sihsp90α significantly downregulated Hsp90α mRNA and protein levels in Glioblastoma Multiforme (GBM). Here we investigated the ability of cell penetrating peptide (Tat48-60 CPP)-mediated siRNA-induced hsp90α knockdown as a single agent and in combination with 17-allylamino-17-demethoxygeldanamycin (17-AAG) to induce tumor growth inhibition in GBM and whether it possessed therapeutic implications. GBM and non-tumorigenic cells exposed to siRNA and/or 17-AAG were subsequently assessed by qRT-PCR, immunofluorescence, FACS analysis, quantitative Akt, LDH leakage and cell viability assays. PAGE was performed for serum stability assessment. A combination of siRNA/17-AAG treatment significantly induced Hsp90α gene and protein knockdown by 95% and 98%, respectively, concomitant to 84% Akt kinase activity attenuation, induced cell cycle arrest and tumor-specific cytotoxicity by 88%. Efficient complex formation between CPP and siRNA exhibited improved serum stability of the siRNA with minimal intrinsic toxicity in vitro. The preliminary in vivo results showed that combination therapy induced hsp90α knockdown and attenuated Akt kinase activity in intracranial glioblastoma mouse models. The results imply that RNAi-mediated hsp90α knockdown increases 17-AAG treatment efficacy in GBM. In addition, the cytotoxic response observed was the consequence of downregulation of hsp90α gene expression, reduced Akt kinase activity and S-G2/M cell cycle arrest. These results are novel and highlight the ability of Tat to efficiently deliver siRNA in GBM and suggest that the dual inhibition of Hsp90 has therapeutic potentials.

  20. NAD(P)H:Quinone Oxidoreductase-1 Expression Sensitizes Malignant Melanoma Cells to the HSP90 Inhibitor 17-AAG.

    Science.gov (United States)

    Kasai, Shuya; Arakawa, Nobuyuki; Okubo, Ayaka; Shigeeda, Wataru; Yasuhira, Shinji; Masuda, Tomoyuki; Akasaka, Toshihide; Shibazaki, Masahiko; Maesawa, Chihaya

    2016-01-01

    The KEAP1-NRF2 pathway regulates cellular redox homeostasis by transcriptional induction of genes associated with antioxidant synthesis and detoxification in response to oxidative stress. Previously, we reported that KEAP1 mutation elicits constitutive NRF2 activation and resistance to cisplatin (CDDP) and dacarbazine (DTIC) in human melanomas. The present study was conducted to clarify whether an HSP90 inhibitor, 17-AAG, efficiently eliminates melanoma with KEAP1 mutation, as the NRF2 target gene, NQO1, is a key enzyme in 17-AAG bioactivation. In melanoma and non-small cell lung carcinoma cell lines with or without KEAP1 mutations, NQO1 expression and 17-AAG sensitivity are inversely correlated. NQO1 is highly expressed in normal melanocytes and in several melanoma cell lines despite the presence of wild-type KEAP1, and the NQO1 expression is dependent on NRF2 activation. Because either CDDP or DTIC produces reactive oxygen species that activate NRF2, we determined whether these agents would sensitize NQO1-low melanoma cells to 17-AAG. Synergistic cytotoxicity of the 17-AAG and CDDP combination was detected in four out of five NQO1-low cell lines, but not in the cell line with KEAP1 mutation. These data indicate that 17-AAG could be a potential chemotherapeutic agent for melanoma with KEAP1 mutation or NQO1 expression.

  1. 17-AAG, an Hsp90 inhibitor, causes kinetochore defects: a novel mechanism by which 17-AAG inhibits cell proliferation.

    Science.gov (United States)

    Niikura, Y; Ohta, S; Vandenbeldt, K J; Abdulle, R; McEwen, B F; Kitagawa, K

    2006-07-13

    The Hsp90 inhibitor 17-allylaminogeldanamycin (17-AAG), which is currently in clinical trials, is thought to exert antitumor activity by simultaneously targeting several oncogenic signaling pathways. Here we report a novel mechanism by which 17-AAG inhibits cell proliferation, and we provide the first evidence that HSP90 is required for the assembly of kinetochore protein complexes in humans. 17-AAG caused delocalization of several kinetochore proteins including CENP-I and CENP-H but excluding CENP-B and CENP-C. Consistently, 17-AAG induced a mitotic arrest that depends on the spindle checkpoint and induced misalignment of chromosomes and aneuploidy. We found that HSP90 associates with SGT1 (suppressor of G2 allele of skp1; SUGT1) in human cells and that depletion of SGT1 sensitizes HeLa cells to 17-AAG. Overexpression of SGT1 restored the localization of specific kinetochore proteins and chromosome alignment in cells treated with 17-AAG. Biochemical and genetic results suggest that HSP90, through its interaction with SGT1 (SUGT1), is required for kinetochore assembly. Furthermore, time-course experiments revealed that transient treatment with 17-AAG between late S and G2/M phases causes substantial delocalization of CENP-H and CENP-I, a finding that strongly suggests that HSP90 participates in kinetochore assembly in a cell cycle-dependent manner.

  2. Hsp90 inhibitor 17-AAG inhibits progression of LuCaP35 xenograft prostate tumors to castration resistance.

    Science.gov (United States)

    O'Malley, Katherine J; Langmann, Gabrielle; Ai, Junkui; Ramos-Garcia, Raquel; Vessella, Robert L; Wang, Zhou

    2012-07-01

    Advanced prostate cancer is currently treated with androgen deprivation therapy (ADT). ADT initially results in tumor regression; however, all patients eventually relapse with castration-resistant prostate cancer. New approaches to delay the progression of prostate cancer to castration resistance are in desperate need. This study addresses whether targeting Heat shock protein 90 (HSP90) regulation of androgen receptor (AR) can inhibit prostate cancer progression to castration resistance. The HSP90 inhibitor 17-AAG was injected intraperitoneally into nude mice bearing LuCaP35 xenograft tumors to determine the effect of HSP90 inhibition on prostate cancer progression to castration resistance and host survival. Administration of 17-AAG maintained androgen-sensitivity, delayed the progression of LuCaP35 xenograft tumors to castration resistance, and prolonged the survival of host. In addition, 17-AAG prevented nuclear localization of endogenous AR in LuCaP35 xenograft tumors in castrated nude mice. Targeting Hsp90 or the mechanism by which HSP90 regulates androgen-independent AR nuclear localization and activation may lead to new approaches to prevent and/or treat castration-resistant prostate cancer. Copyright © 2011 Wiley Periodicals, Inc.

  3. Wild-type EGFR Is Stabilized by Direct Interaction with HSP90 in Cancer Cells and Tumors

    Directory of Open Access Journals (Sweden)

    Aarif Ahsan

    2012-08-01

    Full Text Available The epidermal growth factor receptor (EGFR has been targeted for inhibition using tyrosine kinase inhibitors and monoclonal antibodies, with improvement in outcome in subsets of patients with head and neck, lung, and colorectal carcinomas. We have previously found that EGFR stability plays a key role in cell survival after chemotherapy and radiotherapy. Heat shock protein 90 (HSP90 is known to stabilize mutant EGFR and ErbB2, but its role in cancers with wild-type (WT WT-EGFR is unclear. In this report, we demonstrate that fully mature, membrane-bound WT-EGFR interacts with HSP90 independent of ErbB2. Further, the HSP90 inhibitors geldanamycin (GA and AT13387 cause a decrease in WT-EGFR in cultured head and neck cancer cells. This decrease results from a significantly reduced half-life of WT-EGFR. WT-EGFR was also lost in head and neck xenograft specimens after treatment with AT13387 under conditions that inhibited tumor growth and prolonged survival of the mice. Our findings demonstrate that WT-EGFR is a client protein of HSP90 and that their interaction is critical for maintaining both the stability of the receptor as well as the growth of EGFR-dependent cancers. Furthermore, these findings support the search for specific agents that disrupt HSP90's ability to act as an EGFR chaperone.

  4. Tumour eradication using synchronous thermal ablation and Hsp90 chemotherapy with protein engineered triblock biopolymer-geldanamycin conjugates.

    Science.gov (United States)

    Chen, Yizhe; Youn, Pilju; Pysher, Theodore J; Scaife, Courtney L; Furgeson, Darin Y

    2014-12-01

    Hepatocellular carcinoma (HCC) suffers high tumour recurrence rate after thermal ablation. Heat shock protein 90 (Hsp90) induced post-ablation is critical for tumour survival and progression. A combination therapy of thermal ablation and polymer conjugated Hsp90 chemotherapy was designed and evaluated for complete tumour eradication of HCC. A thermo-responsive, elastin-like polypeptide (ELP)-based tri-block biopolymer was developed and conjugated with a potent Hsp90 inhibitor, geldanamycin (GA). The anti-cancer efficacy of conjugates was evaluated in HCC cell cultures with and without hyperthermia (43 °C). The conjugates were also administered twice weekly in a murine HCC model as a single treatment or in combination with single electrocautery as the ablation method. ELP-GA conjugates displayed enhanced cytotoxicity in vitro and effective heat shock inhibition under hyperthermia. The conjugates alone significantly slowed the tumour growth without systemic toxicity. Four doses of thermo-responsive ELP-GA conjugates with concomitant simple electrocautery accomplished significant Hsp90 inhibition and sustained tumour suppression. Hsp90 inhibition plays a key role in preventing the recurrence of HCC, and the combination of ablation with targeted therapy holds great potential to improve prognosis and survival of HCC patients.

  5. Pharmacological targeting of HSP90 with 17-AAG induces apoptosis of myogenic cells through activation of the intrinsic pathway.

    Science.gov (United States)

    Wagatsuma, Akira; Takayama, Yuzo; Hoshino, Takayuki; Shiozuka, Masataka; Yamada, Shigeru; Matsuda, Ryoichi; Mabuchi, Kunihiko

    2017-12-16

    We have shown that pharmacological inhibition of HSP90 ATPase activity induces apoptosis of myoblasts during their differentiation. However, the signaling pathways remain not fully characterized. We report that pharmacological targeting of HSP90 with 17-AAG activates the intrinsic pathway including caspase-dependent and caspase-independent pathways. 17-AAG induces the typical apoptotic phenotypes including PARP cleavage, chromatin condensation, and nuclear fragmentation with mitochondrial release of cytochrome c, Smac/DIABLO, procaspase-9 processing, and caspase-3 activation. AIF and EndoG redistribute from the mitochondria into the cytosol and are partially translocated to the nucleus in 17-AAG-treated cells. These results suggest that caspase-dependent and caspase-independent pathways should be considered in apoptosis of myogenic cells induced by inhibition of HSP90 ATPase activity.

  6. RECEPTOR SUPERFAMILY OF TUMOR NECROSIS FACTOR Α, AND HSP90 HEAT SHOCK PROTEIN: A MOLECULAR BASIS FOR INTERACTIONS

    Directory of Open Access Journals (Sweden)

    N. V. Ryazantseva

    2011-01-01

    Full Text Available Abstract.  A  study  was  performed  aiming  to  investigate  interactions  between  TNFα  receptor  (TNF1 superfamily and heat shock protein Hsp90, using a Jurkat tumor cell line. The tumor cells cultured in presence of Hsp90 inhibitor (17-AAG showed increased numbers of cells, presenting surface TNFR1 and FasR, which facilitate  triggering  of  programmed  cell  death.  It  was  also  revealed  that  Hsp90  blockage  under  the  in  vitro conditions causes a decrease in FasL, while not affecting TNFα and sTNFR1 production by the tumor cells. (Med. Immunol., 2011, vol. 13, N 2-3, pp 247-252 

  7. Hsc70/Hsp90 chaperone machinery mediates ATP-dependent RISC loading of small RNA duplexes.

    Science.gov (United States)

    Iwasaki, Shintaro; Kobayashi, Maki; Yoda, Mayuko; Sakaguchi, Yuriko; Katsuma, Susumu; Suzuki, Tsutomu; Tomari, Yukihide

    2010-07-30

    Small silencing RNAs--small interfering RNAs (siRNAs) or microRNAs (miRNAs)--direct posttranscriptional gene silencing of their mRNA targets as guides for the RNA-induced silencing complex (RISC). Both siRNAs and miRNAs are born double stranded. Surprisingly, loading these small RNA duplexes into Argonaute proteins, the core components of RISC, requires ATP, whereas separating the two small RNA strands within Argonaute does not. Here we show that the Hsc70/Hsp90 chaperone machinery is required to load small RNA duplexes into Argonaute proteins, but not for subsequent strand separation or target cleavage. We envision that the chaperone machinery uses ATP and mediates a conformational opening of Ago proteins so that they can receive bulky small RNA duplexes. Our data suggest that the chaperone machinery may serve as the driving force for the RISC assembly pathway. Copyright 2010 Elsevier Inc. All rights reserved.

  8. The molecular chaperone Hsp90 is required for cell cycle exit in Drosophila melanogaster.

    Directory of Open Access Journals (Sweden)

    Jennifer L Bandura

    Full Text Available The coordination of cell proliferation and differentiation is crucial for proper development. In particular, robust mechanisms exist to ensure that cells permanently exit the cell cycle upon terminal differentiation, and these include restraining the activities of both the E2F/DP transcription factor and Cyclin/Cdk kinases. However, the full complement of mechanisms necessary to restrain E2F/DP and Cyclin/Cdk activities in differentiating cells are not known. Here, we have performed a genetic screen in Drosophila melanogaster, designed to identify genes required for cell cycle exit. This screen utilized a PCNA-miniwhite(+ reporter that is highly E2F-responsive and results in a darker red eye color when crossed into genetic backgrounds that delay cell cycle exit. Mutation of Hsp83, the Drosophila homolog of mammalian Hsp90, results in increased E2F-dependent transcription and ectopic cell proliferation in pupal tissues at a time when neighboring wild-type cells are postmitotic. Further, these Hsp83 mutant cells have increased Cyclin/Cdk activity and accumulate proteins normally targeted for proteolysis by the anaphase-promoting complex/cyclosome (APC/C, suggesting that APC/C function is inhibited. Indeed, reducing the gene dosage of an inhibitor of Cdh1/Fzr, an activating subunit of the APC/C that is required for timely cell cycle exit, can genetically suppress the Hsp83 cell cycle exit phenotype. Based on these data, we propose that Cdh1/Fzr is a client protein of Hsp83. Our results reveal that Hsp83 plays a heretofore unappreciated role in promoting APC/C function during cell cycle exit and suggest a mechanism by which Hsp90 inhibition could promote genomic instability and carcinogenesis.

  9. The molecular chaperone Hsp90 is required for cell cycle exit in Drosophila melanogaster.

    Science.gov (United States)

    Bandura, Jennifer L; Jiang, Huaqi; Nickerson, Derek W; Edgar, Bruce A

    2013-01-01

    The coordination of cell proliferation and differentiation is crucial for proper development. In particular, robust mechanisms exist to ensure that cells permanently exit the cell cycle upon terminal differentiation, and these include restraining the activities of both the E2F/DP transcription factor and Cyclin/Cdk kinases. However, the full complement of mechanisms necessary to restrain E2F/DP and Cyclin/Cdk activities in differentiating cells are not known. Here, we have performed a genetic screen in Drosophila melanogaster, designed to identify genes required for cell cycle exit. This screen utilized a PCNA-miniwhite(+) reporter that is highly E2F-responsive and results in a darker red eye color when crossed into genetic backgrounds that delay cell cycle exit. Mutation of Hsp83, the Drosophila homolog of mammalian Hsp90, results in increased E2F-dependent transcription and ectopic cell proliferation in pupal tissues at a time when neighboring wild-type cells are postmitotic. Further, these Hsp83 mutant cells have increased Cyclin/Cdk activity and accumulate proteins normally targeted for proteolysis by the anaphase-promoting complex/cyclosome (APC/C), suggesting that APC/C function is inhibited. Indeed, reducing the gene dosage of an inhibitor of Cdh1/Fzr, an activating subunit of the APC/C that is required for timely cell cycle exit, can genetically suppress the Hsp83 cell cycle exit phenotype. Based on these data, we propose that Cdh1/Fzr is a client protein of Hsp83. Our results reveal that Hsp83 plays a heretofore unappreciated role in promoting APC/C function during cell cycle exit and suggest a mechanism by which Hsp90 inhibition could promote genomic instability and carcinogenesis.

  10. Combined BRAF and HSP90 inhibition in patients with unresectable BRAF V600E mutant melanoma.

    Science.gov (United States)

    Eroglu, Zeynep; Chen, Yian Ann; Gibney, Geoffrey T; Weber, Jeffrey S; Kudchadkar, Ragini R; Khushalani, Nikhil I; Markowitz, Joseph; Brohl, Andrew S; Tetteh, Leticia F; Ramadan, Howida; Arnone, Gina; Li, Jiannong; Zhao, Xiuhua; Sharma, Ritin; Darville, Lancia N F; Fang, Bin; Smalley, Inna; Messina, Jane L; Koomen, John M; Sondak, Vernon K; Smalley, Keiran S M

    2018-04-19

    BRAF inhibitors are clinically active in patients with advanced BRAF V600 -mutant melanoma, although acquired resistance remains common. Preclinical studies demonstrated that resistance could be overcome using concurrent treatment with the HSP90 inhibitor XL888. Vemurafenib (960 mg PO BID) combined with escalating doses of XL888 (30, 45, 90 or 135 mg PO twice weekly) was investigated in 21 patients with advanced BRAF V600 -mutant melanoma. Primary endpoints were safety and determination of a maximum tolerated dose. Correlative proteomic studies were performed to confirm HSP inhibitor activity. Objective responses were observed in 15/20 evaluable patients (75%; 95% CI: 51-91%), with 3 complete and 12 partial responses. Median progression-free and overall survival were 9.2 months (95% CI: 3.8-not reached) and 34.6 months (6.2-not reached), respectively. The most common grade 3/4 toxicities were skin toxicities such as rash (n=4, 19%) and cutaneous squamous cell carcinomas (n=3, 14%), along with diarrhea (n=3, 14%). Pharmacodynamic analysis of patients' PBMCs showed increased day 8 HSP70 expression compared to baseline in the three cohorts with XL888 doses ≥45 mg. Diverse effects of vemurafenib-XL888 upon intratumoral HSP-client protein expression were noted, with the expression of multiple proteins (including ERBB3 and BAD) modulated on therapy. XL888 in combination with vemurafenib has clinical activity in patients with advanced BRAF V600 -mutant melanoma, with a tolerable side-effect profile. HSP90 inhibitors warrant further evaluation in combination with current standard-of-care BRAF plus MEK inhibitors in BRAF V600 -mutant melanoma. Copyright ©2018, American Association for Cancer Research.

  11. Andrographolide downregulates the v-Src and Bcr-Abl oncoproteins and induces Hsp90 cleavage in the ROS-dependent suppression of cancer malignancy.

    Science.gov (United States)

    Liu, Sheng-Hung; Lin, Chao-Hsiung; Liang, Fong-Ping; Chen, Pei-Fen; Kuo, Cheng-Deng; Alam, Mohd Mujahid; Maiti, Barnali; Hung, Shih-Kai; Chi, Chin-Wen; Sun, Chung-Ming; Fu, Shu-Ling

    2014-01-15

    Andrographolide is a diterpenoid compound isolated from Andrographis paniculata that exhibits anticancer activity. We previously reported that andrographolide suppressed v-Src-mediated cellular transformation by promoting the degradation of Src. In the present study, we demonstrated the involvement of Hsp90 in the andrographolide-mediated inhibition of Src oncogenic activity. Using a proteomics approach, a cleavage fragment of Hsp90α was identified in andrographolide-treated cells. The concentration- and time-dependent induction of Hsp90 cleavage that accompanied the reduction in Src was validated in RK3E cells transformed with either v-Src or a human truncated c-Src variant and treated with andrographolide. In cancer cells, the induction of Hsp90 cleavage by andrographolide and its structural derivatives correlated well with decreased Src levels, the suppression of transformation, and the induction of apoptosis. Moreover, the andrographolide-induced Hsp90 cleavage, Src degradation, inhibition of transformation, and induction of apoptosis were abolished by a ROS inhibitor, N-acetyl-cysteine. Notably, Hsp90 cleavage, decreased levels of Bcr-Abl (another known Hsp90 client protein), and the induction of apoptosis were also observed in human K562 leukemia cells treated with andrographolide or its active derivatives. Together, we demonstrated a novel mechanism by which andrographolide suppressed cancer malignancy that involved inhibiting Hsp90 function and reducing the levels of Hsp90 client proteins. Our results broaden the molecular basis of andrographolide-mediated anticancer activity. Copyright © 2013 Elsevier Inc. All rights reserved.

  12. The Hsp90 inhibitor 17-AAG represses calcium-induced cytokeratin 1 and 10 expression in HaCaT keratinocytes.

    Science.gov (United States)

    Miyoshi, Sadanori; Yamazaki, Shota; Uchiumi, Asato; Katagata, Yohtaro

    2012-01-01

    Hsp90 is essential for maintaining the activity of numerous signaling factors, and plays a key role in cellular signal transduction networks. 17-Allylamino-17-demethoxygeldanamycin (17-AAG) is an ansamycin antibiotic that binds to Hsp90 and inhibits its function. HaCaT human keratinocytes were used to investigate the cellular and molecular functions of Hsp90 in keratinocyte differentiation. Inhibition of Hsp90 by 17-AAG leads to downregulation of the differentiation markers cytokeratin 1 and cytokeratin 10 at the protein and mRNA levels.

  13. In vitro study comparing the efficacy of the water-soluble HSP90 inhibitors, 17-AEPGA and 17-DMAG, with that of the non‑water-soluble HSP90 inhibitor, 17-AAG, in breast cancer cell lines.

    Science.gov (United States)

    Ghadban, Tarik; Jessen, André; Reeh, Matthias; Dibbern, Judith L; Mahner, Sven; Mueller, Volkmar; Wellner, Ulrich F; Güngör, Cenap; Izbicki, Jakob R; Vashist, Yogesh K

    2016-10-01

    Heat shock protein (HSP)90 has emerged as an important target in cancer therapeutics. Diverse HSP90 inhibitors are under evaluation. The aim of the present study was to investigate the growth inhibitory effects of the newly developed water-soluble HSP90 inhibitors, 17-[2-(Pyrrolidin-1-yl)ethyl]amino-17-demethoxygeldanamycin (17-AEPGA) and 17-dimethylaminoethylamino-17-demethoxygeldanamycin (17-DMAG), compared to that of the non-water-soluble HSP90 inhibitor, 17-allylamino-17-demethoxygeldanamycin (17-AAG). The anti-proliferative effects of the 3 drugs on the human breast cancer cell lines, MCF-7, SKBR-3 and MDA-MB-231, were examined in vitro. In addition, tumor progression factors, including human epidermal growth factor receptor 2 (HER2), epidermal growth factor receptor 1 (EGFR1) and insulin-like growth factor type 1 receptor (IGF1R), as well as apoptotic markers were analysed. We found a time- and dose-dependent effect in all the tested cell lines. The effects of 17-AEPGA and 17-DMAG were equal or superior to those of 17-AAG. The 50% growth inhibition concentration was AAG.

  14. Orchestrating innovation

    NARCIS (Netherlands)

    Berkers, F.T.H.M.; Klein Woolthuis, R.J.A.; Boer, J. de

    2015-01-01

    Orchestrating Innovation increases the probability of success, minimizing the probability of failure of technological innovations by creating sustained societal and economic value. Orchestrating innovation propagates to take into account and actively involve all relevant stakeholders of the (future)

  15. Repercussion of mitochondria deformity induced by anti-Hsp90 drug 17AAG in human tumor cells

    KAUST Repository

    Vishal, Chaturvedi

    2011-06-07

    Inhibiting Hsp90 chaperone roles using 17AAG induces cytostasis or apoptosis in tumor cells through destabilization of several mutated cancer promoting proteins. Although mitochondria are central in deciding the fate of cells, 17AAG induced effects on tumor cell mitochondria were largely unknown. Here, we show that Hsp90 inhibition with 17AAG first affects mitochondrial integrity in different human tumor cells, neuroblastoma, cervical cancer and glial cells. Using human neuroblastoma tumor cells, we found the early effects associated with a change in mitochondrial membrane potential, elongation and engorgement of mitochondria because of an increased matrix vacuolization. These effects are specific to Hsp90 inhibition as other chemotherapeutic drugs did not induce similar mitochondrial deformity. Further, the effects are independent of oxidative damage and cytoarchitecture destabilization since cytoskeletal disruptors and mitochondrial metabolic inhibitors also do not induce similar deformity induced by 17AAG. The 1D PAGE LC MS/ MS mitochondrial proteome analysis of 17AAG treated human neuroblastoma cells showed a loss of 61% proteins from membrane, metabolic, chaperone and ribonucleoprotein families. About 31 unmapped protein IDs were identified from proteolytic processing map using Swiss-Prot accession number, and converted to the matching gene name searching the ExPASy proteomics server. Our studies display that Hsp90 inhibition effects at first embark on mitochondria of tumor cells and compromise mitochondrial integrity. the author(s), publisher and licensee Libertas Academica Ltd.

  16. The Stoichiometric Interaction of the Hsp90-Sgt1-Rar1 Complex by CD and SRCD Spectroscopy

    Directory of Open Access Journals (Sweden)

    Giuliano Siligardi

    2018-01-01

    Full Text Available While the molecular details by which Hsp90 interacts with Sgt1 and Rar1 were previously described the exact stoichiometric complex that is formed remains elusive. Several possibilities remain that include two asymmetric complexes, Sgt12-Hsp902-Rar12 (two molecules of Sgt1 and Rar1 and one Hsp90 dimer or Sgt12-Hsp902-Rar11 (with a single Rar1 molecule and an asymmetric complex (Sgt11-Hsp902-Rar11. The Hsp90-mediated activation of NLR receptors (Nucleotide-binding domain and Leucine-rich Repeat in the innate immunity of both plants and animals is dependent on the co-chaperone Sgt1 and in plants on Rar1, a cysteine- and histidine-rich domain (CHORD-containing protein. The exact stoichiometry of such a complex may have a direct impact on NLR protein oligomerization and thus ultimately on the mechanism by which NLRs are activated. CD spectroscopy was successfully used to determine the stoichiometry of a ternary protein complex among Hsp90, Sgt1, and Rar1 in the presence of excess ADP. The results indicated that a symmetric Sgt12-Hsp902-Rar11 complex was formed that could allow two NLR molecules to simultaneously bind. The stoichiometry of this complex has implications on, and might promote, the dimerization of NLR proteins following their activation.

  17. RNAi knockdown of Hop (Hsp70/Hsp90 organising protein) decreases invasion via MMP-2 down regulation.

    LENUS (Irish Health Repository)

    Walsh, Naomi

    2011-07-28

    We previously identified Hop as over expressed in invasive pancreatic cancer cell lines and malignant tissues of pancreatic cancer patients, suggesting an important role for Hop in the biology of invasive pancreatic cancer. Hop is a co-chaperone protein that binds to both Hsp70\\/Hsp90. We hypothesised that by targeting Hop, signalling pathways modulating invasion and client protein stabilisation involving Hsp90-dependent complexes may be altered. In this study, we show that Hop knockdown by small interfering (si)RNA reduces the invasion of pancreatic cancer cells, resulting in decreased expression of the downstream target gene, matrix metalloproteinases-2 (MMP-2). Hop in conditioned media co-immunoprecipitates with MMP-2, implicating a possible extracellular function for Hop. Knockdown of Hop expression also reduced expression levels of Hsp90 client proteins, HER2, Bcr-Abl, c-MET and v-Src. Furthermore, Hop is strongly expressed in high grade PanINs compared to lower PanIN grades, displaying differential localisation in invasive ductal pancreatic cancer, indicating that the localisation of Hop is an important factor in pancreatic tumours. Our data suggests that the attenuation of Hop expression inactivates key signal transduction proteins which may decrease the invasiveness of pancreatic cancer cells possibly through the modulation of Hsp90 activity. Therefore, targeting Hop in pancreatic cancer may constitute a viable strategy for targeted cancer therapy.

  18. Assessment and Reconstruction of Novel HSP90 Genes: Duplications, Gains and Losses in Fungal and Animal Lineages

    Czech Academy of Sciences Publication Activity Database

    Pantzartzi, Chrysoula; Drosopoulou, E.; Scouras, Z.

    2013-01-01

    Roč. 8, č. 9 (2013), s. 1-11 E-ISSN 1932-6203 R&D Projects: GA MŠk(CZ) ED1.1.00/02.0109 Institutional support: RVO:68378050 Keywords : Hsp90s * Fungi * duplication events Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.534, year: 2013

  19. Synthesis of 19-substituted geldanamycins with altered conformations and their binding to heat shock protein Hsp90

    Science.gov (United States)

    Kitson, Russell R. A.; Chang, Chuan-Hsin; Xiong, Rui; Williams, Huw E. L.; Davis, Adrienne L.; Lewis, William; Dehn, Donna L.; Siegel, David; Roe, S. Mark; Prodromou, Chrisostomos; Ross, David; Moody, Christopher J.

    2013-01-01

    The benzoquinone ansamycin geldanamycin and its derivatives are inhibitors of heat shock protein Hsp90, an emerging target for novel therapeutic agents both in cancer and in neurodegeneration. However, toxicity of these compounds to normal cells has been ascribed to reaction with thiol nucleophiles at the quinone 19-position. We reasoned that blocking this position would ameliorate toxicity, and that it might also enforce a favourable conformational switch of the trans-amide group into the cis-form required for protein binding. We report here an efficient synthesis of such 19-substituted compounds and realization of our hypotheses. Protein crystallography established that the new compounds bind to Hsp90 with, as expected, a cis-amide conformation. Studies on Hsp90 inhibition in cells demonstrated the molecular signature of Hsp90 inhibitors: decreases in client proteins with compensatory increases in other heat shock proteins in both human breast cancer and dopaminergic neural cells, demonstrating their potential for use in the therapy of cancer or neurodegenerative diseases. PMID:23511419

  20. A member of the HSP90 family from ovine Babesia in China: molecular characterization, phylogenetic analysis and antigenicity.

    Science.gov (United States)

    Guan, Guiquan; Liu, Junlong; Liu, Aihong; Li, Youquan; Niu, Qingli; Gao, Jinliang; Luo, Jianxun; Chauvin, Alain; Yin, Hong; Moreau, Emmanuelle

    2015-09-01

    Heat shock protein 90 (HSP90) is a key component of the molecular chaperone complex essential for activating many signalling proteins involved in the development and progression of pathogenic cellular transformation. A Hsp90 gene (BQHsp90) was cloned and characterized from Babesia sp. BQ1 (Lintan), an ovine Babesia isolate belonging to Babesia motasi-like group, by screening a cDNA expression library and performing rapid amplification of cDNA ends. The full-length cDNA of BQHsp90 is 2399 bp with an open reading frame of 2154 bp encoding a predicted 83 kDa polypeptide with 717 amino acid residues. It shows significant homology and similar structural characteristics to Hsp90 of other apicomplex organisms. Phylogenetic analysis, based on the HSP90 amino acid sequences, showed that the Babesia genus is clearly separated from other apicomplexa genera. Five Chinese ovine Babesia isolates were divided into 2 phylogenetic clusters, namely Babesia sp. Xinjiang (previously designated a new species) cluster and B. motasi-like cluster which could be further divided into 2 subclusters (Babesia sp. BQ1 (Lintan)/Babesia sp. Tianzhu and Babesia sp. BQ1 (Ningxian)/Babesia sp. Hebei). Finally, the antigenicity of rBQHSP90 protein from prokaryotic expression was also evaluated using western blot and enzyme-linked immunosorbent assay (ELISA).

  1. GRP94: An HSP90-like protein specialized for protein folding and quality control in the endoplasmic reticulum

    DEFF Research Database (Denmark)

    Marzec, Michal; Eletto, Davide; Argon, Yair

    2012-01-01

    Glucose-regulated protein 94 is the HSP90-like protein in the lumen of the endoplasmic reticulum and therefore it chaperones secreted and membrane proteins. It has essential functions in development and physiology of multicellular organisms, at least in part because of this unique clientele. GRP94...

  2. Hsp90 inhibition differentially destabilises MAP kinase and TGF-beta signalling components in cancer cells revealed by kinase-targeted chemoproteomics

    International Nuclear Information System (INIS)

    Haupt, Armin; Dahl, Andreas; Lappe, Michael; Lehrach, Hans; Gonzalez, Cayetano; Drewes, Gerard; Lange, Bodo MH; Joberty, Gerard; Bantscheff, Marcus; Fröhlich, Holger; Stehr, Henning; Schweiger, Michal R; Fischer, Axel; Kerick, Martin; Boerno, Stefan T

    2012-01-01

    The heat shock protein 90 (Hsp90) is required for the stability of many signalling kinases. As a target for cancer therapy it allows the simultaneous inhibition of several signalling pathways. However, its inhibition in healthy cells could also lead to severe side effects. This is the first comprehensive analysis of the response to Hsp90 inhibition at the kinome level. We quantitatively profiled the effects of Hsp90 inhibition by geldanamycin on the kinome of one primary (Hs68) and three tumour cell lines (SW480, U2OS, A549) by affinity proteomics based on immobilized broad spectrum kinase inhibitors ('kinobeads'). To identify affected pathways we used the KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway classification. We combined Hsp90 and proteasome inhibition to identify Hsp90 substrates in Hs68 and SW480 cells. The mutational status of kinases from the used cell lines was determined using next-generation sequencing. A mutation of Hsp90 candidate client RIPK2 was mapped onto its structure. We measured relative abundances of > 140 protein kinases from the four cell lines in response to geldanamycin treatment and identified many new potential Hsp90 substrates. These kinases represent diverse families and cellular functions, with a strong representation of pathways involved in tumour progression like the BMP, MAPK and TGF-beta signalling cascades. Co-treatment with the proteasome inhibitor MG132 enabled us to classify 64 kinases as true Hsp90 clients. Finally, mutations in 7 kinases correlate with an altered response to Hsp90 inhibition. Structural modelling of the candidate client RIPK2 suggests an impact of the mutation on a proposed Hsp90 binding domain. We propose a high confidence list of Hsp90 kinase clients, which provides new opportunities for targeted and combinatorial cancer treatment and diagnostic applications

  3. HSP90 inhibitors potentiate PGF2α-induced IL-6 synthesis via p38 MAP kinase in osteoblasts.

    Directory of Open Access Journals (Sweden)

    Kazuhiko Fujita

    Full Text Available Heat shock protein 90 (HSP90 that is ubiquitously expressed in various tissues, is recognized to be a major molecular chaperone. We have previously reported that prostaglandin F2α (PGF2α, a potent bone remodeling mediator, stimulates the synthesis of interleukin-6 (IL-6 through p44/p42 mitogen-activated protein (MAP kinase and p38 MAP kinase in osteoblast-like MC3T3-E1 cells, and that Rho-kinase acts at a point upstream of p38 MAP kinase. In the present study, we investigated the involvement of HSP90 in the PGF2α-stimulated IL-6 synthesis and the underlying mechanism in MC3T3-E1 cells. Geldanamycin, an inhibitor of HSP90, significantly amplified both the PGF2α-stimulated IL-6 release and the mRNA expression levels. In addition, other HSP90 inhibitors, 17-allylamino-17demethoxy-geldanamycin (17-AAG and 17-dimethylamino-ethylamino-17-demethoxy-geldanamycin (17-DMAG and onalespib, enhanced the PGF2α-stimulated IL-6 release. Geldanamycin, 17-AAG and onalespib markedly strengthened the PGF2α-induced phosphorylation of p38 MAP kinase. Geldanamycin and 17-AAG did not affect the PGF2α-induced phosphorylation of p44/p42 MAP kinase and myosin phosphatase targeting subunit (MYPT-1, a substrate of Rho-kinase, and the protein levels of RhoA and Rho-kinase. In addition, HSP90-siRNA enhanced the PGF2α-induced phosphorylation of p38 MAP kinase. Furthermore, SB203580, an inhibitor of p38 MAP kinase, significantly suppressed the amplification by geldanamycin, 17-AAG or 17-DMAG of the PGF2α-stimulated IL-6 release. Our results strongly suggest that HSP90 negatively regulates the PGF2α-stimulated IL-6 synthesis in osteoblasts, and that the effect of HSP90 is exerted through regulating p38 MAP kinase activation.

  4. Exposure to non-ionizing radiation provokes changes in rat thyroid morphology and expression of HSP-90.

    Science.gov (United States)

    Misa-Agustiño, Maria J; Jorge-Mora, Teresa; Jorge-Barreiro, Francisco J; Suarez-Quintanilla, Juan; Moreno-Piquero, Eduardo; Ares-Pena, Francisco J; López-Martín, Elena

    2015-09-01

    Non-ionizing radiation at 2.45 GHz may modify the morphology and expression of genes that codify heat shock proteins (HSP) in the thyroid gland. Diathermy is the therapeutic application of non-ionizing radiation to humans for its beneficial effects in rheumatological and musculo-skeletal pain processes. We used a diathermy model on laboratory rats subjected to maximum exposure in the left front leg, in order to study the effects of radiation on the nearby thyroid tissue. Fifty-six rats were individually exposed once or repeatedly (10 times in two weeks) for 30 min to 2.45 GHz radiation in a commercial chamber at different non-thermal specific absorption rates (SARs), which were calculated using the finite difference time domain technique. We used immunohistochemistry methods to study the expression of HSP-90 and morphological changes in thyroid gland tissues. Ninety minutes after radiation with the highest SAR, the central and peripheral follicles presented increased size and the thickness of the peripheral septa had decreased. Twenty-four hours after radiation, only peripheral follicles radiated at 12 W were found to be smaller. Peripheral follicles increased in size with repeated exposure at 3 W power. Morphological changes in the thyroid tissue may indicate a glandular response to acute or repeated stress from radiation in the hypothalamic-pituitary-thyroid axis. Further research is needed to determine if the effect of this physical agent over time may cause disease in the human thyroid gland. © 2015 by the Society for Experimental Biology and Medicine.

  5. Exposure to non-ionizing radiation provokes changes in rat thyroid morphology and expression of HSP-90

    Science.gov (United States)

    Misa-Agustiño, Maria J; Jorge-Mora, Teresa; Jorge-Barreiro, Francisco J; Suarez-Quintanilla, Juan; Moreno-Piquero, Eduardo; Ares-Pena, Francisco J

    2015-01-01

    Non-ionizing radiation at 2.45 GHz may modify the morphology and expression of genes that codify heat shock proteins (HSP) in the thyroid gland. Diathermy is the therapeutic application of non-ionizing radiation to humans for its beneficial effects in rheumatological and musculo-skeletal pain processes. We used a diathermy model on laboratory rats subjected to maximum exposure in the left front leg, in order to study the effects of radiation on the nearby thyroid tissue. Fifty-six rats were individually exposed once or repeatedly (10 times in two weeks) for 30 min to 2.45 GHz radiation in a commercial chamber at different non-thermal specific absorption rates (SARs), which were calculated using the finite difference time domain technique. We used immunohistochemistry methods to study the expression of HSP-90 and morphological changes in thyroid gland tissues. Ninety minutes after radiation with the highest SAR, the central and peripheral follicles presented increased size and the thickness of the peripheral septa had decreased. Twenty-four hours after radiation, only peripheral follicles radiated at 12 W were found to be smaller. Peripheral follicles increased in size with repeated exposure at 3 W power. Morphological changes in the thyroid tissue may indicate a glandular response to acute or repeated stress from radiation in the hypothalamic–pituitary–thyroid axis. Further research is needed to determine if the effect of this physical agent over time may cause disease in the human thyroid gland. PMID:25649190

  6. Repression of Proteases and Hsp90 Chaperone Expression Induced by an Antiretroviral in Virulent Environmental Strains of Cryptococcus neoformans.

    Science.gov (United States)

    Serafin, Cleber Fernando; Paris, Ana Paula; Paula, Claudete Rodrigues; Simão, Rita Cássia Garcia; Gandra, Rinaldo Ferreira

    2017-04-01

    This study evaluated the effect of the antiretroviral ritonavir on protease secretion in different strains of Cryptococcus neoformans isolated from the environment and investigated the expression of heat shock protein (Hsp90), classically described virulence factors in other yeast in the presence of the same antiretroviral. The presence of the enzyme was detected by the formation of a degradation of the halo around the colonies. The results were classified as follows: level 1 (without proteases), level 2 (positive for proteases), and level 3 (strongly positive for proteases). Total protein extract isolated from the cell walls of the 12 strains incubated in the absence and presence of ritonavir (0.3125 mg mL -1 ) were resolved by SDS-PAGE and analyzed by Western blot assays using an antiserum against Hsp90 from Blastocladiella emersonii. All strains tested showed inhibition of proteinase activity in the presence of ritonavir at 0.3125 to 1.25 mg mL -1 . High levels of Hsp90 were observed in the absence of ritonavir (0.3125 mg mL -1 ), except for the non-virulent control cells. In contrast, in the presence of the antiretroviral, a drastic reduction in the expression of the chaperone was observed. The data suggest that ritonavir, in addition to containing viral replication, could inhibit the expression of virulence factors in opportunistic yeast, as proteases and Hsp90. According to our current knowledge, this is the first time that the inhibition of Hsp90 by an antiretroviral was reported for environmental isolates of C. neoformans.

  7. Hsp90 interacts specifically with viral RNA and differentially regulates replication initiation of Bamboo mosaic virus and associated satellite RNA.

    Directory of Open Access Journals (Sweden)

    Ying Wen Huang

    Full Text Available Host factors play crucial roles in the replication of plus-strand RNA viruses. In this report, a heat shock protein 90 homologue of Nicotiana benthamiana, NbHsp90, was identified in association with partially purified replicase complexes from BaMV-infected tissue, and shown to specifically interact with the 3' untranslated region (3' UTR of BaMV genomic RNA, but not with the 3' UTR of BaMV-associated satellite RNA (satBaMV RNA or that of genomic RNA of other viruses, such as Potato virus X (PVX or Cucumber mosaic virus (CMV. Mutational analyses revealed that the interaction occurs between the middle domain of NbHsp90 and domain E of the BaMV 3' UTR. The knockdown or inhibition of NbHsp90 suppressed BaMV infectivity, but not that of satBaMV RNA, PVX, or CMV in N. benthamiana. Time-course analysis further revealed that the inhibitory effect of 17-AAG is significant only during the immediate early stages of BaMV replication. Moreover, yeast two-hybrid and GST pull-down assays demonstrated the existence of an interaction between NbHsp90 and the BaMV RNA-dependent RNA polymerase. These results reveal a novel role for NbHsp90 in the selective enhancement of BaMV replication, most likely through direct interaction with the 3' UTR of BaMV RNA during the initiation of BaMV RNA replication.

  8. Extracellular Hsp90 serves as a co-factor for MAPK activation and latent viral gene expression during de novo infection by KSHV

    International Nuclear Information System (INIS)

    Qin Zhiqiang; DeFee, Michael; Isaacs, Jennifer S.; Parsons, Chris

    2010-01-01

    The Kaposi's sarcoma-associated herpesvirus (KSHV) is the causative agent of Kaposi's sarcoma (KS), an important cause of morbidity and mortality in immunocompromised patients. KSHV interaction with the cell membrane triggers activation of specific intracellular signal transduction pathways to facilitate virus entry, nuclear trafficking, and ultimately viral oncogene expression. Extracellular heat shock protein 90 localizes to the cell surface (csHsp90) and facilitates signal transduction in cancer cell lines, but whether csHsp90 assists in the coordination of KSHV gene expression through these or other mechanisms is unknown. Using a recently characterized non-permeable inhibitor specifically targeting csHsp90 and Hsp90-specific antibodies, we show that csHsp90 inhibition suppresses KSHV gene expression during de novo infection, and that this effect is mediated largely through the inhibition of mitogen-activated protein kinase (MAPK) activation by KSHV. Moreover, we show that targeting csHsp90 reduces constitutive MAPK expression and the release of infectious viral particles by patient-derived, KSHV-infected primary effusion lymphoma cells. These data suggest that csHsp90 serves as an important co-factor for KSHV-initiated MAPK activation and provide proof-of-concept for the potential benefit of targeting csHsp90 for the treatment or prevention of KSHV-associated illnesses.

  9. [Gene Expression Profile of Apoptosis in Leukemia Cells Induced by Hsp90 Selective inhibitor 17-AAG].

    Science.gov (United States)

    Wang, Na-Na; Li, Zhi-Heng; Tao, Yan-Fang; Xu, Li-Xiao; Pan, Jian; Hu, Shao-Yan

    2016-06-01

    To investigate the apoptotic effects of Hsp90 selective inhibitor 17-AAG on human leukemia HL-60 and NB4 cells and analyse its possible mechanism. CCK-8 assay was used to quantify the growth inhibition of cells after exposure to 17-AAG for 24 hours. Flow cytometrve with annexin V/propidium iodide staining was used to detect apoptosis of leukemia cells. Then Western blot was used to detect the activation of apoptosis related protein caspase-3 and PARP level. Gene expression profile of NB4 cells treated with 17-AAG was analyzed with real-time PCR arrays. The inhibition of leukemia cell proliferation displayed a dose-dependent manner. Annexin V assay, cell cycle analysis and activation of PARP demonstrate that 17-AAG induced apoptosis leukemia cells. Real-time PCR array analysis showed that expression of 56 genes significantly up-regulated and expression of 23 genes were significantly down-regulated after 17-AAG treatment. The 17-AAG can inhibit the proliferation and induce the apoptosis of leukemia cells. After leukemia cells are treated with 17-AAG, the significant changes of apoptosis-related genes occured, and the cell apoptosis occurs via activating apoptosis related signaling pathway.

  10. Hsp10, Hsp70, and Hsp90 immunohistochemical levels change in ulcerative colitis after therapy

    Directory of Open Access Journals (Sweden)

    G. Tomasello

    2011-10-01

    Full Text Available Ulcerative colitis (UC is a form of inflammatory bowel disease (IBD characterized by damage of large bowel mucosa and frequent extra-intestinal autoimmune comorbidities. The role played in IBD pathogenesis by molecular chaperones known to interact with components of the immune system involved in inflammation is unclear. We previously demonstrated that mucosal Hsp60 decreases in UC patients treated with conventional therapies (mesalazine, probiotics, suggesting that this chaperonin could be a reliable biomarker useful for monitoring response to treatment, and that it might play a role in pathogenesis. In the present work we investigated three other heat shock protein/molecular chaperones: Hsp10, Hsp70, and Hsp90. We found that the levels of these proteins are increased in UC patients at the time of diagnosis and decrease after therapy, supporting the notion that these proteins deserve attention in the study of the mechanisms that promote the development and maintenance of IBD, and as biomarkers of this disease (e.g., to monitor response to treatment at the histological level.

  11. Increased radiosensitivity and radiothermosensitivity of human pancreatic MIA PaCa-2 and U251 glioblastoma cell lines treated with the novel Hsp90 inhibitor NVP-HSP990

    International Nuclear Information System (INIS)

    Milanović, Dušan; Firat, Elke; Grosu, Anca Ligia; Niedermann, Gabriele

    2013-01-01

    Heat shock Protein 90 (Hsp90) is a molecular chaperone that folds, stabilizes, and functionally regulates many cellular proteins involved in oncogenic signaling and in the regulation of radiosensitivity. It is upregulated in response to stress such a heat. Hyperthermia is a potent radiosensitizer, but induction of Hsp90 may potentially limit its efficacy. Our aim was to investigate whether the new Hsp90 inhibitor NVP-HSP990 increases radiosensitivity, thermosensitivity and radiothermosensitivity of human tumor cell lines. U251 glioblastoma and MIA PaCa-2 pancreatic carcinoma cells were used. To determine clonogenic survival, colony forming assays were performed. Cell viability and proliferation were assesed by Trypan blue staining. Cell cycle and apoptosis analyses were performed by flow cytometry. DAPI staining was used to detect mitotic catastrophe. NVP-HSP990 increased the thermosensitivity, radiosensitivity and radio-thermosensitivity of both cell lines in clonogenic assays. 72 hours after irradiation with 4 Gy, a significant reduction in cell number associated with considerable G2/M acumulation and mitotic catastrophe as well as cell death by apoptosis/necrosis was observed. Treatment with NVP-HSP990 strongly sensitized U251 and MIA PaCa-2 cells to hyperthermia and ionizing radiation or combination thereof through augmentation of G2/M arrest, mitotic catastrophe and associated apoptosis

  12. HSP90 and pCREB alterations are linked to mancozeb-dependent behavioral and neurodegenerative effects in a marine teleost

    Energy Technology Data Exchange (ETDEWEB)

    Zizza, Merylin [Comparative Neuroanatomy Laboratory, Biology, Ecology and Earth Science Department (DiBEST), University of Calabria, Arcavacata of Rende, 87036, CS (Italy); Di Lorenzo, Mariana; Laforgia, Vincenza [Department of Biology, Section of Evolutionary and Comparative Biology, University of Naples Federico II, 80134 Naples (Italy); Furia, Emilia; Sindona, Giovanni [Department of Chemistry and Chemical Technologies (DCTC), University of Calabria, Arcavacata of Rende, 87036, CS (Italy); Canonaco, Marcello [Comparative Neuroanatomy Laboratory, Biology, Ecology and Earth Science Department (DiBEST), University of Calabria, Arcavacata of Rende, 87036, CS (Italy); Facciolo, Rosa Maria, E-mail: rm.facciolo@unical.it [Comparative Neuroanatomy Laboratory, Biology, Ecology and Earth Science Department (DiBEST), University of Calabria, Arcavacata of Rende, 87036, CS (Italy)

    2017-05-15

    The pesticide mancozeb (mz) is recognized as a potent inducer of oxidative stress due to its ability to catalyze the production of reactive oxygen species plus inhibiting mitochondrial respiration thus becoming an environmental risk for neurodegenerative diseases. Despite numerous toxicological studies on mz have been directed to mammals, attention on marine fish is still lacking. Thus, it was our intention to evaluate neurobehavioral activities of ornate wrasses (Thalassoma pavo) exposed to 0.2 mg/l of mz after a preliminary screening test (0.07–0.3 mg/l). Treated fish exhibited an evident (p < 0.001) latency to reach T-maze arms (> 1000%) while exploratory attitudes (total arm entries) diminished (− 50%; p < 0.05) versus controls during spontaneous exploration tests. Moreover, they showed evident enhancements (+ 111%) of immobility in the cylinder test. Contextually, strong (− 88%; p < 0.01) reductions of permanence in light zone of the Light/Dark apparatus along with diminished crossings (− 65%) were also detected. Conversely, wrasses displayed evident enhancements (160%) of risk assessment consisting of fast entries in the dark side of this apparatus. From a molecular point of view, a notable activation (p < 0.005) of the brain transcription factor pCREB occurred during mz-exposure. Similarly, in situ hybridization supplied increased HSP90 mRNAs in most brain areas such as the lateral part of the dorsal telencephalon (Dl; + 68%) and valvula of the cerebellum (VCe; + 35%) that also revealed evident argyrophilic signals. Overall, these first indications suggest a possible protective role of the early biomarkers pCREB and HSP90 against fish toxicity. - Highlights: • Fish exposed to mancozeb exhibited an evident latency to reach T-maze arms. • Mancozeb caused immobility and reduction of explorative attitudes. • Fish exposed to mancozeb showed anxiogenic performances in the Light/Dark apparatus. • The brain of fish exposed to mancozeb supplied p

  13. HSP90 and pCREB alterations are linked to mancozeb-dependent behavioral and neurodegenerative effects in a marine teleost

    International Nuclear Information System (INIS)

    Zizza, Merylin; Di Lorenzo, Mariana; Laforgia, Vincenza; Furia, Emilia; Sindona, Giovanni; Canonaco, Marcello; Facciolo, Rosa Maria

    2017-01-01

    The pesticide mancozeb (mz) is recognized as a potent inducer of oxidative stress due to its ability to catalyze the production of reactive oxygen species plus inhibiting mitochondrial respiration thus becoming an environmental risk for neurodegenerative diseases. Despite numerous toxicological studies on mz have been directed to mammals, attention on marine fish is still lacking. Thus, it was our intention to evaluate neurobehavioral activities of ornate wrasses (Thalassoma pavo) exposed to 0.2 mg/l of mz after a preliminary screening test (0.07–0.3 mg/l). Treated fish exhibited an evident (p < 0.001) latency to reach T-maze arms (> 1000%) while exploratory attitudes (total arm entries) diminished (− 50%; p < 0.05) versus controls during spontaneous exploration tests. Moreover, they showed evident enhancements (+ 111%) of immobility in the cylinder test. Contextually, strong (− 88%; p < 0.01) reductions of permanence in light zone of the Light/Dark apparatus along with diminished crossings (− 65%) were also detected. Conversely, wrasses displayed evident enhancements (160%) of risk assessment consisting of fast entries in the dark side of this apparatus. From a molecular point of view, a notable activation (p < 0.005) of the brain transcription factor pCREB occurred during mz-exposure. Similarly, in situ hybridization supplied increased HSP90 mRNAs in most brain areas such as the lateral part of the dorsal telencephalon (Dl; + 68%) and valvula of the cerebellum (VCe; + 35%) that also revealed evident argyrophilic signals. Overall, these first indications suggest a possible protective role of the early biomarkers pCREB and HSP90 against fish toxicity. - Highlights: • Fish exposed to mancozeb exhibited an evident latency to reach T-maze arms. • Mancozeb caused immobility and reduction of explorative attitudes. • Fish exposed to mancozeb showed anxiogenic performances in the Light/Dark apparatus. • The brain of fish exposed to mancozeb supplied p

  14. The novel Hsp90 inhibitor NXD30001 induces tumor regression in a genetically engineered mouse model of glioblastoma multiforme.

    Science.gov (United States)

    Zhu, Haihao; Woolfenden, Steve; Bronson, Roderick T; Jaffer, Zahara M; Barluenga, Sofia; Winssinger, Nicolas; Rubenstein, Allan E; Chen, Ruihong; Charest, Al

    2010-09-01

    Glioblastoma multiforme (GBM) has an abysmal prognosis. We now know that the epidermal growth factor receptor (EGFR) signaling pathway and the loss of function of the tumor suppressor genes p16Ink4a/p19ARF and PTEN play a crucial role in GBM pathogenesis: initiating the early stages of tumor development, sustaining tumor growth, promoting infiltration, and mediating resistance to therapy. We have recently shown that this genetic combination is sufficient to promote the development of GBM in adult mice. Therapeutic agents raised against single targets of the EGFR signaling pathway have proven rather inefficient in GBM therapy, showing the need for combinatorial therapeutic approaches. An effective strategy for concurrent disruption of multiple signaling pathways is via the inhibition of the molecular chaperone heat shock protein 90 (Hsp90). Hsp90 inhibition leads to the degradation of so-called client proteins, many of which are key effectors of GBM pathogenesis. NXD30001 is a novel second generation Hsp90 inhibitor that shows improved pharmacokinetic parameters. Here we show that NXD30001 is a potent inhibitor of GBM cell growth in vitro consistent with its capacity to inhibit several key targets and regulators of GBM biology. We also show the efficacy of NXD30001 in vivo in an EGFR-driven genetically engineered mouse model of GBM. Our findings establish that the Hsp90 inhibitor NXD30001 is a therapeutically multivalent molecule, whose actions strike GBM at the core of its drivers of tumorigenesis and represent a compelling rationale for its use in GBM treatment.

  15. Development of Tethered Hsp90 Inhibitors Carrying Radioiodinated Probes to Specifically Discriminate and Kill Malignant Breast Tumor Cells

    Science.gov (United States)

    2017-05-01

    properties and the causes of poor solubility and poor permeability . Journal of pharmacological and toxicological methods 44, 235-249. Liu, S., Li, D., Zhang...250,000 women respectively. Unfortunately, the sensitivity but low specificity of screening has led to concerns about over treatment of indolent...cancer deaths annually. Clinical data indicate a strong link between high expression/activation of Heat shock protein 90 (Hsp90) with poor prognosis

  16. Active Ingredients of Epimedii Folium and Ligustri Lucidi Fructus Balanced GR/HSP90 to Improve the Sensitivity of Asthmatic Rats to Budesonide

    Directory of Open Access Journals (Sweden)

    Xiufeng Tang

    2017-01-01

    Full Text Available This study aimed to investigate the possible molecular mechanisms of active ingredients of Epimedii Folium (EF and Ligustri Lucidi Fructus (LLF combined with Budesonide (Bun in asthmatic rats. Rats were divided into 5 groups, including normal group, asthma model group, Bun group, group of active ingredients of EL and LLF (EL, and group of coadministration of Bun with EL (Bun&EL. The asthmatic model was prepared by ovalbumin sensitizing and challenging. Lymphocyte apoptosis, GR protein and binding, and the protein and mRNA of GRα, GRβ, and HSP90 were tested. The results showed that Bun&EL ① markedly increased lymphocyte apoptosis, GR and HSP90 protein, and GR binding in BALF and ② enhanced the expressions of GRα and HSP90 and the ratio of GRα to GRβ or to HSP90 both in protein and in mRNA levels in lung, ③ while decrease occurred in GRβ mRNA and the mRNA ratio of GRβ to HSP90 compared with asthma or Bun group. Moreover, there was a significant correlation between GRα and GRβ in protein level, or between GRα and HSP90 both in protein and in mRNA levels. EL may effectively enhance the sensitivity of asthmatic rats to Bun via balancing GR/HSP90. And these findings will be beneficial for the treatment of asthma in the future.

  17. HSP90 Inhibitors, Geldanamycin and Radicicol, Enhance Fisetin-Induced Cytotoxicity via Induction of Apoptosis in Human Colonic Cancer Cells

    Directory of Open Access Journals (Sweden)

    Ming-Shun Wu

    2013-01-01

    Full Text Available We revealed the cytotoxic effect of the flavonoid, fisetin (FIS, on human COLO205 colon cancer cells in the presence and absence of the HSP90 inhibitors, geldanamycin (GA and radicicol (RAD. Compared to FIS treatment alone of COLO205 cells, GA and RAD significantly enhanced FIS-induced cytotoxicity, increased expression of cleaved caspase-3 and the PAPR protein, and produced a greater density of DNA ladder formation. GA and RAD also reduced the MMPs with induction of caspase-9 protein cleavage in FIS-treated COLO205 cells. Increased caspase-3 and -9 activities were detected in COLO205 cells treated with FIS+GA or FIS+RAD, and the intensity of DNA ladder formation induced by FIS+GA was reduced by adding the caspase-3 inhibitor, DEVD-FMK. A decrease in Bcl-2 but not Bcl-XL or Bax protein by FIS+GA or FIS+RAD was identified in COLO205 cells by Western blotting. A reduction in p53 protein with increased ubiquitin-tagged proteins was observed in COLO205 cells treated with FIS+GA or FIS+RAD. Furthermore, GA and RAD reduced the stability of the p53 protein in COLO205 cells under FIS stimulation. The evidence supports HSP90 inhibitors possibly sensitizing human colon cancer cells to FIS-induced apoptosis, and treating colon cancer by combining HSP90 inhibitors with FIS deserves further in vivo study.

  18. Development and characterization of a novel C-terminal inhibitor of Hsp90 in androgen dependent and independent prostate cancer cells

    International Nuclear Information System (INIS)

    Eskew, Jeffery D; Rajewski, Roger A; Blagg, Brian SJ; Manjarrez, Jacob R; Matts, Robert L; Holzbeierlein, Jeffrey M; Vielhauer, George A; Sadikot, Takrima; Morales, Pedro; Duren, Alicia; Dunwiddie, Irene; Swink, Megan; Zhang, Xiaoying; Hembruff, Stacey; Donnelly, Alison

    2011-01-01

    The molecular chaperone, heat shock protein 90 (Hsp90) has been shown to be overexpressed in a number of cancers, including prostate cancer, making it an important target for drug discovery. Unfortunately, results with N-terminal inhibitors from initial clinical trials have been disappointing, as toxicity and resistance resulting from induction of the heat shock response (HSR) has led to both scheduling and administration concerns. Therefore, Hsp90 inhibitors that do not induce the heat shock response represent a promising new direction for the treatment of prostate cancer. Herein, the development of a C-terminal Hsp90 inhibitor, KU174, is described, which demonstrates anti-cancer activity in prostate cancer cells in the absence of a HSR and describe a novel approach to characterize Hsp90 inhibition in cancer cells. PC3-MM2 and LNCaP-LN3 cells were used in both direct and indirect in vitro Hsp90 inhibition assays (DARTS, Surface Plasmon Resonance, co-immunoprecipitation, luciferase, Western blot, anti-proliferative, cytotoxicity and size exclusion chromatography) to characterize the effects of KU174 in prostate cancer cells. Pilot in vivo efficacy studies were also conducted with KU174 in PC3-MM2 xenograft studies. KU174 exhibits robust anti-proliferative and cytotoxic activity along with client protein degradation and disruption of Hsp90 native complexes without induction of a HSR. Furthermore, KU174 demonstrates direct binding to the Hsp90 protein and Hsp90 complexes in cancer cells. In addition, in pilot in-vivo proof-of-concept studies KU174 demonstrates efficacy at 75 mg/kg in a PC3-MM2 rat tumor model. Overall, these findings suggest C-terminal Hsp90 inhibitors have potential as therapeutic agents for the treatment of prostate cancer

  19. Development and characterization of a novel C-terminal inhibitor of Hsp90 in androgen dependent and independent prostate cancer cells

    Directory of Open Access Journals (Sweden)

    Eskew Jeffery D

    2011-10-01

    Full Text Available Abstract Background The molecular chaperone, heat shock protein 90 (Hsp90 has been shown to be overexpressed in a number of cancers, including prostate cancer, making it an important target for drug discovery. Unfortunately, results with N-terminal inhibitors from initial clinical trials have been disappointing, as toxicity and resistance resulting from induction of the heat shock response (HSR has led to both scheduling and administration concerns. Therefore, Hsp90 inhibitors that do not induce the heat shock response represent a promising new direction for the treatment of prostate cancer. Herein, the development of a C-terminal Hsp90 inhibitor, KU174, is described, which demonstrates anti-cancer activity in prostate cancer cells in the absence of a HSR and describe a novel approach to characterize Hsp90 inhibition in cancer cells. Methods PC3-MM2 and LNCaP-LN3 cells were used in both direct and indirect in vitro Hsp90 inhibition assays (DARTS, Surface Plasmon Resonance, co-immunoprecipitation, luciferase, Western blot, anti-proliferative, cytotoxicity and size exclusion chromatography to characterize the effects of KU174 in prostate cancer cells. Pilot in vivo efficacy studies were also conducted with KU174 in PC3-MM2 xenograft studies. Results KU174 exhibits robust anti-proliferative and cytotoxic activity along with client protein degradation and disruption of Hsp90 native complexes without induction of a HSR. Furthermore, KU174 demonstrates direct binding to the Hsp90 protein and Hsp90 complexes in cancer cells. In addition, in pilot in-vivo proof-of-concept studies KU174 demonstrates efficacy at 75 mg/kg in a PC3-MM2 rat tumor model. Conclusions Overall, these findings suggest C-terminal Hsp90 inhibitors have potential as therapeutic agents for the treatment of prostate cancer.

  20. Orchestrating innovation

    OpenAIRE

    Berkers, F.T.H.M.; Klein Woolthuis, R.J.A.; Boer, J. de

    2015-01-01

    Orchestrating Innovation increases the probability of success, minimizing the probability of failure of technological innovations by creating sustained societal and economic value. Orchestrating innovation propagates to take into account and actively involve all relevant stakeholders of the (future) ecosystem in which the innovation will, can or has to be adopted.

  1. The Noncompetitive Effect of Gambogic Acid Displaces Fluorescence-Labeled ATP but Requires ATP for Binding to Hsp90/HtpG.

    Science.gov (United States)

    Yue, Qing; Stahl, Frank; Plettenburg, Oliver; Kirschning, Andreas; Warnecke, Athanasia; Zeilinger, Carsten

    2018-05-08

    The heat shock protein 90 (Hsp90) family plays a critical role in maintaining the homeostasis of the intracellular environment for human and prokaryotic cells. Hsp90 orthologues were identified as important target proteins for cancer and plant disease therapies. It was shown that gambogic acid (GBA) has the potential to inhibit human Hsp90. However, it is unknown whether it is also able to act on the bacterial high-temperature protein (HtpG) analogue. In this work, we screened GBA and nine other novel potential Hsp90 inhibitors using a miniaturized high-throughput protein microarray-based assay and found that GBA shows an inhibitory effect on different Hsp90s after dissimilarity analysis of the protein sequence alignment. The dissociation constant of GBA and HtpG Xanthomonas (XcHtpG) computed from microscale thermophoresis is 682.2 ± 408 μM in the presence of ATP, which is indispensable for the binding of GBA to XcHtpG. Our results demonstrate that GBA is a promising Hsp90/HtpG inhibitor. The work further demonstrates that our assay concept has great potential for finding new potent Hsp/HtpG inhibitors.

  2. PRDM14 directly interacts with heat shock proteins HSP90α and glucose-regulated protein 78.

    Science.gov (United States)

    Moriya, Chiharu; Taniguchi, Hiroaki; Nagatoishi, Satoru; Igarashi, Hisayoshi; Tsumoto, Kouhei; Imai, Kohzoh

    2018-02-01

    PRDM14 is overexpressed in various cancers and can regulate cancer phenotype under certain conditions. Inhibiting PRDM14 expression in breast and pancreatic cancers has been reported to reduce cancer stem-like phenotypes, which are associated with aggressive tumor properties. Therefore, PRDM14 is considered a promising target for cancer therapy. To develop a pharmaceutical treatment, the mechanism and interacting partners of PRDM14 need to be clarified. Here, we identified the proteins interacting with PRDM14 in triple-negative breast cancer (TNBC) cells, which do not express the three most common types of receptor (estrogen receptors, progesterone receptors, and HER2). We obtained 13 candidates that were pulled down with PRDM14 in TNBC HCC1937 cells and identified them by mass spectrometry. Two candidates-glucose-regulated protein 78 (GRP78) and heat shock protein 90-α (HSP90α)-were confirmed in immunoprecipitation assay in two TNBC cell lines (HCC1937 and MDA-MB231). Surface plasmon resonance analysis using GST-PRDM14 showed that these two proteins directly interacted with PRDM14 and that the interactions required the C-terminal region of PRDM14, which includes zinc finger motifs. We also confirmed the interactions in living cells by NanoLuc luciferase-based bioluminescence resonance energy transfer (NanoBRET) assay. Moreover, HSP90 inhibitors (17DMAG and HSP990) significantly decreased breast cancer stem-like CD24 -  CD44 + and side population (SP) cells in HCC1937 cells, but not in PRDM14 knockdown HCC1937 cells. The combination of the GRP78 inhibitor HA15 and PRDM14 knockdown significantly decreased cell proliferation and SP cell number in HCC1937 cells. These results suggest that HSP90α and GRP78 interact with PRDM14 and participate in cancer regulation. © 2017 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

  3. Expression of focal adhesion kinase in uveal melanoma and the effects of Hsp90 inhibition by 17-AAG.

    Science.gov (United States)

    Faingold, Dana; Filho, Vasco Bravo; Fernandes, Bruno; Jagan, Lisa; de Barros, Alexandre M; Orellana, Maria Eugenia; Antecka, Emilia; Burnier, Miguel N

    2014-11-01

    Focal adhesion kinase (FAK) is implicated in tumor progression and metastatic cascade, and has been shown to be overexpressed in a variety of human cancers. However, the role of FAK in human uveal melanoma (UM) is not well defined. The purpose of this study was to evaluate the expression of FAK in UM tumors and normal eyes, and to determine the effect of Hsp90 inhibition on FAK expression in UM cells. FAK expression was assessed in 39 UM specimens, FAK[pY397] expression was assessed in 51 UM specimens, and both FAK and FAK[pY397] expression were assessed in 20 normal eyes. The expression of FAK and FAK[pY397] was detected by Western blot in five UM cell lines after treatment with 10 μmol/L of 17-AAG. FAK was positive in 87.2% and FAK[pY397] in 90% of UM specimens. Low FAK expression was detected in non-tumor structures and in normal eyes. The cell lines with the most proliferative, invasive phenotype (92.1, SP6.5 and MKT-BR) displayed high expression of FAK[pY397], and the levels of FAK and FAK[pY397] were decreased in the presence of 17-AAG starting with 24 h of exposure. FAK and FAK[pY397] were overexpressed in human UM tumors compared to normal ocular tissue and high levels of FAK[pY397] were seen in the most aggressive UM cell lines. Hsp90 inhibition led to downregulation of FAK expression. We propose a role for FAK in the pathogenesis of UM. Future studies are needed to explore the use of Hsp90 inhibitors as a feasible approach for modulating FAK in UM. Copyright © 2014 Elsevier GmbH. All rights reserved.

  4. Efficacy of the HSP90 inhibitor 17-AAG in human glioma cell lines and tumorigenic glioma stem cells.

    Science.gov (United States)

    Sauvageot, Claire Marie-Elisabeth; Weatherbee, Jessica Leigh; Kesari, Santosh; Winters, Susan Elizabeth; Barnes, Jessica; Dellagatta, Jamie; Ramakrishna, Naren Raj; Stiles, Charles Dean; Kung, Andrew Li-Jen; Kieran, Mark W; Wen, Patrick Yung Chih

    2009-04-01

    Glioblastoma multiforme (GBM) arises from genetic and signaling abnormalities in components of signal transduction pathways involved in proliferation, survival, and the cell cycle axis. Studies to date with single-agent targeted molecular therapy have revealed only modest effects in attenuating the growth of these tumors, suggesting that targeting multiple aberrant pathways may be more beneficial. Heat-shock protein 90 (HSP90) is a molecular chaperone that is involved in the conformational maturation of a defined group of client proteins, many of which are deregulated in GBM. 17-allylamino-17-demethoxygeldanamycin (17-AAG) is a well-characterized HSP90 inhibitor that should be able to target many of the aberrant signal transduction pathways in GBM. We assessed the ability of 17-AAG to inhibit the growth of glioma cell lines and glioma stem cells both in vitro and in vivo and assessed its ability to synergize with radiation and/or temozolomide, the standard therapies for GBM. Our results reveal that 17-AAG is able to inhibit the growth of both human glioma cell lines and glioma stem cells in vitro and is able to target the appropriate proteins within these cells. In addition, 17-AAG can inhibit the growth of intracranial tumors and can synergize with radiation both in tissue culture and in intracranial tumors. This compound was not found to synergize with temozolomide in any of our models of gliomas. Our results suggest that HSP90 inhibitors like 17-AAG may have therapeutic potential in GBM, either as a single agent or in combination with radiation.

  5. Phase I evaluation of STA-1474, a prodrug of the novel HSP90 inhibitor ganetespib, in dogs with spontaneous cancer.

    Directory of Open Access Journals (Sweden)

    Cheryl A London

    Full Text Available The novel water soluble compound STA-1474 is metabolized to ganetespib (formerly STA-9090, a potent HSP90 inhibitor previously shown to kill canine tumor cell lines in vitro and inhibit tumor growth in the setting of murine xenografts. The purpose of the following study was to extend these observations and investigate the safety and efficacy of STA-1474 in dogs with spontaneous tumors.This was a Phase 1 trial in which dogs with spontaneous tumors received STA-1474 under one of three different dosing schemes. Pharmacokinetics, toxicities, biomarker changes, and tumor responses were assessed. Twenty-five dogs with a variety of cancers were enrolled. Toxicities were primarily gastrointestinal in nature consisting of diarrhea, vomiting, inappetence and lethargy. Upregulation of HSP70 protein expression was noted in both tumor specimens and PBMCs within 7 hours following drug administration. Measurable objective responses were observed in dogs with malignant mast cell disease (n = 3, osteosarcoma (n = 1, melanoma (n = 1 and thyroid carcinoma (n = 1, for a response rate of 24% (6/25. Stable disease (>10 weeks was seen in 3 dogs, for a resultant overall biological activity of 36% (9/25.This study provides evidence that STA-1474 exhibits biologic activity in a relevant large animal model of cancer. Given the similarities of canine and human cancers with respect to tumor biology and HSP90 activation, it is likely that STA-1474 and ganetespib will demonstrate comparable anti-cancer activity in human patients.

  6. [HSP90 Inhibitor 17-AAG Inhibits Multiple Myeloma Cell Proliferation by Down-regulating Wnt/β-Catenin Signaling Pathway].

    Science.gov (United States)

    Chen, Kan-Kan; He, Zheng-Mei; Ding, Bang-He; Chen, Yue; Zhang, Li-Juan; Yu, Liang; Gao, Jian

    2016-02-01

    To investigate the inhibitory effect of HSP90 inhibitory 17-AAG on proliferation of multiple myeloma cells and its main mechanism. The multiple myeloma cells U266 were treated with 17-AAG of different concentrations (200, 400, 600 and 800 nmol/L) for 24, 48, and 72 hours respectively, then the proliferation rate, expression levels of β-catenin and C-MYC protein, as well as cell cycle of U266 cells were treated with 17-AAG and were detected by MTT method, Western blot and flow cytometry, respectively. The 17-AAG showed inhibitory effect on the proliferation of U266 cells in dose- and time-depetent manners (r = -0.518, P AAG displayed no inhibitory effect on proliferation of U266 cells (P > 0.05). The result of culturing U266 cells for 72 hours by 17-AAG of different concentrations showed that the more high of 17-AAG concentration, the more low level of β-catenin and C-MYC proteins (P AAG concentration, the more high of cell ratio in G1 phase (P AAG, the more long time of culture, the more high of cell ratio in G1 phase (P AAG can inhibit the proliferation of multiple myeloma cells, the down-regulation of Wnt/β-catenin signaling pathway and inhibition of HSP90 expression may be the main mechnisms of 17-AAG effect.

  7. Inhibition of homologous recombination repair in irradiated tumor cells pretreated with Hsp90 inhibitor 17-allylamino-17-demethoxygeldanamycin

    International Nuclear Information System (INIS)

    Noguchi, Miho; Yu, Dong; Hirayama, Ryoichi; Ninomiya, Yasuharu; Sekine, Emiko; Kubota, Nobuo; Ando, Koichi; Okayasu, Ryuichi

    2006-01-01

    In order to investigate the mechanism of radio-sensitization by an Hsp90 inhibitor 17-allylamino-17-demethoxygeldanamycin (17-AAG), we studied repair of DNA double strand breaks (DSBs) in irradiated human cells pre-treated with 17-AAG. DSBs are thought to be the critical target for radiation-induced cell death. Two human tumor cell lines DU145 and SQ-5 which showed clear radio-sensitization by 17-AAG revealed a significant inhibition of DSB repair, while normal human cells which did not show radio-sensitization by the drug indicated no change in the DSB repair kinetics with 17-AAG. We further demonstrated that BRCA2 was a novel client protein for Hsp90, and 17-AAG caused the degradation of BRCA2 and in turn altered the behavior of Rad51, a critical protein for homologous recombination (HR) pathway of DSB repair. Our data demonstrate for the first time that 17-AAG inhibits the HR repair process and could provide a new therapeutic strategy to selectively result in higher tumor cell killing

  8. Cellular growth kinetics distinguish a cyclophilin inhibitor from an HSP90 inhibitor as a selective inhibitor of hepatitis C virus.

    Directory of Open Access Journals (Sweden)

    Rudolf K F Beran

    Full Text Available During antiviral drug discovery, it is critical to distinguish molecules that selectively interrupt viral replication from those that reduce virus replication by adversely affecting host cell viability. In this report we investigate the selectivity of inhibitors of the host chaperone proteins cyclophilin A (CypA and heat-shock protein 90 (HSP90 which have each been reported to inhibit replication of hepatitis C virus (HCV. By comparing the toxicity of the HSP90 inhibitor, 17-(Allylamino-17-demethoxygeldanamycin (17-AAG to two known cytostatic compounds, colchicine and gemcitabine, we provide evidence that 17-AAG exerts its antiviral effects indirectly through slowing cell growth. In contrast, a cyclophilin inhibitor, cyclosporin A (CsA, exhibited selective antiviral activity without slowing cell proliferation. Furthermore, we observed that 17-AAG had little antiviral effect in a non-dividing cell-culture model of HCV replication, while CsA reduced HCV titer by more than two orders of magnitude in the same model. The assays we describe here are useful for discriminating selective antivirals from compounds that indirectly affect virus replication by reducing host cell viability or slowing cell growth.

  9. The role of secreted heat shock protein-90 (Hsp90) in wound healing - how could it shape future therapeutics?

    Science.gov (United States)

    Guo, Jiacong; Chang, Cheng; Li, Wei

    2017-08-01

    Defects in tissue repair or wound healing pose a clinical, economic and social problem worldwide. Despite decades of studies, there have been few effective therapeutic treatments. Areas covered: We discuss the possible reasons for why growth factor therapy did not succeed. We point out the lack of human disorder-relevant animal models as another blockade for therapeutic development. We summarize the recent discovery of secreted heat shock protein-90 (Hsp90) as a novel wound healing agent. Expert commentary: Wound healing is a highly complex and multistep process that requires participations of many cell types, extracellular matrices and soluble molecules to work together in a spatial and temporal fashion within the wound microenvironment. The time that wounds remain open directly correlates with the clinical mortality associated with wounds. This time urgency makes the healing process impossible to regenerate back to the unwounded stage, rather forces it to take many shortcuts in order to protect life. Therefore, for therapeutic purpose, it is crucial to identify so-called 'driver genes' for the life-saving phase of wound closure. Keratinocyte-secreted Hsp90α was discovered in 2007 and has shown the promise by overcoming several key hurdles that have blocked the effectiveness of growth factors during wound healing.

  10. MicroPET/CT Imaging of AXL Downregulation by HSP90 Inhibition in Triple-Negative Breast Cancer

    Directory of Open Access Journals (Sweden)

    Wanqin Wang

    2017-01-01

    Full Text Available AXL receptor tyrosine kinase is overexpressed in a number of solid tumor types including triple-negative breast cancer (TNBC. AXL is considered an important regulator of epithelial-to-mesenchymal transition (EMT and a potential therapeutic target for TNBC. In this work, we used microPET/CT with 64Cu-labeled anti-human AXL antibody (64Cu-anti-hAXL to noninvasively interrogate the degradation of AXL in vivo in response to 17-allylamino-17-demethoxygeldanamycin (17-AAG, a potent inhibitor of HSP90. 17-AAG treatment caused significant decline in AXL expression in orthotopic TNBC MDA-MB-231 tumors, inhibited EMT, and delayed tumor growth in vivo, resulting in significant reduction in tumor uptake of 64Cu-anti-hAXL as clearly visualized by microPET/CT. Our data indicate that 64Cu-anti-hAXL can be useful for monitoring anti-AXL therapies and for assessing inhibition of HSP90 molecular chaperone using AXL as a molecular surrogate.

  11. Drug-Induced HSP90 Inhibition Alleviates Pain in Monoarthritic Rats and Alters the Expression of New Putative Pain Players at the DRG.

    Science.gov (United States)

    Nascimento, Diana Sofia Marques; Potes, Catarina Soares; Soares, Miguel Luz; Ferreira, António Carlos; Malcangio, Marzia; Castro-Lopes, José Manuel; Neto, Fani Lourença Moreira

    2018-05-01

    Purinergic receptors (P2XRs) have been widely associated with pain states mostly due to their involvement in neuron-glia communication. Interestingly, we have previously shown that satellite glial cells (SGC), surrounding dorsal root ganglia (DRG) neurons, become activated and proliferate during monoarthritis (MA) in the rat. Here, we demonstrate that P2X7R expression increases in ipsilateral DRG after 1 week of disease, while P2X3R immunoreactivity decreases. We have also reported a significant induction of the activating transcriptional factor 3 (ATF3) in MA. In this study, we show that ATF3 knocked down in DRG cell cultures does not affect the expression of P2X7R, P2X3R, or glial fibrillary acidic protein (GFAP). We suggest that P2X7R negatively regulates P2X3R, which, however, is unlikely mediated by ATF3. Interestingly, we found that ATF3 knockdown in vitro induced significant decreases in the heat shock protein 90 (HSP90) expression. Thus, we evaluated in vivo the involvement of HSP90 in MA and demonstrated that the HSP90 messenger RNA levels increase in ipsilateral DRG of inflamed animals. We also show that HSP90 is mostly found in a cleaved form in this condition. Moreover, administration of a HSP90 inhibitor, 17-dimethylaminoethylamino-17-demethoxygeldanamycin (17-DMAG), attenuated MA-induced mechanical allodynia in the first hours. The drug also reversed the HSP90 upregulation and cleavage. 17-DMAG seemed to attenuate glial activation and neuronal sensitization (as inferred by downregulation of GFAP and P2X3R in ipsilateral DRG) which might correlate with the observed pain alleviation. Our data indicate a role of HSP90 in MA pathophysiology, but further investigation is necessary to clarify the underlying mechanisms.

  12. Chaperone-Mediated Regulation of Choline Acetyltransferase Protein Stability and Activity by HSC/HSP70, HSP90, and p97/VCP

    Directory of Open Access Journals (Sweden)

    Trevor M. Morey

    2017-12-01

    Full Text Available Choline acetyltransferase (ChAT synthesizes the neurotransmitter acetylcholine in cholinergic neurons, and mutations of this enzyme are linked to the neuromuscular disorder congenital myasthenic syndrome (CMS. One CMS-related mutation, V18M, reduces ChAT enzyme activity and cellular protein levels, and is located within a highly-conserved N-terminal proline-rich motif at residues 14PKLPVPP20. We showed previously that disruption of this proline-rich motif by either proline-to-alanine mutation (P17A/P19A or mutation of residue Val18 (V18M enhances ubiquitination and degradation of these mutant ChAT proteins expressed in cholinergic SN56 cells by an unknown mechanism. In this study, using proximity-dependent biotin identification (BioID, co-immunoprecipitation and in situ proximity-ligation assay (PLA, we identified the heat shock proteins (HSPs HSC/HSP70 and HSP90 as novel ChAT protein-interactors. These molecular chaperones are well-known for promoting the folding and stabilization of cellular proteins. Thus, we found that inhibition of HSPs by treatment of cells with either the HSC/HSP70 inhibitors 2-phenylethynesulfonamide (PES or VER-155008, or the HSP90 inhibitor 17-AAG reduced cellular ChAT activity and solubility, and enhanced the ubiquitination and proteasome-dependent loss of ChAT protein. Importantly, the effects of HSP inhibition were greater for mutant ChAT proteins (P17A/P19A-ChAT and CMS-related V18M- and A513T-ChAT compared to wild-type ChAT. HSPs can promote ubiquitination and degradation of terminally misfolded proteins through cooperative interaction with the E3 ubiquitin ligase CHIP/Stub1, and while we show that ChAT interacts with CHIP in situ, siRNA-mediated knock-down of CHIP had no effect on either wild-type or mutant ChAT protein levels. However, inhibition of the endoplasmic reticulum (ER- and HSP-associated co-chaperone p97/VCP prevented degradation of ubiquitinated ChAT. Together, these results identify novel mechanisms

  13. [The Role of Membrane-Bound Heat Shock Proteins Hsp90 in Migration of Tumor Cells in vitro and Involvement of Cell Surface Heparan Sulfate Proteoglycans in Protein Binding to Plasma Membrane].

    Science.gov (United States)

    Snigireva, A V; Vrublevskaya, V V; Skarga, Y Y; Morenkov, O S

    2016-01-01

    Heat shock protein Hsp90, detected in the extracellular space and on the membrane of cells, plays an important role in cell motility, migration, invasion and metastasis of tumor cells. At present, the functional role and molecular mechanisms of Hsp90 binding to plasma membrane are not elucidated. Using isoform-specific antibodies against Hsp90, Hsp9α and Hsp90β, we showed that membrane-bound Hsp90α and Hsp90β play a significant role in migration of human fibrosarcoma (HT1080) and glioblastoma (A-172) cells in vitro. Disorders of sulfonation of cell heparan sulfates, cleavage of cell heparan. sulfates by heparinase I/III as well as treatment of cells with heparin lead to an abrupt reduction in the expression level of Hsp90 isoforms. Furthermore, heparin significantly inhibits tumor cell migration. The results obtained demonstrate that two isoforms of membrane-bound Hsp90 are involved in migration of tumor cells in vitro and that cell surface heparan sulfate proteoglycans play a pivotal role in the "anchoring" of Hsp90α and Hsp90β to the plasma membrane.

  14. Orchestrating Docker

    CERN Document Server

    Holla, Shrikrishna

    2015-01-01

    If you are a competent developer or DevOps with a good understanding of Linux filesystems but want to manage and orchestrate Docker services, images, and products using a multitude of techniques, then this book is for you. No prior knowledge of Docker or container virtualization is required.

  15. Expression of hsp70, hsp90 and hsf1 in the reef coral Acropora digitifera under prospective acidified conditions over the next several decades

    Directory of Open Access Journals (Sweden)

    Masako Nakamura

    2012-02-01

    Ocean acidification is an ongoing threat for marine organisms due to the increasing atmospheric CO2 concentration. Seawater acidification has a serious impact on physiologic processes in marine organisms at all life stages. On the other hand, potential tolerance to external pH changes has been reported in coral larvae. Information about the possible mechanisms underlying such tolerance responses, however, is scarce. In the present study, we examined the effects of acidified seawater on the larvae of Acropora digitifera at the molecular level. We targeted two heat shock proteins, Hsp70 and Hsp90, and a heat shock transcription factor, Hsf1, because of their importance in stress responses and in early life developmental stages. Coral larvae were maintained under the ambient and elevated CO2 conditions that are expected to occur within next 100 years, and then we evaluated the expression of hsps and hsf1 by quantitative real-time polymerase chain reaction (PCR. Expression levels of these molecules significantly differed among target genes, but they did not change significantly between CO2 conditions. These findings indicate that the expression of hsps is not changed due to external pH changes, and suggest that tolerance to acidified seawater in coral larvae may not be related to hsp expression.

  16. BAG3-dependent expression of Mcl-1 confers resistance of mutant KRAS colon cancer cells to the HSP90 inhibitor AUY922.

    Science.gov (United States)

    Wang, Chun Yan; Guo, Su Tang; Croft, Amanda; Yan, Xu Guang; Jin, Lei; Zhang, Xu Dong; Jiang, Chen Chen

    2018-02-01

    Past studies have shown that mutant KRAS colon cancer cells are susceptible to apoptosis induced by the HSP90 inhibitor AUY922. Nevertheless, intrinsic and acquired resistance remains an obstacle for the potential application of the inhibitor in the treatment of the disease. Here we report that Mcl-1 is important for survival of colon cancer cells in the presence of AUY922. Mcl-1 was upregulated in mutant KRAS colon cancer cells selected for resistance to AUY922-induced apoptosis. This was due to its increased stability mediated by Bcl-2-associated athanogene domain 3 (BAG3), which was also increased in resistant colon cancer cells by heat shock factor 1 (HSF1) as a result of chronic endoplasmic reticulum (ER) stress. Functional investigations demonstrated that inhibition of Mcl-1, BAG3, or HSF1 triggered apoptosis in resistant colon cancer cells, and rendered AUY922-naïve colon cancer cells more sensitive to the inhibitor. Together, these results identify that the HSF1-BAG3-Mcl-1 signal axis is critical for protection of mutant KRAS colon cancer cells from AUY922-induced apoptosis, with potential implications for targeting HSF1/BAG3/Mcl-1 to improve the efficacy of AUY922 in the treatment of colon cancer. © 2017 Wiley Periodicals, Inc.

  17. Integration of gene dosage and gene expression in non-small cell lung cancer, identification of HSP90 as potential target.

    Directory of Open Access Journals (Sweden)

    Mariëlle I Gallegos Ruiz

    Full Text Available BACKGROUND: Lung cancer causes approximately 1.2 million deaths per year worldwide, and non-small cell lung cancer (NSCLC represents 85% of all lung cancers. Understanding the molecular events in non-small cell lung cancer (NSCLC is essential to improve early diagnosis and treatment for this disease. METHODOLOGY AND PRINCIPAL FINDINGS: In an attempt to identify novel NSCLC related genes, we performed a genome-wide screening of chromosomal copy number changes affecting gene expression using microarray based comparative genomic hybridization and gene expression arrays on 32 radically resected tumor samples from stage I and II NSCLC patients. An integrative analysis tool was applied to determine whether chromosomal copy number affects gene expression. We identified a deletion on 14q32.2-33 as a common alteration in NSCLC (44%, which significantly influenced gene expression for HSP90, residing on 14q32. This deletion was correlated with better overall survival (P = 0.008, survival was also longer in patients whose tumors had low expression levels of HSP90. We extended the analysis to three independent validation sets of NSCLC patients, and confirmed low HSP90 expression to be related with longer overall survival (P = 0.003, P = 0.07 and P = 0.04. Furthermore, in vitro treatment with an HSP90 inhibitor had potent antiproliferative activity in NSCLC cell lines. CONCLUSIONS: We suggest that targeting HSP90 will have clinical impact for NSCLC patients.

  18. [HSP90 inhibitor 17-AAG plays an important role in JAK3/STAT5 signaling pathways in HTLV-1 infection cell line HUT-102].

    Science.gov (United States)

    Yang, Q Q; Tan, H; Fu, Z P; Ma, Q; Song, J L

    2017-08-14

    Objective: To analyze whether heat-shock protein 90 (HSP90) be involved in a permanently abnormal activated JAK/STAT signaling in ATL cells in vitro. Methods: The effect of 17-AAG on proliferation of ATL cell lines HUT-102 was assessed using CCK8 at different time points. Cell apoptosis was measured by flow cytometry. The specific proteins HSP90, STAT5, p-STAT5 and JAK3 were detected by Western blotting. Results: Overexpression of HSP90 in HUT-102 cell lines was disclosed ( P AAG led to reduced cell proliferation, but there was no significant change in terms of cell proliferation when the concentration of 17-AAG between 2 000-8 000 nmol/L ( P >0.05) . 17-AAG induced cell apoptosis in different time-points and concentrations. 17-AAG don't affect the expression of JAK3 gene. Conclusion: This study indicated that JAK3 as HSP90 client protein was aberrantly activated in HTLV-1-infected T-cell lines, leading to constitutive activation of p-STAT5 in JAK/STAT signal pathway, which demonstrated that HSP90-inhibitors 17-AAG inhibited the growth of HTLV-1-infected T-cell lines by reducing cell proliferation and inducing cell apoptosis.

  19. Structural Bioinformatics and Protein Docking Analysis of the Molecular Chaperone-Kinase Interactions: Towards Allosteric Inhibition of Protein Kinases by Targeting the Hsp90-Cdc37 Chaperone Machinery

    Directory of Open Access Journals (Sweden)

    Gennady Verkhivker

    2013-11-01

    Full Text Available A fundamental role of the Hsp90-Cdc37 chaperone system in mediating maturation of protein kinase clients and supporting kinase functional activity is essential for the integrity and viability of signaling pathways involved in cell cycle control and organism development. Despite significant advances in understanding structure and function of molecular chaperones, the molecular mechanisms and guiding principles of kinase recruitment to the chaperone system are lacking quantitative characterization. Structural and thermodynamic characterization of Hsp90-Cdc37 binding with protein kinase clients by modern experimental techniques is highly challenging, owing to a transient nature of chaperone-mediated interactions. In this work, we used experimentally-guided protein docking to probe the allosteric nature of the Hsp90-Cdc37 binding with the cyclin-dependent kinase 4 (Cdk4 kinase clients. The results of docking simulations suggest that the kinase recognition and recruitment to the chaperone system may be primarily determined by Cdc37 targeting of the N-terminal kinase lobe. The interactions of Hsp90 with the C-terminal kinase lobe may provide additional “molecular brakes” that can lock (or unlock kinase from the system during client loading (release stages. The results of this study support a central role of the Cdc37 chaperone in recognition and recruitment of the kinase clients. Structural analysis may have useful implications in developing strategies for allosteric inhibition of protein kinases by targeting the Hsp90-Cdc37 chaperone machinery.

  20. 2.4 Å resolution crystal structure of human TRAP1 NM , the Hsp90 paralog in the mitochondrial matrix

    Energy Technology Data Exchange (ETDEWEB)

    Sung, Nuri; Lee, Jungsoon; Kim, Ji-Hyun; Chang, Changsoo; Tsai, Francis T. F.; Lee, Sukyeong

    2016-07-13

    TRAP1 is an organelle-specific Hsp90 paralog that is essential for neoplastic growth. As a member of the Hsp90 family, TRAP1 is presumed to be a general chaperone facilitating the late-stage folding of Hsp90 client proteins in the mitochondrial matrix. Interestingly, TRAP1 cannot replace cytosolic Hsp90 in protein folding, and none of the known Hsp90 co-chaperones are found in mitochondria. Thus, the three-dimensional structure of TRAP1 must feature regulatory elements that are essential to the ATPase activity and chaperone function of TRAP1. Here, the crystal structure of a human TRAP1NMdimer is presented, featuring an intact N-domain and M-domain structure, bound to adenosine 5'-β,γ-imidotriphosphate (ADPNP). The crystal structure together with epitope-mapping results shows that the TRAP1 M-domain loop 1 contacts the neighboring subunit and forms a previously unobserved third dimer interface that mediates the specific interaction with mitochondrial Hsp70.

  1. 2.4 Å resolution crystal structure of human TRAP1NM, the Hsp90 paralog in the mitochondrial matrix.

    Science.gov (United States)

    Sung, Nuri; Lee, Jungsoon; Kim, Ji Hyun; Chang, Changsoo; Tsai, Francis T F; Lee, Sukyeong

    2016-08-01

    TRAP1 is an organelle-specific Hsp90 paralog that is essential for neoplastic growth. As a member of the Hsp90 family, TRAP1 is presumed to be a general chaperone facilitating the late-stage folding of Hsp90 client proteins in the mitochondrial matrix. Interestingly, TRAP1 cannot replace cytosolic Hsp90 in protein folding, and none of the known Hsp90 co-chaperones are found in mitochondria. Thus, the three-dimensional structure of TRAP1 must feature regulatory elements that are essential to the ATPase activity and chaperone function of TRAP1. Here, the crystal structure of a human TRAP1NM dimer is presented, featuring an intact N-domain and M-domain structure, bound to adenosine 5'-β,γ-imidotriphosphate (ADPNP). The crystal structure together with epitope-mapping results shows that the TRAP1 M-domain loop 1 contacts the neighboring subunit and forms a previously unobserved third dimer interface that mediates the specific interaction with mitochondrial Hsp70.

  2. 3D-QSAR, molecular docking, and molecular dynamic simulations for prediction of new Hsp90 inhibitors based on isoxazole scaffold.

    Science.gov (United States)

    Abbasi, Maryam; Sadeghi-Aliabadi, Hojjat; Amanlou, Massoud

    2018-05-01

    Heat shock protein 90(Hsp90), as a molecular chaperone, play a crucial role in folding and proper function of many proteins. Hsp90 inhibitors containing isoxazole scaffold are currently being used in the treatment of cancer as tumor suppressers. Here in the present studies, new compounds based on isoxazole scaffold were predicted using a combination of molecular modeling techniques including three-dimensional quantitative structure-activity relationship (3D-QSAR), molecular docking and molecular dynamic (MD) simulations. Comparative molecular field analysis (CoMFA) and comparative molecular similarity indices analysis (CoMSIA) were also done. The steric and electrostatic contour map of CoMFA and CoMSIA were created. Hydrophobic, hydrogen bond donor and acceptor of CoMSIA model also were generated, and new compounds were predicted by CoMFA and CoMSIA contour maps. To investigate the binding modes of the predicted compounds in the active site of Hsp90, a molecular docking simulation was carried out. MD simulations were also conducted to evaluate the obtained results on the best predicted compound and the best reported Hsp90 inhibitors in the 3D-QSAR model. Findings indicate that the predicted ligands were stable in the active site of Hsp90.

  3. Hsp90 inhibitor 17-AAG sensitizes Bcl-2 inhibitor (-)-gossypol by suppressing ERK-mediated protective autophagy and Mcl-1 accumulation in hepatocellular carcinoma cells.

    Science.gov (United States)

    Wang, Bin; Chen, Linfeng; Ni, Zhenhong; Dai, Xufang; Qin, Liyan; Wu, Yaran; Li, Xinzhe; Xu, Liang; Lian, Jiqin; He, Fengtian

    2014-11-01

    Natural BH3-memitic (-)-gossypol shows promising antitumor efficacy in several kinds of cancer. However, our previous studies have demonstrated that protective autophagy decreases the drug sensitivities of Bcl-2 inhibitors in hepatocellular carcinoma (HCC) cells. In the present study, we are the first to report that Hsp90 inhibitor 17-AAG enhanced (-)-gossypol-induced apoptosis via suppressing (-)-gossypol-triggered protective autophagy and Mcl-1 accumulation. The suppression effect of 17-AAG on autophagy was mediated by inhibiting ERK-mediated Bcl-2 phosphorylation while was not related to Beclin1 or LC3 protein instability. Meanwhile, 17-AAG downregulated (-)-gossypol-triggered Mcl-1 accumulation by suppressing Mcl-1(Thr163) phosphorylation and promoting protein degradation. Collectively, our study indicates that Hsp90 plays an important role in tumor maintenance and inhibition of Hsp90 may become a new strategy for sensitizing Bcl-2-targeted chemotherapies in HCC cells. Copyright © 2014 Elsevier Inc. All rights reserved.

  4. Acquired resistance to HSP90 inhibitor 17-AAG and increased metastatic potential are associated with MUC1 expression in colon carcinoma cells.

    Science.gov (United States)

    Liu, Xin; Ban, Li-Li; Luo, Gang; Li, Zhi-Yao; Li, Yun-Feng; Zhou, Yong-Chun; Wang, Xi-Cai; Jin, Cong-Guo; Ye, Jia-Gui; Ma, Ding-Ding; Xie, Qing; Huang, You-Guang

    2016-06-01

    Heat shock protein 90 (HSP90) is a molecular chaperone required for the stability and function of many proteins. The chaperoning of oncoproteins by HSP90 enhances the survival, growth, and invasive potential of cancer cells. HSP90 inhibitors are promising new anticancer agents, in which the benzoquinone ansamycin 17-allylamino-17-demethoxygeldanamycin (17-AAG) is currently in clinical evaluation. However, the implications of acquired resistance to this class of drug remain largely unexplored. In the present study, we have generated isogenic human colon cancer cell lines that are resistant to 17-AAG by continued culturing in the compound. Cross-resistance was found with another HSP90 inhibitor 17-dimethylaminoethylamino-17-demethoxygeldanamycin. The resistant cells showed obvious morphology changes with a metastatic phenotype and significant increases in migration and adhesion to collagens. Western blotting analysis of epithelial-mesenchymal transition molecular markers found that expression of E-cadherin downregulated, whereas expression of N-cadherin and β-catenin upregulated in the resistant cells. Mucin 1 (MUC1) has been reported to mediate metastasis as well as chemical resistance in many cancers. Here, we found that MUC1 expression was significantly elevated in the acquired drug resistance cells. 17-AAG treatment could decrease MUC1 more in parental cells than in acquired 17-AAG-resistant cells. Further study found that knockdown of MUC1 expression by small interfering RNA could obviously re-sensitize the resistant cells to 17-AAG treatment, and decrease the cell migration and adhesion. These were coupled with a downregulation in N-cadherin and β-catenin. The results indicate that HSP90 inhibitor therapies in colon carcinomas could generate resistance and increase metastatic potential that might mediated by upregulation of MUC1 expression. Findings from this study further our understanding of the potential clinical effects of HSP90-directed therapies in

  5. Comparative analysis of novel and conventional Hsp90 inhibitors on HIF activity and angiogenic potential in clear cell renal cell carcinoma: implications for clinical evaluation

    International Nuclear Information System (INIS)

    Bohonowych, Jessica ES; Peng, Shuping; Gopal, Udhayakumar; Hance, Michael W; Wing, Shane B; Argraves, Kelley M; Lundgren, Karen; Isaacs, Jennifer S

    2011-01-01

    Perturbing Hsp90 chaperone function targets hypoxia inducible factor (HIF) function in a von Hippel-Lindau (VHL) independent manner, and represents an approach to combat the contribution of HIF to cell renal carcinoma (CCRCC) progression. However, clinical trials with the prototypic Hsp90 inhibitor 17-AAG have been unsuccessful in halting the progression of advanced CCRCC. Here we evaluated a novel next generation small molecule Hsp90 inhibitor, EC154, against HIF isoforms and HIF-driven molecular and functional endpoints. The effects of EC154 were compared to those of the prototypic Hsp90 inhibitor 17-AAG and the histone deacetylase (HDAC) inhibitor LBH589. The findings indicate that EC154 is a potent inhibitor of HIF, effective at doses 10-fold lower than 17-AAG. While EC154, 17-AAG and the histone deacetylase (HDAC) inhibitor LBH589 impaired HIF transcriptional activity, CCRCC cell motility, and angiogenesis; these effects did not correlate with their ability to diminish HIF protein expression. Further, our results illustrate the complexity of HIF targeting, in that although these agents suppressed HIF transcripts with differential dynamics, these effects were not predictive of drug efficacy in other relevant assays. We provide evidence for EC154 targeting of HIF in CCRCC and for LBH589 acting as a suppressor of both HIF-1 and HIF-2 activity. We also demonstrate that 17-AAG and EC154, but not LBH589, can restore endothelial barrier function, highlighting a potentially new clinical application for Hsp90 inhibitors. Finally, given the discordance between HIF activity and protein expression, we conclude that HIF expression is not a reliable surrogate for HIF activity. Taken together, our findings emphasize the need to incorporate an integrated approach in evaluating Hsp90 inhibitors within the context of HIF suppression

  6. Targets Fishing and Identification of Calenduloside E as Hsp90AB1: Design, Synthesis, and Evaluation of Clickable Activity-Based Probe

    Science.gov (United States)

    Wang, Shan; Tian, Yu; Zhang, Jing-Yi; Xu, Hui-Bo; Zhou, Ping; Wang, Min; Lu, Sen-Bao; Luo, Yun; Wang, Min; Sun, Gui-Bo; Xu, Xu-Dong; Sun, Xiao-Bo

    2018-01-01

    Calenduloside E (CE), a natural triterpenoid compound isolated from Aralia elata, can protect against ox-LDL-induced human umbilical vein endothelial cell (HUVEC) injury in our previous reports. However, the exact targets and mechanisms of CE remain elusive. For the sake of resolving this question, we designed and synthesized a clickable activity-based probe (CE-P), which could be utilized to fish the functional targets in HUVECs using a gel-based strategy. Based on the previous studies of the structure-activity relationship (SAR), we introduced an alkyne moiety at the C-28 carboxylic group of CE, which kept the protective and anti-apoptosis activity. Via proteomic approach, one of the potential proteins bound to CE-P was identified as Hsp90AB1, and further verification was performed by pure recombinant Hsp90AB1 and competitive assay. These results demonstrated that CE could bind to Hsp90AB1. We also found that CE could reverse the Hsp90AB1 decrease after ox-LDL treatment. To make our results more convincing, we performed SPR analysis and the affinity kinetic assay showed that CE/CE-P could bind to Hsp90AB1 in a dose-dependent manner. Taken together, our research showed CE could probably bind to Hsp90AB1 to protect the cell injury, which might provide the basis for the further exploration of its cardiovascular protective mechanisms. For the sake of resolving this question, we designed and synthesized a clickable activity-based probe (CE-P), which could be utilized to fish the functional targets in HUVECs using a gel-based strategy. PMID:29875664

  7. Loss of Smyhc1 or Hsp90alpha1 function results in different effects on myofibril organization in skeletal muscles of zebrafish embryos.

    Directory of Open Access Journals (Sweden)

    Marta Codina

    Full Text Available BACKGROUND: Myofibrillogenesis requires the correct folding and assembly of sarcomeric proteins into highly organized sarcomeres. Heat shock protein 90alpha1 (Hsp90alpha1 has been implicated as a myosin chaperone that plays a key role in myofibrillogenesis. Knockdown or mutation of hsp90alpha1 resulted in complete disorganization of thick and thin filaments and M- and Z-line structures. It is not clear whether the disorganization of these sarcomeric structures is due to a direct effect from loss of Hsp90alpha1 function or indirectly through the disorganization of myosin thick filaments. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we carried out a loss-of-function analysis of myosin thick filaments via gene-specific knockdown or using a myosin ATPase inhibitor BTS (N-benzyl-p-toluene sulphonamide in zebrafish embryos. We demonstrated that knockdown of myosin heavy chain 1 (myhc1 resulted in sarcomeric defects in the thick and thin filaments and defective alignment of Z-lines. Similarly, treating zebrafish embryos with BTS disrupted thick and thin filament organization, with little effect on the M- and Z-lines. In contrast, loss of Hsp90alpha1 function completely disrupted all sarcomeric structures including both thick and thin filaments as well as the M- and Z-lines. CONCLUSION/SIGNIFICANCE: Together, these studies indicate that the hsp90alpha1 mutant phenotype is not simply due to disruption of myosin folding and assembly, suggesting that Hsp90alpha1 may play a role in the assembly and organization of other sarcomeric structures.

  8. Comparative analysis of novel and conventional Hsp90 inhibitors on HIF activity and angiogenic potential in clear cell renal cell carcinoma: implications for clinical evaluation

    Directory of Open Access Journals (Sweden)

    Bohonowych Jessica ES

    2011-12-01

    Full Text Available Abstract Background Perturbing Hsp90 chaperone function targets hypoxia inducible factor (HIF function in a von Hippel-Lindau (VHL independent manner, and represents an approach to combat the contribution of HIF to cell renal carcinoma (CCRCC progression. However, clinical trials with the prototypic Hsp90 inhibitor 17-AAG have been unsuccessful in halting the progression of advanced CCRCC. Methods Here we evaluated a novel next generation small molecule Hsp90 inhibitor, EC154, against HIF isoforms and HIF-driven molecular and functional endpoints. The effects of EC154 were compared to those of the prototypic Hsp90 inhibitor 17-AAG and the histone deacetylase (HDAC inhibitor LBH589. Results The findings indicate that EC154 is a potent inhibitor of HIF, effective at doses 10-fold lower than 17-AAG. While EC154, 17-AAG and the histone deacetylase (HDAC inhibitor LBH589 impaired HIF transcriptional activity, CCRCC cell motility, and angiogenesis; these effects did not correlate with their ability to diminish HIF protein expression. Further, our results illustrate the complexity of HIF targeting, in that although these agents suppressed HIF transcripts with differential dynamics, these effects were not predictive of drug efficacy in other relevant assays. Conclusions We provide evidence for EC154 targeting of HIF in CCRCC and for LBH589 acting as a suppressor of both HIF-1 and HIF-2 activity. We also demonstrate that 17-AAG and EC154, but not LBH589, can restore endothelial barrier function, highlighting a potentially new clinical application for Hsp90 inhibitors. Finally, given the discordance between HIF activity and protein expression, we conclude that HIF expression is not a reliable surrogate for HIF activity. Taken together, our findings emphasize the need to incorporate an integrated approach in evaluating Hsp90 inhibitors within the context of HIF suppression.

  9. Involvement of YAP, TAZ and HSP90 in contact guidance and intercellular junction formation in corneal epithelial cells.

    Directory of Open Access Journals (Sweden)

    Vijay Krishna Raghunathan

    Full Text Available The extracellular environment possesses a rich milieu of biophysical and biochemical signaling cues that are simultaneously integrated by cells and influence cellular phenotype. Yes-associated protein (YAP and transcriptional co-activator with PDZ-binding motif (WWTR1; TAZ, two important signaling molecules of the Hippo pathway, have been recently implicated as nuclear relays of cytoskeletal changes mediated by substratum rigidity and topography. These proteins intersect with other important intracellular signaling pathways (e.g. Wnt and TGFβ. In the cornea, epithelial cells adhere to the stroma through a 3-dimensional topography-rich basement membrane, with features in the nano-submicron size-scale that are capable of profoundly modulating a wide range of fundamental cell behaviors. The influences of substratum-topography, YAP/TAZ knockdown, and HSP90 inhibition on cell morphology, YAP/TAZ localization, and the expression of TGFβ2 and CTGF, were investigated. The results demonstrate (a that knockdown of TAZ enhances contact guidance in a YAP dependent manner, (b that CTGF is predominantly regulated by YAP and not TAZ, and (c that TGFβ2 is regulated by both YAP and TAZ in these cells. Additionally, inhibition of HSP90 resulted in nuclear localization and subsequent transcriptional-activation of YAP, formation of cell-cell junctions and co-localization of E-cadherin and β-catenin at adherens junctions. Results presented in this study reflect the complexities underlying the molecular relationships between the cytoskeleton, growth factors, heat shock proteins, and co-activators of transcription that impact mechanotransduction. The data reveal the importance of YAP/TAZ on the cell behaviors, and gene and protein expression.

  10. Phase I Evaluation of STA-1474, a Prodrug of the Novel HSP90 Inhibitor Ganetespib, in Dogs with Spontaneous Cancer

    Science.gov (United States)

    London, Cheryl A.; Bear, Misty D.; McCleese, Jennifer; Foley, Kevin P.; Paalangara, Reji; Inoue, Takayo; Ying, Weiwen; Barsoum, James

    2011-01-01

    Background The novel water soluble compound STA-1474 is metabolized to ganetespib (formerly STA-9090), a potent HSP90 inhibitor previously shown to kill canine tumor cell lines in vitro and inhibit tumor growth in the setting of murine xenografts. The purpose of the following study was to extend these observations and investigate the safety and efficacy of STA-1474 in dogs with spontaneous tumors. Methods and Findings This was a Phase 1 trial in which dogs with spontaneous tumors received STA-1474 under one of three different dosing schemes. Pharmacokinetics, toxicities, biomarker changes, and tumor responses were assessed. Twenty-five dogs with a variety of cancers were enrolled. Toxicities were primarily gastrointestinal in nature consisting of diarrhea, vomiting, inappetence and lethargy. Upregulation of HSP70 protein expression was noted in both tumor specimens and PBMCs within 7 hours following drug administration. Measurable objective responses were observed in dogs with malignant mast cell disease (n = 3), osteosarcoma (n = 1), melanoma (n = 1) and thyroid carcinoma (n = 1), for a response rate of 24% (6/25). Stable disease (>10 weeks) was seen in 3 dogs, for a resultant overall biological activity of 36% (9/25). Conclusions This study provides evidence that STA-1474 exhibits biologic activity in a relevant large animal model of cancer. Given the similarities of canine and human cancers with respect to tumor biology and HSP90 activation, it is likely that STA-1474 and ganetespib will demonstrate comparable anti-cancer activity in human patients. PMID:22073242

  11. Discovery of 2',4'-dimethoxychalcone as a Hsp90 inhibitor and its effect on iressa-resistant non-small cell lung cancer (NSCLC).

    Science.gov (United States)

    Seo, Young Ho

    2015-10-01

    Heat shock protein 90 (Hsp90) is a ATP dependent molecular chaperone and has emerged as an attractive therapeutic target in the war on cancer due to its role in regulating maturation and stabilization of numerous oncogenic proteins. In this study, we discovered that 2',4'-dimethoxychalcone (1b) disrupted Hsp90 chaperoning function and inhibited the growth of iressa-resistant non-small cell lung cancer (NSCLC, H1975). The result suggested that 2',4'-dimethoxychalcone (1b) could serve as a potential therapeutic lead to circumvent the drug resistance acquired by EGFR mutation and Met amplification.

  12. The Hsp90 Complex in Microbes and Man | Center for Cancer Research

    Science.gov (United States)

    Why would cancer researchers be interested in how a bacteria named Escherichia coli (E. coli) rebuilds its cellular proteins after they have been inactivated by environmental stress such as heat?  The answer lies in a protein remodeling mechanism that is shared by microbes and man.

  13. Clinical Development of Gamitrinib, a Novel Mitochondrial-Targeted Small Molecule Hsp90 Inhibitor

    Science.gov (United States)

    2016-09-01

    controls the ATP supply in cancer cells? Biochemistry lessons to understand cancer energy metabolism. Int J BiochemCell Biol. 2014; 50:10–23. Epub 2014...silencing of Akt1 or Akt2 (Figure 6G) in hypoxia increased the phosphorylation of the energy stress sensor, AMP -regulated kinase (AMPK). This response was

  14. Clinical Development of Gamitrinib, a Novel Mitochondrial-Targeted Small Molecule Hsp90 Inhibitor

    Science.gov (United States)

    2015-09-01

    and Languino LR. β1 integrins mediate cell proliferation in three-dimensional cultures by regulating expression of the sonic hedgehog effector protein...unknown, but protein folding quality control within the unique anatomy of mitochondria (18) is required to buffer the risk of proteotoxic stress (19

  15. Clinical Development of Gamitrinib, a Novel Mitochondrial-Targeted Small Molecule Hsp90 Inhibitor

    Science.gov (United States)

    2014-09-01

    experiments, exposure of tumour cells to concentrations 450 mM of the oxidative agent, hydrogen peroxide (H2O2), reduced SDHB levels (Fig. 3d). siRNA...and potentially ‘ addictive ’ survival factor for these cells. There is now intense interest in pursuing aberrant tumour cell metabolism for cancer...cellular homeostasis (Ellis, 2007) by buffering proteotoxic stress and ensuring flexible adaptation to environmental cues (Balch et al., 2008). In

  16. 17-Allylamino-17-demethoxygeldanamycin induces downregulation of critical Hsp90 protein clients and results in cell cycle arrest and apoptosis of human urinary bladder cancer cells

    International Nuclear Information System (INIS)

    Karkoulis, Panagiotis K; Stravopodis, Dimitrios J; Margaritis, Lukas H; Voutsinas, Gerassimos E

    2010-01-01

    17-Allylamino-17-demethoxygeldanamycin (17-AAG), a benzoquinone ansamycin antibiotic, specifically targets heat shock protein 90 (Hsp90) and interferes with its function as a molecular chaperone that maintains the structural and functional integrity of various protein clients involved in cellular signaling. In this study, we have investigated the effect of 17-AAG on the regulation of Hsp90-dependent signaling pathways directly implicated in cell cycle progression, survival and motility of human urinary bladder cancer cell lines. We have used MTT-based assays, FACS analysis, Western blotting, semi-quantitative RT-PCR, immunocytochemistry and scratch-wound assay in RT4, RT112 and T24 human urinary bladder cancer cell lines. We have demonstrated that, upon 17-AAG treatment, bladder cancer cells are arrested in the G1 phase of the cell cycle and eventually undergo apoptotic cell death in a dose-dependent manner. Furthermore, 17-AAG administration was shown to induce a pronounced downregulation of multiple Hsp90 protein clients and other downstream effectors, such as IGF-IR, Akt, IKK-α, IKK-β, FOXO1, ERK1/2 and c-Met, resulting in sequestration-mediated inactivation of NF-κB, reduced cell proliferation and decline of cell motility. In total, we have clearly evinced a dose-dependent and cell type-specific effect of 17-AAG on cell cycle progression, survival and motility of human bladder cancer cells, due to downregulation of multiple Hsp90 clients and subsequent disruption of signaling integrity

  17. Mode of cell death induced by the HSP90 inhibitor 17-AAG (tanespimycin) is dependent on the expression of pro-apoptotic BAX.

    Science.gov (United States)

    Powers, Marissa V; Valenti, Melanie; Miranda, Susana; Maloney, Alison; Eccles, Suzanne A; Thomas, George; Clarke, Paul A; Workman, Paul

    2013-11-01

    Inhibitors of the molecular chaperone heat shock protein 90 (HSP90) are of considerable current interest as targeted cancer therapeutic agents because of the ability to destabilize multiple oncogenic client proteins. Despite their resulting pleiotropic effects on multiple oncogenic pathways and hallmark traits of cancer, resistance to HSP90 inhibitors is possible and their ability to induce apoptosis is less than might be expected. Using an isogenic model for BAX knockout in HCT116 human colon carcinoma cells, we demonstrate the induction of BAX-dependent apoptosis at pharmacologically relevant concentrations of the HSP90 inhibitor 17-AAG both in vitro and in tumor xenografts in vivo. Removal of BAX expression by homologous recombination reduces apoptosis in vitro and in vivo but allows a lower level of cell death via a predominantly necrotic mechanism. Despite reducing apoptosis, the loss of BAX does not alter the overall sensitivity to 17-AAG in vitro or in vivo. The results indicate that 17-AAG acts predominantly to cause a cytostatic antiproliferative effect rather than cell death and further suggest that BAX status may not alter the overall clinical response to HSP90 inhibitors. Other agents may be required in combination to enhance tumor-selective killing by these promising drugs. In addition, there are implications for the use of apoptotic endpoints in the assessment of the activity of molecularly targeted agents.

  18. Hsp90α forms a stable complex at the cilium neck for the interaction of signalling molecules in IGF-1 receptor signalling.

    Science.gov (United States)

    Wang, Hongzhong; Zou, Xinle; Wei, Zhuang; Wu, Yuan; Li, Rongxia; Zeng, Rong; Chen, Zhengjun; Liao, Kan

    2015-01-01

    The primary cilium is composed of an axoneme that protrudes from the cell surface, a basal body beneath the membrane and a transition neck in between. It is a sensory organelle on the plasma membrane, involved in mediating extracellular signals. In the transition neck region of the cilium, the microtubules change from triplet to doublet microtubules. This region also contains the transition fibres that crosslink the axoneme with the membrane and the necklace proteins that regulate molecules being transported into and out of the cilium. In this protein-enriched, complex area it is important to maintain the correct assembly of all of these proteins. Here, through immunofluorescent staining and protein isolation, we identify the molecular chaperone Hsp90α clustered at the periciliary base. At the transition neck region, phosphorylated Hsp90α forms a stable ring around the axoneme. Heat shock treatment causes Hsp90α to dissipate and induces resorption of cilia. We further identify that Hsp90α at the transition neck region represents a signalling platform on which IRS-1 interacts with intracellular downstream signalling molecules involved in IGF-1 receptor signalling. © 2015. Published by The Company of Biologists Ltd.

  19. Inhibition of Hsp90 acts synergistically with topoisomerase II poisons to increase the apoptotic killing of cells due to an increase in topoisomerase II mediated DNA damage.

    Science.gov (United States)

    Barker, Catherine R; McNamara, Anne V; Rackstraw, Stephen A; Nelson, David E; White, Mike R; Watson, Alastair J M; Jenkins, John R

    2006-01-01

    Topoisomerase II plays a crucial role during chromosome condensation and segregation in mitosis and meiosis and is a highly attractive target for chemotherapeutic agents. We have identified previously topoisomerase II and heat shock protein 90 (Hsp90) as part of a complex. In this paper we demonstrate that drug combinations targeting these two enzymes cause a synergistic increase in apoptosis. The objective of our study was to identify the mode of cell killing and the mechanism behind the increase in topoisomerase II mediated DNA damage. Importantly we demonstrate that Hsp90 inhibition results in an increased topoiosmerase II activity but not degradation of topoisomerase II and it is this, in the presence of a topoisomerase II poison that causes the increase in cell death. Our results suggest a novel mechanism of action where the inhibition of Hsp90 disrupts the Hsp90-topoisomerase II interaction leading to an increase in and activation of unbound topoisomerase II, which, in the presence of a topoisomerase II poison leads to the formation of an increased number of cleavable complexes ultimately resulting in rise in DNA damage and a subsequent increase cell death.

  20. Establish an automated flow injection ESI-MS method for the screening of fragment based libraries: Application to Hsp90.

    Science.gov (United States)

    Riccardi Sirtori, Federico; Caronni, Dannica; Colombo, Maristella; Dalvit, Claudio; Paolucci, Mauro; Regazzoni, Luca; Visco, Carlo; Fogliatto, Gianpaolo

    2015-08-30

    ESI-MS is a well established technique for the study of biopolymers (nucleic acids, proteins) and their non covalent adducts, due to its capacity to detect ligand-target complexes in the gas phase and allows inference of ligand-target binding in solution. In this article we used this approach to investigate the interaction of ligands to the Heat Shock Protein 90 (Hsp90). This enzyme is a molecular chaperone involved in the folding and maturation of several proteins which has been subjected in the last years to intensive drug discovery efforts due to its key role in cancer. In particular, reference compounds, with a broad range of dissociation constants from 40pM to 100μM, were tested to assess the reliability of ESI-MS for the study of protein-ligand complexes. A good agreement was found between the values measured with a fluorescence polarization displacement assay and those determined by mass spectrometry. After this validation step we describe the setup of a medium throughput screening method, based on ESI-MS, suitable to explore interactions of therapeutic relevance biopolymers with chemical libraries. Our approach is based on an automated flow injection ESI-MS method (AFI-MS) and has been applied to screen the Nerviano Medical Sciences proprietary fragment library of about 2000 fragments against Hsp90. In order to discard false positive hits and to discriminate those of them interacting with the N-terminal ATP binding site, competition experiments were performed using a reference inhibitor. Gratifyingly, this group of hits matches with the ligands previously identified by NMR FAXS techniques and confirmed by X-ray co-crystallization experiments. These results support the use of AFI-MS for the screening of medium size libraries, including libraries of small molecules with low affinity typically used in fragment based drug discovery. AFI-MS is a valid alternative to other techniques with the additional opportunities to identify compounds interacting with

  1. Computational modeling of allosteric regulation in the hsp90 chaperones: a statistical ensemble analysis of protein structure networks and allosteric communications.

    Directory of Open Access Journals (Sweden)

    Kristin Blacklock

    2014-06-01

    Full Text Available A fundamental role of the Hsp90 chaperone in regulating functional activity of diverse protein clients is essential for the integrity of signaling networks. In this work we have combined biophysical simulations of the Hsp90 crystal structures with the protein structure network analysis to characterize the statistical ensemble of allosteric interaction networks and communication pathways in the Hsp90 chaperones. We have found that principal structurally stable communities could be preserved during dynamic changes in the conformational ensemble. The dominant contribution of the inter-domain rigidity to the interaction networks has emerged as a common factor responsible for the thermodynamic stability of the active chaperone form during the ATPase cycle. Structural stability analysis using force constant profiling of the inter-residue fluctuation distances has identified a network of conserved structurally rigid residues that could serve as global mediating sites of allosteric communication. Mapping of the conformational landscape with the network centrality parameters has demonstrated that stable communities and mediating residues may act concertedly with the shifts in the conformational equilibrium and could describe the majority of functionally significant chaperone residues. The network analysis has revealed a relationship between structural stability, global centrality and functional significance of hotspot residues involved in chaperone regulation. We have found that allosteric interactions in the Hsp90 chaperone may be mediated by modules of structurally stable residues that display high betweenness in the global interaction network. The results of this study have suggested that allosteric interactions in the Hsp90 chaperone may operate via a mechanism that combines rapid and efficient communication by a single optimal pathway of structurally rigid residues and more robust signal transmission using an ensemble of suboptimal multiple

  2. The HSP90 inhibitor 17-AAG synergizes with doxorubicin and U0126 in anaplastic large cell lymphoma irrespective of ALK expression.

    Science.gov (United States)

    Georgakis, Georgios V; Li, Yang; Rassidakis, Georgios Z; Medeiros, L Jeffrey; Younes, Anas

    2006-12-01

    Heat shock protein 90 (HSP90) chaperones and maintains the molecular integrity of a variety of signal transduction proteins, including the nucleophosmin-anaplastic lymphoma kinase (NPM-ALK) oncogenic protein, a genetic abnormality that is frequently observed in anaplastic large cell lymphoma (ALCL) cells. Here we demonstrate that HSP90 is overexpressed in primary and cultured ALK-positive and ALK-negative ALCL cells, and we evaluate the potential role of the small molecule inhibitor of HSP90, 17-allylamino-17-demethoxygeldanamycin (17-AAG) in treating ALCL. The antiproliferative effect of 17-AAG-cultured cells was determined by MTS assay. Apoptosis and cell-cycle arrest were determined by Annexin-V/propidium iodide and propidium iodide staining, respectively, and fluorescein-activated cell sorting analysis. Expression of HSP90 was evaluated by immunohistochemistry, and molecular changes were determined by Western blot. Treatment of cultured ALCL cells with 17-AAG induced cell-cycle arrest and apoptosis, irrespective of ALK expression. At the molecular level, 17-AAG induced degradation of ALK and Akt proteins, dephosphorylated extracellular signal-regulated kinase, and degraded the cell-cycle regulatory protein cyclin D1 and its cyclin-dependent kinases, CDK4 and CDK6, but had a differential effect on p27 and p53 proteins. Inhibition of extracellular signal-regulated kinase phosphorylation by the mitogen activated protein kinase inhibitor U0126 induced cell death in all ALCL cell lines, and sublethal concentration 17-AAG showed synergistic antiproliferative effects when combined with U0126 or doxorubicin. Our data demonstrate that targeting HSP90 function by 17-AAG may offer a novel therapeutic strategy for ALCL, either as single-agent activity or by combining 17-AAG with conventional or targeted therapeutic schemes.

  3. TAS-116, a novel Hsp90 inhibitor, selectively enhances radio-sensitivity of human cancer cells to X-rays and carbon ion radiation

    Science.gov (United States)

    Lee, Younghyun; Sunada, Shigeaki; Hirakawa, Hirokazu; Fujimori, Akira; Nickoloff, Jac A.; Okayasu, Ryuichi

    2016-01-01

    Hsp90 inhibitors have been investigated as cancer therapeutics in mono-therapy and to augment radiotherapy, however serious adverse effects of early generation Hsp90 inhibitors limited their development. TAS-116 is a novel Hsp90 inhibitor with lower adverse effects than other Hsp90 inhibitors, and here we investigated the radio-sensitizing effects of TAS-116 in low LET X-ray, and high LET carbon ion irradiated human cancer cells and mouse tumor xenografts. TAS-116 decreased cell survival of both X-ray and carbon ion-irradiated human cancer cell lines (HeLa and H1299 cells), and similar to other Hsp90 inhibitors, it did not affect radiosensitivity of non-cancerous human fibroblasts. TAS-116 increased the number of radiation-induced γ-H2AX foci, and delayed the repair of DNA double-strand breaks (DSBs). TAS-116 reduced the expression of proteins that mediate repair of DSBs by homologous recombination (RAD51) and non-homologous end joining (Ku, DNA-PKcs), and suppressed formation of RAD51 foci and phosphorylation/activation of DNA-PKcs. TAS-116 also decreased expression of the cdc25 cell cycle progression marker, markedly increasing G2/M arrest. Combined treatment of mouse tumor xenografts with carbon ions and TAS-116 showed promising delay in tumor growth compared to either individual treatment. These results demonstrate that TAS-116 radio-sensitizes human cancer cells to both X rays and carbon ions by inhibiting the two major DSB repair pathways, and these effects were accompanied by marked cell cycle arrest. The promising results of combination TAS-116 + carbon ion radiation therapy of tumor xenografts justify further exploration of TAS-116 as an adjunct to radiotherapy using low or high LET radiation. PMID:28062703

  4. ATM Is Required for the Prolactin-Induced HSP90-Mediated Increase in Cellular Viability and Clonogenic Growth After DNA Damage.

    Science.gov (United States)

    Karayazi Atici, Ödül; Urbanska, Anna; Gopinathan, Sesha Gopal; Boutillon, Florence; Goffin, Vincent; Shemanko, Carrie S

    2018-02-01

    Prolactin (PRL) acts as a survival factor for breast cancer cells, but the PRL signaling pathway and the mechanism are unknown. Previously, we identified the master chaperone, heat shock protein 90 (HSP90) α, as a prolactin-Janus kinase 2 (JAK2)-signal transducer and activator of transcription 5 (STAT5) target gene involved in survival, and here we investigated the role of HSP90 in the mechanism of PRL-induced viability in response to DNA damage. The ataxia-telangiectasia mutated kinase (ATM) protein plays a critical role in the cellular response to double-strand DNA damage. We observed that PRL increased viability of breast cancer cells treated with doxorubicin or etoposide. The increase in cellular resistance is specific to the PRL receptor, because the PRL receptor antagonist, Δ1-9-G129R-hPRL, prevented the increase in viability. Two different HSP90 inhibitors, 17-allylamino-17-demethoxygeldanamycin and BIIB021, reduced the PRL-mediated increase in cell viability of doxorubicin-treated cells and led to a decrease in JAK2, ATM, and phosphorylated ATM protein levels. Inhibitors of JAK2 (G6) and ATM (KU55933) abolished the PRL-mediated increase in cell viability of DNA-damaged cells, supporting the involvement of each, as well as the crosstalk of ATM with the PRL pathway in the context of DNA damage. Drug synergism was detected between the ATM inhibitor (KU55933) and doxorubicin and between the HSP90 inhibitor (BIIB021) and doxorubicin. Short interfering RNA directed against ATM prevented the PRL-mediated increase in cell survival in two-dimensional cell culture, three-dimensional collagen gel cultures, and clonogenic cell survival, after doxorubicin treatment. Our results indicate that ATM contributes to the PRL-JAK2-STAT5-HSP90 pathway in mediating cellular resistance to DNA-damaging agents. Copyright © 2018 Endocrine Society.

  5. The HSP90 inhibitor NVP-AUY922 radiosensitizes by abrogation of homologous recombination resulting in mitotic entry with unresolved DNA damage.

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    Shane Zaidi

    Full Text Available Heat shock protein 90 (HSP90 is a molecular chaperone responsible for the conformational maintenance of a number of client proteins that play key roles in cell cycle arrest, DNA damage repair and apoptosis following radiation. HSP90 inhibitors exhibit antitumor activity by modulating the stabilisation and activation of HSP90 client proteins. We sought to evaluate NVP-AUY922, the most potent HSP90 inhibitor yet reported, in preclinical radiosensitization studies.NVP-AUY922 potently radiosensitized cells in vitro at low nanomolar concentrations with a concurrent depletion of radioresistance-linked client proteins. Radiosensitization by NVP-AUY922 was verified for the first time in vivo in a human head and neck squamous cell carcinoma xenograft model in athymic mice, as measured by delayed tumor growth and increased surrogate end-point survival (p = <0.0001. NVP-AUY922 was shown to ubiquitously inhibit resolution of dsDNA damage repair correlating to delayed Rad51 foci formation in all cell lines tested. Additionally, NVP-AUY922 induced a stalled mitotic phenotype, in a cell line-dependent manner, in HeLa and HN5 cell lines irrespective of radiation exposure. Cell cycle analysis indicated that NVP-AUY922 induced aberrant mitotic entry in all cell lines tested in the presence of radiation-induced DNA damage due to ubiquitous CHK1 depletion, but resultant downstream cell cycle effects were cell line dependent.These results identify NVP-AUY922 as the most potent HSP90-mediated radiosensitizer yet reported in vitro, and for the first time validate it in a clinically relevant in vivo model. Mechanistic analysis at clinically achievable concentrations demonstrated that radiosensitization is mediated by the combinatorial inhibition of cell growth and survival pathways, ubiquitous delay in Rad51-mediated homologous recombination and CHK1-mediated G(2/M arrest, but that the contribution of cell cycle perturbation to radiosensitization may be cell line

  6. Hsp40s specify functions of Hsp104 and Hsp90 protein chaperone machines.

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    Michael Reidy

    2014-10-01

    Full Text Available Hsp100 family chaperones of microorganisms and plants cooperate with the Hsp70/Hsp40/NEF system to resolubilize and reactivate stress-denatured proteins. In yeast this machinery also promotes propagation of prions by fragmenting prion polymers. We previously showed the bacterial Hsp100 machinery cooperates with the yeast Hsp40 Ydj1 to support yeast thermotolerance and with the yeast Hsp40 Sis1 to propagate [PSI+] prions. Here we find these Hsp40s similarly directed specific activities of the yeast Hsp104-based machinery. By assessing the ability of Ydj1-Sis1 hybrid proteins to complement Ydj1 and Sis1 functions we show their C-terminal substrate-binding domains determined distinctions in these and other cellular functions of Ydj1 and Sis1. We find propagation of [URE3] prions was acutely sensitive to alterations in Sis1 activity, while that of [PIN+] prions was less sensitive than [URE3], but more sensitive than [PSI+]. These findings support the ideas that overexpressing Ydj1 cures [URE3] by competing with Sis1 for interaction with the Hsp104-based disaggregation machine, and that different prions rely differently on activity of this machinery, which can explain the various ways they respond to alterations in chaperone function.

  7. [Effects of HSP90 inhibitor 17-AAG on cell cycle and apoptosis of human gastric cancer cell lines SGC-7901].

    Science.gov (United States)

    Chen, Meini; Xu, Jinghong; Zhao, Jumei

    2013-02-01

    To study the effect of the HSP90 inhibitor, 17-allylamino-17-demethoxygeldanamycin (17-AAG), on cell proliferation and apoptosis of human cancer SGC-7901 cells and explore the mechanisms. The inhibitory effect of 17-AAG on the proliferation and morphology of SGC-7901 cells was assessed with MTT assay and DNA-PI staining, respectively. Flow cytometry was employed to analyze the changes in cell cycle and apoptosis of the cells following 17-AAG exposure. The cellular expression of Fas protein was detected by immunohistochemistry. 17-AAG significantly suppressed the proliferation of SGC-7901 cells in a time- and dose-dependent manner. After treatment with 17-AAG for 48 h, SGC-7901 cells showed cell cycle arrested at G(2)/M stage, and the cell apoptosis rate increased with the 17-AAG concentration. The expression of Fas protein in the cytoplasm of SGC-7901 cells increased gradually with the increase of 17-AAG concentration. 17-AAG can induce apoptosis, alters the cell cycle distribution and up-regulates the expression of Fas protein in SGC-7901 cells to suppress the cell proliferation.

  8. Neratinib induces ErbB2 ubiquitylation and endocytic degradation via HSP90 dissociation in breast cancer cells.

    Science.gov (United States)

    Zhang, Yingqiu; Zhang, Jinrui; Liu, Congcong; Du, Sha; Feng, Lu; Luan, Xuelin; Zhang, Yayun; Shi, Yulin; Wang, Taishu; Wu, Yue; Cheng, Wei; Meng, Songshu; Li, Man; Liu, Han

    2016-11-28

    Receptor tyrosine kinase ErbB2/HER2 is frequently observed to be overexpressed in human cancers, leading to over activation of downstream signaling modules. HER2 positive is a major type of breast cancer for which ErbB2 targeting is already proving to be an effective therapeutic strategy. Apart from antibodies against ErbB2, the small molecule tyrosine kinase inhibitor lapatinib has had successful clinical outcomes, and other inhibitors such as neratinib are currently undergoing clinical investigations. In this study we report the effects of lapatinib and neratinib on the mRNA and protein levels of the ErbB2 receptor. We provide evidence that neratinib-induced down regulation of ErbB2 occurs through ubiquitin-mediated endocytic sorting and lysosomal degradation. At the mechanistic level, neratinib treatment leads to HSP90 release from ErbB2 and its subsequent ubiquitylation and endocytic degradation. Our findings provide novel insights into the mechanism of ErbB2 inhibition by neratinib. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  9. Down-regulation of cellular FLICE-inhibitory protein (Long Form contributes to apoptosis induced by Hsp90 inhibition in human lung cancer cells

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    Wang Qilin

    2012-12-01

    Full Text Available Abstract Background Cellular FLICE-Inhibitory Protein (long form, c-FLIPL is a critical negative regulator of death receptor-mediated apoptosis. Overexpression of c-FLIPL has been reported in many cancer cell lines and is associated with chemoresistance. In contrast, down-regulation of c-FLIP may drive cancer cells into cellular apoptosis. This study aims to demonstrate that inhibition of the heat shock protein 90 (Hsp90 either by inhibitors geldanamycin/17-N-Allylamino-17-demethoxygeldanamycin (GA/17-AAG or siRNA technique in human lung cancer cells induces c-FLIPL degradation and cellular apoptosis through C-terminus of Hsp70-interacting protein (CHIP-mediated mechanisms. Methods Calu-1 and H157 cell lines (including H157-c-FLIPL overexpressing c-FLIPL and control cell H157-lacZ were treated with 17-AAG and the cell lysates were prepared to detect the given proteins by Western Blot and the cell survival was assayed by SRB assay. CHIP and Hsp90 α/β proteins were knocked down by siRNA technique. CHIP and c-FLIPL plasmids were transfected into cells and immunoprecipitation experiments were performed to testify the interactions between c-FLIPL, CHIP and Hsp90. Results c-FLIPL down-regulation induced by 17-AAG can be reversed with the proteasome inhibitor MG132, which suggested that c-FLIPL degradation is mediated by a ubiquitin-proteasome system. Inhibition of Hsp90α/β reduced c-FLIPL level, whereas knocking down CHIP expression with siRNA technique inhibited c-FLIPL degradation. Furthermore, c-FLIPL and CHIP were co-precipitated in the IP complexes. In addition, overexpression of c-FLIPL can rescue cancer cells from apoptosis. When 17-AAG was combined with an anti-cancer agent celecoxib(CCB, c-FLIPL level declined further and there was a higher degree of caspase activation. Conclusion We have elucidated c-FLIPL degradation contributes to apoptosis induced by Hsp90 inhibition, suggesting c-FLIP and Hsp90 may be the promising combined targets

  10. Histoplasma capsulatum heat-shock 60 orchestrates the adaptation of the fungus to temperature stress.

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    Allan Jefferson Guimarães

    2011-02-01

    Full Text Available Heat shock proteins (Hsps are among the most widely distributed and evolutionary conserved proteins. Hsps are essential regulators of diverse constitutive metabolic processes and are markedly upregulated during stress. A 62 kDa Hsp (Hsp60 of Histoplasma capsulatum (Hc is an immunodominant antigen and the major surface ligand to CR3 receptors on macrophages. However little is known about the function of this protein within the fungus. We characterized Hc Hsp60-protein interactions under different temperature to gain insights of its additional functions oncell wall dynamism, heat stress and pathogenesis. We conducted co-immunoprecipitations with antibodies to Hc Hsp60 using cytoplasmic and cell wall extracts. Interacting proteins were identified by shotgun proteomics. For the cell wall, 84 common interactions were identified among the 3 growth conditions, including proteins involved in heat-shock response, sugar and amino acid/protein metabolism and cell signaling. Unique interactions were found at each temperature [30°C (81 proteins, 37°C (14 and 37/40°C (47]. There were fewer unique interactions in cytoplasm [30°C (6, 37°C (25 and 37/40°C (39] and four common interactions, including additional Hsps and other known virulence factors. These results show the complexity of Hsp60 function and provide insights into Hc biology, which may lead to new avenues for the management of histoplasmosis.

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

    Science.gov (United States)

    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.

  12. Organoids with cancer stem cell-like properties secrete exosomes and HSP90 in a 3D nanoenvironment

    Science.gov (United States)

    Okusha, Yuka; Uchibe, Kenta; Iinuma, Ryosuke; Ono, Kisho; Nakano, Keisuke; Murakami, Jun; Itoh, Manabu; Arai, Kazuya; Fujiwara, Toshifumi; Namba, Yuri; Murata, Yoshiki; Ohyama, Kazumi; Shimomura, Manami; Okamura, Hirohiko; Takigawa, Masaharu; Nakatsura, Tetsuya; Kozaki, Ken-ichi; Okamoto, Kuniaki; Calderwood, Stuart K.

    2018-01-01

    Ability to form cellular aggregations such as tumorspheres and spheroids have been used as a morphological marker of malignant cancer cells and in particular cancer stem cells (CSC). However, the common definition of the types of cellular aggregation formed by cancer cells has not been available. We examined morphologies of 67 cell lines cultured on three dimensional morphology enhancing NanoCulture Plates (NCP) and classified the types of cellular aggregates that form. Among the 67 cell lines, 49 cell lines formed spheres or spheroids, 8 cell lines formed grape-like aggregation (GLA), 8 cell lines formed other types of aggregation, and 3 cell lines formed monolayer sheets. Seven GLA-forming cell lines were derived from adenocarcinoma among the 8 lines. A neuroendocrine adenocarcinoma cell line PC-3 formed asymmetric GLA with ductal structures on the NCPs and rapidly growing asymmetric tumors that metastasized to lymph nodes in immunocompromised mice. In contrast, another adenocarcinoma cell line DU-145 formed spheroids in vitro and spheroid-like tumors in vivo that did not metastasize to lymph nodes until day 50 after transplantation. Culture in the 3D nanoenvironment and in a defined stem cell medium enabled the neuroendocrine adenocarcinoma cells to form slowly growing large organoids that expressed multiple stem cell markers, neuroendocrine markers, intercellular adhesion molecules, and oncogenes in vitro. In contrast, the more commonly used 2D serum-contained environment reduced intercellular adhesion and induced mesenchymal transition and promoted rapid growth of the cells. In addition, the 3D stemness nanoenvironment promoted secretion of HSP90 and EpCAM-exosomes, a marker of CSC phenotype, from the neuroendocrine organoids. These findings indicate that the NCP-based 3D environment enables cells to form stem cell tumoroids with multipotency and model more accurately the in vivo tumor status at the levels of morphology and gene expression. PMID:29415026

  13. Differential Proteomics Identification of HSP90 as Potential Serum Biomarker in Hepatocellular Carcinoma by Two-dimensional Electrophoresis and Mass Spectrometry

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    Yiyi Sun

    2010-03-01

    Full Text Available The aim of the current study is to identify the potential biomarkers involved in Hepatocellular carcinoma (HCC carcinogenesis. A comparative proteomics approach was utilized to identify the differentially expressed proteins in the serum of 10 HCC patients and 10 controls. A total of 12 significantly altered proteins were identified by mass spectrometry. Of the 12 proteins identified, HSP90 was one of the most significantly altered proteins and its over-expression in the serum of 20 HCC patients was confirmed using ELISA analysis. The observations suggest that HSP90 might be a potential biomarker for early diagnosis, prognosis, and monitoring in the therapy of HCC. This work demonstrates that a comprehensive strategy of proteomic identification combined with further validation should be adopted in the field of cancer biomarker discovery.

  14. Receptor ligand-triggered resistance to alectinib and its circumvention by Hsp90 inhibition in EML4-ALK lung cancer cells.

    Science.gov (United States)

    Tanimoto, Azusa; Yamada, Tadaaki; Nanjo, Shigeki; Takeuchi, Shinji; Ebi, Hiromichi; Kita, Kenji; Matsumoto, Kunio; Yano, Seiji

    2014-07-15

    Alectinib is a new generation ALK inhibitor with activity against the gatekeeper L1196M mutation that showed remarkable activity in a phase I/II study with echinoderm microtubule associated protein-like 4 (EML4)--anaplastic lymphoma kinase (ALK) non-small cell lung cancer (NSCLC) patients. However, alectinib resistance may eventually develop. Here, we found that EGFR ligands and HGF, a ligand of the MET receptor, activate EGFR and MET, respectively, as alternative pathways, and thereby induce resistance to alectinib. Additionally, the heat shock protein 90 (Hsp90) inhibitor suppressed protein expression of ALK, MET, EGFR, and AKT, and thereby induced apoptosis in EML4-ALK NSCLC cells, even in the presence of EGFR ligands or HGF. These results suggest that Hsp90 inhibitors may overcome ligand-triggered resistance to new generation ALK inhibitors and may result in more successful treatment of NSCLC patients with EML4-ALK.

  15. Chromosomal assignment of six genes (EIF4G3, HSP90, RBBP6, IL8, TERT, and TERC) in four species of the genus Equus.

    Science.gov (United States)

    Vidale, Pamela; Piras, Francesca M; Nergadze, Solomon G; Bertoni, Livia; Verini-Supplizi, Andrea; Adelson, David; Guérin, Gérard; Giulotto, Elena

    2011-01-01

    We mapped six genes (EIF4G3, HSP90, RBBP6, IL8, TERT, and TERC) on the chromosomes of Equus caballus, Equus asinus, Equus grevyi, and Equus burchelli by fluorescence in situ hybridization. Our results add six type I markers to the cytogenetic map of these species and provide new information on the comparative genomics of the genus Equus. Copyright © Taylor & Francis Group, LLC

  16. Reactive oxygen species-dependent HSP90 protein cleavage participates in arsenical As+3- and MMA+3-induced apoptosis through inhibition of telomerase activity via JNK activation

    International Nuclear Information System (INIS)

    Shen, S.-C.; Yang, L.-Y.; Lin, H.-Y.; Wu, C.-Y.; Su, T.-H.; Chen, Y.-C.

    2008-01-01

    The effects of six arsenic compounds including As +3 , MMA +3 , DMA +3 , As +5 , MMA +5 , and DMA +5 on the viability of NIH3T3 cells were examined. As +3 and MMA +3 , but not the others, exhibited significant cytotoxic effects in NIH3T3 cells through apoptosis induction. The apoptotic events such as DNA fragmentation and chromosome condensation induced by As +3 and MMA +3 were prevented by the addition of NAC and CAT, and induction of HO-1 gene expression in accordance with cleavage of the HSP90 protein, and suppression of telomerase activity were observed in NIH3T3 cells under As +3 and MMA +3 treatments. An increase in the intracellular peroxide level was examined in As +3 - and MMA +3 -treated NIH3T3 cells, and As +3 - and MMA +3 -induced apoptotic events were blocked by NAC, CAT, and DPI addition. HSP90 inhibitors, GA and RD, significantly attenuated the telomerase activity in NIH3T3 cells with an enhancement of As +3 - and MMA +3 -induced cytotoxicity. Suppression of JNKs significantly inhibited As +3 - and MMA +3 -induced apoptosis by blocking HSP90 protein cleavage and telomerase reduction in NIH3T3 cells. Furthermore, Hb, SnPP, and dexferosamine showed no effect against As +3 - and MMA +3 -induced apoptosis, and overexpression of HO-1 protein or inhibition of HO-1 protein expression did not affect the apoptosis induced by As +3 or MMA +3 . These data provide the first evidence to indicate that apoptosis induced by As +3 and MMA +3 is mediated by an ROS-dependent degradation of HSP90 protein and reduction of telomerase via JNK activation, and HO-1 induction might not be involved

  17. Inhibition of Hsp90 acts synergistically with topoisomerase II poisons to increase the apoptotic killing of cells due to an increase in topoisomerase II mediated DNA damage

    OpenAIRE

    Barker, Catherine R.; McNamara, Anne V.; Rackstraw, Stephen A.; Nelson, David E.; White, Mike R.; Watson, Alastair J. M.; Jenkins, John R.

    2006-01-01

    Topoisomerase II plays a crucial role during chromosome condensation and segregation in mitosis and meiosis and is a highly attractive target for chemotherapeutic agents. We have identified previously topoisomerase II and heat shock protein 90 (Hsp90) as part of a complex. In this paper we demonstrate that drug combinations targeting these two enzymes cause a synergistic increase in apoptosis. The objective of our study was to identify the mode of cell killing and the mechanism behind the inc...

  18. Apigenin inhibits proliferation and induces apoptosis in human multiple myeloma cells through targeting the trinity of CK2, Cdc37 and Hsp90

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    Xu Yuan-Ji

    2011-08-01

    Full Text Available Abstract Background Multiple myeloma (MM is a B-cell malignancy that is largely incurable and is characterized by the accumulation of malignant plasma cells in the bone marrow. Apigenin, a common flavonoid, has been reported to suppress proliferation in a wide variety of solid tumors and hematological cancers; however its mechanism is not well understood and its effect on MM cells has not been determined. Results In this study, we investigated the effects of apigenin on MM cell lines and on primary MM cells. Cell viability assays demonstrated that apigenin exhibited cytotoxicity against both MM cell lines and primary MM cells but not against normal peripheral blood mononuclear cells. Together, kinase assays, immunoprecipitation and western blot analysis showed that apigenin inhibited CK2 kinase activity, decreased phosphorylation of Cdc37, disassociated the Hsp90/Cdc37/client complex and induced the degradation of multiple kinase clients, including RIP1, Src, Raf-1, Cdk4 and AKT. By depleting these kinases, apigenin suppressed both constitutive and inducible activation of STAT3, ERK, AKT and NF-κB. The treatment also downregulated the expression of the antiapoptotic proteins Mcl-1, Bcl-2, Bcl-xL, XIAP and Survivin, which ultimately induced apoptosis in MM cells. In addition, apigenin had a greater effects in depleting Hsp90 clients when used in combination with the Hsp90 inhibitor geldanamycin and the histone deacetylase inhibitor vorinostat. Conclusions Our results suggest that the primary mechanisms by which apigenin kill MM cells is by targeting the trinity of CK2-Cdc37-Hsp90, and this observation reveals the therapeutic potential of apigenin in treating multiple myeloma.

  19. Tuning Hsf1 levels drives distinct fungal morphogenetic programs with depletion impairing Hsp90 function and overexpression expanding the target space

    Science.gov (United States)

    Miao, Zhengqiang; Tan, Kaeling; Vyas, Valmik K.; Whiteway, Malcolm; Robbins, Nicole; Wong, Koon Ho; Cowen, Leah E.

    2018-01-01

    The capacity to respond to temperature fluctuations is critical for microorganisms to survive within mammalian hosts, and temperature modulates virulence traits of diverse pathogens. One key temperature-dependent virulence trait of the fungal pathogen Candida albicans is its ability to transition from yeast to filamentous growth, which is induced by environmental cues at host physiological temperature. A key regulator of temperature-dependent morphogenesis is the molecular chaperone Hsp90, which has complex functional relationships with the transcription factor Hsf1. Although Hsf1 controls global transcriptional remodeling in response to heat shock, its impact on morphogenesis remains unknown. Here, we establish an intriguing paradigm whereby overexpression or depletion of C. albicans HSF1 induces morphogenesis in the absence of external cues. HSF1 depletion compromises Hsp90 function, thereby driving filamentation. HSF1 overexpression does not impact Hsp90 function, but rather induces a dose-dependent expansion of Hsf1 direct targets that drives overexpression of positive regulators of filamentation, including Brg1 and Ume6, thereby bypassing the requirement for elevated temperature during morphogenesis. This work provides new insight into Hsf1-mediated environmentally contingent transcriptional control, implicates Hsf1 in regulation of a key virulence trait, and highlights fascinating biology whereby either overexpression or depletion of a single cellular regulator induces a profound developmental transition. PMID:29590106

  20. Tuning Hsf1 levels drives distinct fungal morphogenetic programs with depletion impairing Hsp90 function and overexpression expanding the target space.

    Directory of Open Access Journals (Sweden)

    Amanda O Veri

    2018-03-01

    Full Text Available The capacity to respond to temperature fluctuations is critical for microorganisms to survive within mammalian hosts, and temperature modulates virulence traits of diverse pathogens. One key temperature-dependent virulence trait of the fungal pathogen Candida albicans is its ability to transition from yeast to filamentous growth, which is induced by environmental cues at host physiological temperature. A key regulator of temperature-dependent morphogenesis is the molecular chaperone Hsp90, which has complex functional relationships with the transcription factor Hsf1. Although Hsf1 controls global transcriptional remodeling in response to heat shock, its impact on morphogenesis remains unknown. Here, we establish an intriguing paradigm whereby overexpression or depletion of C. albicans HSF1 induces morphogenesis in the absence of external cues. HSF1 depletion compromises Hsp90 function, thereby driving filamentation. HSF1 overexpression does not impact Hsp90 function, but rather induces a dose-dependent expansion of Hsf1 direct targets that drives overexpression of positive regulators of filamentation, including Brg1 and Ume6, thereby bypassing the requirement for elevated temperature during morphogenesis. This work provides new insight into Hsf1-mediated environmentally contingent transcriptional control, implicates Hsf1 in regulation of a key virulence trait, and highlights fascinating biology whereby either overexpression or depletion of a single cellular regulator induces a profound developmental transition.

  1. Tuning Hsf1 levels drives distinct fungal morphogenetic programs with depletion impairing Hsp90 function and overexpression expanding the target space.

    Science.gov (United States)

    Veri, Amanda O; Miao, Zhengqiang; Shapiro, Rebecca S; Tebbji, Faiza; O'Meara, Teresa R; Kim, Sang Hu; Colazo, Juan; Tan, Kaeling; Vyas, Valmik K; Whiteway, Malcolm; Robbins, Nicole; Wong, Koon Ho; Cowen, Leah E

    2018-03-01

    The capacity to respond to temperature fluctuations is critical for microorganisms to survive within mammalian hosts, and temperature modulates virulence traits of diverse pathogens. One key temperature-dependent virulence trait of the fungal pathogen Candida albicans is its ability to transition from yeast to filamentous growth, which is induced by environmental cues at host physiological temperature. A key regulator of temperature-dependent morphogenesis is the molecular chaperone Hsp90, which has complex functional relationships with the transcription factor Hsf1. Although Hsf1 controls global transcriptional remodeling in response to heat shock, its impact on morphogenesis remains unknown. Here, we establish an intriguing paradigm whereby overexpression or depletion of C. albicans HSF1 induces morphogenesis in the absence of external cues. HSF1 depletion compromises Hsp90 function, thereby driving filamentation. HSF1 overexpression does not impact Hsp90 function, but rather induces a dose-dependent expansion of Hsf1 direct targets that drives overexpression of positive regulators of filamentation, including Brg1 and Ume6, thereby bypassing the requirement for elevated temperature during morphogenesis. This work provides new insight into Hsf1-mediated environmentally contingent transcriptional control, implicates Hsf1 in regulation of a key virulence trait, and highlights fascinating biology whereby either overexpression or depletion of a single cellular regulator induces a profound developmental transition.

  2. A CNS-permeable Hsp90 inhibitor rescues synaptic dysfunction and memory loss in APP-overexpressing Alzheimer's mouse model via an HSF1-mediated mechanism.

    Science.gov (United States)

    Wang, B; Liu, Y; Huang, L; Chen, J; Li, J J; Wang, R; Kim, E; Chen, Y; Justicia, C; Sakata, K; Chen, H; Planas, A; Ostrom, R S; Li, W; Yang, G; McDonald, M P; Chen, R; Heck, D H; Liao, F-F

    2017-07-01

    Induction of neuroprotective heat-shock proteins via pharmacological Hsp90 inhibitors is currently being investigated as a potential treatment for neurodegenerative diseases. Two major hurdles for therapeutic use of Hsp90 inhibitors are systemic toxicity and limited central nervous system permeability. We demonstrate here that chronic treatment with a proprietary Hsp90 inhibitor compound (OS47720) not only elicits a heat-shock-like response but also offers synaptic protection in symptomatic Tg2576 mice, a model of Alzheimer's disease, without noticeable systemic toxicity. Despite a short half-life of OS47720 in mouse brain, a single intraperitoneal injection induces rapid and long-lasting (>3 days) nuclear activation of the heat-shock factor, HSF1. Mechanistic study indicates that the remedial effects of OS47720 depend upon HSF1 activation and the subsequent HSF1-mediated transcriptional events on synaptic genes. Taken together, this work reveals a novel role of HSF1 in synaptic function and memory, which likely occurs through modulation of the synaptic transcriptome.

  3. Application of SGT1-Hsp90 chaperone complex for soluble expression of NOD1 LRR domain in E. coli

    International Nuclear Information System (INIS)

    Hong, Tae-Joon; Hahn, Ji-Sook

    2016-01-01

    NOD1 is an intracellular sensor of innate immunity which is related to a number of inflammatory diseases. NOD1 is known to be difficult to express and purify for structural and biochemical studies. Based on the fact that Hsp90 and its cochaperone SGT1 are necessary for the stabilization and activation of NOD1 in mammals, SGT1 was chosen as a fusion partner of the leucine-rich repeat (LRR) domain of NOD1 for its soluble expression in Escherichia coli. Fusion of human SGT1 (hSGT1) to NOD1 LRR significantly enhanced the solubility, and the fusion protein was stabilized by coexpression of mouse Hsp90α. The expression level of hSGT1-NOD1 LRR was further enhanced by supplementation of rare codon tRNAs and exchange of antibiotic marker genes. - Highlights: • The NOD1 LRR domain was solubilized by SGT1 fusion in E. coli. • The coexpression of HSP90 stabilized the SGT1-NOD1 LRR fusion protein. • Several optimizations could enhance the expression level of the fusion protein.

  4. Calcium/calmodulin kinase1 and its relation to thermotolerance and HSP90 in Sporothrix schenckii: an RNAi and yeast two-hybrid study

    Directory of Open Access Journals (Sweden)

    Gonzalez-Mendez Ricardo

    2011-07-01

    Full Text Available Abstract Background Sporothrix schenckii is a pathogenic dimorphic fungus of worldwide distribution. It grows in the saprophytic form with hyaline, regularly septated hyphae and pyriform conidia at 25°C and as the yeast or parasitic form at 35°C. Previously, we characterized a calcium/calmodulin kinase in this fungus. Inhibitors of this kinase were observed to inhibit the yeast cell cycle in S. schenckii. Results The presence of RNA interference (RNAi mechanism in this fungus was confirmed by the identification of a Dicer-1 homologue in S. schenckii DNA. RNAi technology was used to corroborate the role of calcium/calmodulin kinase I in S. schenckii dimorphism. Yeast cells were transformed with the pSilent-Dual2G (pSD2G plasmid w/wo inserts of the coding region of the calcium/calmodulin kinase I (sscmk1 gene. Transformants were selected at 35°C using resistance to geneticin. Following transfer to liquid medium at 35°C, RNAi transformants developed as abnormal mycelium clumps and not as yeast cells as would be expected. The level of sscmk1 gene expression in RNAi transformants at 35°C was less than that of cells transformed with the empty pSD2G at this same temperature. Yeast two-hybrid analysis of proteins that interact with SSCMK1 identified a homologue of heat shock protein 90 (HSP90 as interacting with this kinase. Growth of the fungus similar to that of the RNAi transformants was observed in medium with geldanamycin (GdA, 10 μM, an inhibitor of HSP90. Conclusions Using the RNAi technology we silenced the expression of sscmk1 gene in this fungus. RNAi transformants were unable to grow as yeast cells at 35°C showing decreased tolerance to this temperature. The interaction of SSCMK1 with HSP90, observed using the yeast two-hybrid assay suggests that this kinase is involved in thermotolerance through its interaction with HSP90. SSCMK1 interacted with the C terminal domain of HSP90 where effector proteins and co-chaperones interact. These

  5. Molecular imaging of EGFR and CD44v6 for prediction and response monitoring of HSP90 inhibition in an in vivo squamous cell carcinoma model

    Energy Technology Data Exchange (ETDEWEB)

    Spiegelberg, Diana; Mortensen, Anja C.; Stenerloew, Bo [Uppsala University, Department of Immunology, Genetics and Pathology, Uppsala (Sweden); Selvaraju, Ram K.; Eriksson, Olof [Uppsala University, Preclinical PET Platform, Uppsala (Sweden); Nestor, Marika [Uppsala University, Department of Immunology, Genetics and Pathology, Uppsala (Sweden); Uppsala University, Unit of Otolaryngology and Head and Neck Surgery, Department of Surgical Sciences, Uppsala (Sweden)

    2016-05-15

    Heat shock protein 90 (HSP90) is essential for the activation and stabilization of numerous oncogenic client proteins. AT13387 is a novel HSP90 inhibitor promoting degradation of oncogenic proteins upon binding, and may also act as a radiosensitizer. For optimal treatment there is, however, the need for identification of biomarkers for patient stratification and therapeutic response monitoring, and to find suitable targets for combination treatments. The aim of this study was to assess the response of surface antigens commonly expressed in squamous cell carcinoma to AT13387 treatment, and to find suitable biomarkers for molecular imaging and radioimmunotherapy in combination with HSP90 inhibition. Cancer cell proliferation and radioimmunoassays were used to evaluate the effect of AT13387 on target antigen expression in vitro. Inhibitor effects were then assessed in vivo in mice-xenografts. Animals were treated with AT13387 (5 x 50 mg/kg), and were imaged with PET using either {sup 18}F-FDG or {sup 124}I-labelled tracers for EGFR and CD44v6, and this was followed by ex-vivo biodistribution analysis and immunohistochemical staining. AT13387 exposure resulted in high cytotoxicity and possible radiosensitization with IC{sub 50} values below 4 nM. Both in vitro and in vivo AT13387 effectively downregulated HSP90 client proteins. PET imaging with {sup 124}I-cetuximab showed a significant decrease of EGFR in AT13387-treated animals compared with untreated animals. In contrast, the squamous cell carcinoma-associated biomarker CD44v6, visualized with {sup 124}I-AbD19384 as well as {sup 18}F-FDG uptake, were not significantly altered by AT13387 treatment. We conclude that AT13387 downregulates HSP90 client proteins, and that molecular imaging of these proteins may be a suitable approach for assessing treatment response. Furthermore, radioimmunotherapy targeting CD44v6 in combination with AT13387 may potentiate the radioimmunotherapy outcome due to radiosensitizing effects of

  6. 17-AAG mediated targeting of Hsp90 limits tert activity in peritoneal sarcoma related malignant ascites by downregulating cyclin D1 during cell cycle entry.

    Science.gov (United States)

    Chaklader, M; Das, P; Pereira, J A; Law, A; Chattopadhyay, S; Chatterjee, R; Mondal, A; Law, S

    2012-07-01

    Peritoneal or retro-peritoneal sarcomatosis related malignant ascites formation is a rare but serious consequence of the locoregional metastatic event. The present work aimed to study the effect of the Hsp90 inhibitor (17-AAG), an ansamycin analog, on cell cycle and DNA replication specific chaperone-clients interaction in the event of peritoneal sarcoma related malignant ascites formation in mouse model at the late stage of malignant growth. We administered 17-AAG, an Hsp90 inhibitor, divided doses (330 μg/kg b.w./day for first five days then next ten days with166 μg/kg b.w./day) through intra-peritoneal route of inbred Swiss albino mice bearing full grown peritoneal malignant ascites of sarcoma-180. Our study was evaluated by peripheral blood hemogram analysis, malignant ascitic cytology, cell viability test, survival time and mitotic indexing. Furthermore, flowcytometric HSP90, TERT, CyclinD1, PCNA and GM-CSF expression analysis has been considered for special objective of the study. Our experimental efforts reduced the aggressive proliferation of malignant ascites by drastic downregulation of TERT and cyclin D1 on the verge of cell cycle entry along with DNA replication processivity factor PCNA by directly modulating their folding machinery - heat shock protein 90. Consequently, we observed that malignant ascitic cells became error prone during the event of karyokinesis and produced micronucleus containing malignant cells with low viability. Peripheral neutrophilia due to over-expression of GM-CSF by the peritoneal malignant ascites were also controlled by the treatment with 17-AAG and overall, the treatment modality improved the median survival time. Finally we can conclude that 17AAG administration might serve as a prospective pharmacological agent for the management of peritoneal sarcoma related malignant ascites and throws light towards prolonged survival of the patients concerned.

  7. The Transient Receptor Potential Melastatin 7 Channel Regulates Pancreatic Cancer Cell Invasion through the Hsp90α/uPA/MMP2 pathway

    Directory of Open Access Journals (Sweden)

    Pierre Rybarczyk

    2017-04-01

    Full Text Available Pancreatic ductal adenocarcinoma (PDAC is an aggressive malignancy with a very poor prognosis. There is an urgent need to better understand the molecular mechanisms that regulate PDAC cell aggressiveness. The transient receptor potential melastatin 7 (TRPM7 is a nonselective cationic channel that mainly conducts Ca2+ and Mg2+. TRPM7 is overexpressed in numerous malignancies including PDAC. In the present study, we used the PANC-1 and MIA PaCa-2 cell lines to specifically assess the role of TRPM7 in cell invasion and matrix metalloproteinase secretion. We show that TRPM7 regulates Mg2+ homeostasis and constitutive cation entry in both PDAC cell lines. Moreover, cell invasion is strongly reduced by TRPM7 silencing without affecting the cell viability. Conditioned media were further studied, by gel zymography, to detect matrix metalloproteinase (MMP secretion in PDAC cells. Our results show that MMP-2, urokinase plasminogen activator (uPA, and heat-shock protein 90α (Hsp90α secretions are significantly decreased in TRPM7-deficient PDAC cells. Moreover, TRPM7 expression in human PDAC lymph node metastasis is correlated to the channel expression in primary tumor. Taken together, our results show that TRPM7 is involved in PDAC cell invasion through regulation of Hsp90α/uPA/MMP-2 proteolytic axis, confirming that this channel could be a promising biomarker and possibly a target for PDAC metastasis therapy.

  8. Fluorescent ligand fishing combination with in-situ imaging and characterizing to screen Hsp 90 inhibitors from Curcuma longa L. based on InP/ZnS quantum dots embedded mesoporous nanoparticles.

    Science.gov (United States)

    Hu, Yue; Fu, Anchen; Miao, Zhaoyi; Zhang, Xiaojing; Wang, Tianlin; Kang, An; Shan, Jinjun; Zhu, Dong; Li, Wei

    2018-02-01

    Although ligand fishing has been shown to be an efficient technique for the identification of bioactive components from complex mixtures such as natural products, it cannot be applied to biomedical image processing. Herein, a specific fluorescent ligand fishing combined with in situ imaging approach is presented for the identification of heat shock protein 90 (Hsp 90) inhibitors from complex matrixes, Curcuma longa L., using N-terminus immobilized Hsp 90α functionalized InP/ZnS quantum dots embedded mesoporous nanoparticles (i.e. Hsp 90α (NT)-FQDNs) as extraction sorbents and fluorescent tracer. The fished ligands were identified by liquid chromatography time-of-flight/mass spectrometry (LC-TOF/MS) and gas chromatography-mass spectrometry (GC-MS). Moreover, in situ imaging by confocal laser scanning microscopy (CLSM) was applied for evaluating the effect of fished-ligands on bioactivity-induced apoptosis morphologically in HeLa cells. MTT assay verified the bioactivity of the ligands and molecular docking results further provided convincing information to verify the feasible binding mode between ligands and protein. Twelve ligands as potential Hsp 90 inhibitors were ultimately fished and identified from Curcuma longa L. crude extracts. The proposed approach based on Hsp 90α functionalized nanocomposites is superior in the combination of highly specific screening efficiency and concurrent visual in situ imaging, which could have great promise for the development of other plant-derived Hsp 90 inhibitors, and providing a rapid and reliable platform for discovering biologically active molecules in natural products. Copyright © 2017 Elsevier B.V. All rights reserved.

  9. Oral administration of an HSP90 inhibitor, 17-DMAG, intervenes tumor-cell infiltration into multiple organs and improves survival period for ATL model mice

    International Nuclear Information System (INIS)

    Ikebe, E; Kawaguchi, A; Tezuka, K; Taguchi, S; Hirose, S; Matsumoto, T; Mitsui, T; Senba, K; Nishizono, A; Hori, M; Hasegawa, H; Yamada, Y; Ueno, T; Tanaka, Y; Sawa, H; Hall, W; Minami, Y; Jeang, K T; Ogata, M; Morishita, K; Hasegawa, H; Fujisawa, J; Iha, H

    2013-01-01

    In the peripheral blood leukocytes (PBLs) from the carriers of the human T-lymphotropic virus type-1 (HTLV-1) or the patients with adult T-cell leukemia (ATL), nuclear factor kappaB (NF-κB)-mediated antiapoptotic signals are constitutively activated primarily by the HTLV-1-encoded oncoprotein Tax. Tax interacts with the I κB kinase regulatory subunit NEMO (NF-κB essential modulator) to activate NF-κB, and this interaction is maintained in part by a molecular chaperone, heat-shock protein 90 (HSP90), and its co-chaperone cell division cycle 37 (CDC37). The antibiotic geldanamycin (GA) inhibits HSP90's ATP binding for its proper interaction with client proteins. Administration of a novel water-soluble and less toxic GA derivative, 17-dimethylaminoethylamino-17-demethoxygeldanamycin hydrochloride (17-DMAG), to Tax-expressing ATL-transformed cell lines, C8166 and MT4, induced significant degradation of Tax. 17-DMAG also facilitated growth arrest and cellular apoptosis to C8166 and MT4 and other ATL cell lines, although this treatment has no apparent effects on normal PBLs. 17-DMAG also downregulated Tax-mediated intracellular signals including the activation of NF-κB, activator protein 1 or HTLV-1 long terminal repeat in Tax-transfected HEK293 cells. Oral administration of 17-DMAG to ATL model mice xenografted with lymphomatous transgenic Lck-Tax (Lck proximal promoter-driven Tax transgene) cells or HTLV-1-producing tumor cells dramatically attenuated aggressive infiltration into multiple organs, inhibited de novo viral production and improved survival period. These observations identified 17-DMAG as a promising candidate for the prevention of ATL progression

  10. International organizations as orchestrators

    CERN Document Server

    Abbott, Kenneth W

    2015-01-01

    International Organizations as Orchestrators reveals how IOs leverage their limited authority and resources to increase their effectiveness, power, and autonomy from states. By 'orchestrating' intermediaries - including NGOs - IOs can shape and steer global governance without engaging in hard, direct regulation. This volume is organized around a theoretical model that emphasizes voluntary collaboration and support. An outstanding group of scholars investigate the significance of orchestration across key issue areas, including trade, finance, environment and labor, and in leading organizations, including the GEF, G20, WTO, EU, Kimberley Process, UNEP and ILO. The empirical studies find that orchestration is pervasive. They broadly confirm the theoretical hypotheses while providing important new insights, especially that states often welcome IO orchestration as achieving governance without creating strong institutions. This volume changes our understanding of the relationships among IOs, nonstate actors and sta...

  11. The targeted inhibition of mitochondrial Hsp90 overcomes the apoptosis resistance conferred by Bcl-2 in Hep3B cells via necroptosis

    Energy Technology Data Exchange (ETDEWEB)

    Yan, Chunlan [Department of Anatomy and Cell Biology, Dong-A University College of Medicine and Mitochondria Hub Regulation Center, Busan, 602-714 (Korea, Republic of); Department of Physiology, Zhejiang University School of Medicine, Hangzhou, Zhejiang 310058 (China); Oh, Joon Seok; Yoo, Seung Hee; Lee, Jee Suk [Department of Anatomy and Cell Biology, Dong-A University College of Medicine and Mitochondria Hub Regulation Center, Busan, 602-714 (Korea, Republic of); Yoon, Young Geol [Department of Anatomy and Cell Biology, Dong-A University College of Medicine and Mitochondria Hub Regulation Center, Busan, 602-714 (Korea, Republic of); Department of Biomedical Science, Institute for Biomedical and Health Sciences, Jungwon University, Chungbuk, 367-805 (Korea, Republic of); Oh, Yoo Jin; Jang, Min Seok [Department of Anatomy and Cell Biology, Dong-A University College of Medicine and Mitochondria Hub Regulation Center, Busan, 602-714 (Korea, Republic of); Lee, Sang Yeob [Department of Rheumatology, Dong-A University College of Medicine, Busan, 602-714 (Korea, Republic of); Yang, Jun [Department of Toxicology, Hangzhou Normal University School of Public Health, Hangzhou, Zhejiang, 310036 China (China); Lee, Sang Hwa [Department of Microbiology and, Dong-A University College of Medicine, Busan, 602-714 (Korea, Republic of); Kim, Hye Young [Department of Anatomy and Cell Biology, Dong-A University College of Medicine and Mitochondria Hub Regulation Center, Busan, 602-714 (Korea, Republic of); Yoo, Young Hyun, E-mail: yhyoo@dau.ac.kr [Department of Anatomy and Cell Biology, Dong-A University College of Medicine and Mitochondria Hub Regulation Center, Busan, 602-714 (Korea, Republic of)

    2013-01-01

    Previous studies have reported that a Gamitrinib variant containing triphenylphosphonium (G-TPP) binds to mitochondrial Hsp90 and rapidly inhibits its activity, thus inducing the apoptotic pathway in the cells. Accordingly, G-TPP shows a potential as a promising drug for the treatment of cancer. A cell can die from different types of cell death such as apoptosis, necrosis, necroptosis, and autophagic cell death. In this study, we further investigated the mechanisms and modes of cell death in the G-TPP-treated Hep3B and U937 cell lines. We discovered that G-TPP kills the U937 cells through the apoptotic pathway and the overexpression of Bcl-2 significantly inhibits U937 cell death to G-TPP. We further discovered that G-TPP kills the Hep3B cells by activating necroptosis in combination with the partial activation of caspase-dependent apoptosis. Importantly, G-TPP overcomes the apoptosis resistance conferred by Bcl-2 in Hep3B cells via necroptosis. We also observed that G-TPP induces compensatory autophagy in the Hep3B cell line. We further found that whereas there is a Bcl-2-Beclin 1 interaction in response to G-TPP, silencing the beclin 1 gene failed to block LC3-II accumulation in the Hep3B cells, indicating that G-TPP triggers Beclin 1-independent protective autophagy in Hep3B cells. Taken together, these data reveal that G-TPP induces cell death through a combination of death pathways, including necroptosis and apoptosis, and overcomes the apoptosis resistance conferred by Bcl-2 in Hep3B cells via necroptosis. These findings are important for the therapeutic exploitation of necroptosis as an alternative cell death program to bypass the resistance to apoptosis. Highlights: ► G-TPP binds to mitochondrial Hsp90. ► G-TPP induces apoptosis in U937 human leukemia cancer cells. ► G-TPP induces combination of death pathways in Hep3B cell. ► G-TPP overcomes the resistance conferred by Bcl-2 in Hep3B cells via necroptosis. ► G-TPP triggers Beclin 1-independent

  12. 17-AAG, a Hsp90 inhibitor, attenuates the hypoxia-induced expression of SDF-1alpha and ILK in mouse RPE cells.

    Science.gov (United States)

    Wang, Ye Qing; Zhang, Xiao Mei; Wang, Xiao Dan; Wang, Bin Jie; Wang, Wei

    2010-03-01

    The aim of this study was to investigate the changes of SDF-1alpha and ILK expression in mouse retinal pigment epithelium (RPE) cells in response to hypoxia, and the effect of 17-Allylamino-17-demethoxygeldanamycin (17-AAG), a heat shock protein 90 (Hsp90) inhibitor, on the hypoxia-induced expression of SDF-1alpha and ILK. RPE cells were cultured with 200 micromol/L cobalt chloride (CoCl(2)) for different times (1, 3, 6, 12, 24, 72 h) to imitate chemical hypoxia. Pretreatment of 17-AAG was 1 h prior to hypoxic insult. Cellular viability after 17-AAG treatment was assessed by MTT assay, and the changes of SDF-1alpha and ILK expression were examined by RT-PCR and Western blot. Up-regulation of SDF-1alpha and ILK expression in response to hypoxia was observed. One hour pretreatment of 17-AAG could remarkably decreased the hypoxia-induced SDF-1alpha and ILK expression in vitro. Our results indicated that SDF-1alpha and ILK involved in the hypoxic response of RPE cells, and 1 h pretreatment of 17-AAG had an inhibitive effect on the hypoxia-induced SDF-1alpha and ILK expression.

  13. Targeting Hsp90 by 17-AAG in leukemia cells: mechanisms for synergistic and antagonistic drug combinations with arsenic trioxide and Ara-C.

    Science.gov (United States)

    Pelicano, H; Carew, J S; McQueen, T J; Andreeff, M; Plunkett, W; Keating, M J; Huang, P

    2006-04-01

    17-Allylamino-17-demethoxygeldanamycin (17-AAG) is a new anticancer agent currently in clinical trials. The ability of 17-AAG to abrogate the function of heat-shock protein Hsp90 and modulate cellular sensitivity to anticancer agents has prompted recent research to use this compound in drug combination therapy. Here we report that 17-AAG has striking opposite effects on the activity of arsenic trioxide (ATO) and ara-C. Combination of 17-AAG with ATO exhibited a synergistic effect in leukemia cells, whereas coincubation of 17-AAG and ara-C showed antagonistic activity. Mechanistic studies revealed that ATO exerted cytotoxic action by reactive oxygen species generation, and activated Akt survival pathway. 17-AAG abrogated Akt activation and enhanced the activity of ATO. In contrast, treatment of leukemia cells with 17-AAG caused a G1 arrest, a decrease in DNA synthesis and reduced ara-C incorporation into DNA, leading to antagonism. The ability of 17-AAG to enhance the antileukemia activity of ATO was further demonstrated in primary leukemia cells isolated from patients with acute myeloid leukemia and chronic lymphocytic leukemia, including cells from refractory patients. Our data suggest that combination of 17-AAG and ATO may be an effective therapeutic regimen. Caution should be exercised in using 17-AAG together with ara-C, as their combination effects are schedule dependent.

  14. Induction of cell cycle arrest and apoptosis with downregulation of Hsp90 client proteins and histone modification by 4β-hydroxywithanolide E isolated from Physalis peruviana.

    Science.gov (United States)

    Park, Eun-Jung; Sang-Ngern, Mayuramas; Chang, Leng Chee; Pezzuto, John M

    2016-06-01

    Physalis peruviana (Solanaceae) is used for culinary and medicinal purposes. We currently report withanolides, isolated from P. peruviana, inhibit the growth of colon cancer monolayer and spheroid cultures. A detailed mechanistic evaluation was performed with 4β-hydroxywithanolide E (4HWE). Treatment of HT-29 cells with low concentrations of 4HWE inhibited growth while enhancing levels of p21(Waf1/Cip1) and reducing levels of several cell cycle-related proteins. Apoptosis was induced at higher concentrations. In addition, 4HWE treatment downregulated the levels of Hsp90 client proteins. Nuclear sirtuin 1 (SIRT1) was increased and histone H3 acetylated at lysine 9 was decreased. An additional consequence of SIRT1 elevation in the nucleus may be inhibition of c-Jun activity. The expression of 21 genes was altered, including downregulation of PTGS2, and this correlated with reduced protein levels of cyclooxygenase-2 (COX-2). Overall, efficacious induction of G0/G1 cell cycle arrest at low concentrations, and induction of apoptosis at higher concentrations are interesting 4HWE-mediated phenomena that are accompanied by a complex array of molecular events. Considering the worldwide prevalence of colon cancer, and the unique mode of action mediated by 4HWE, it is reasonable to investigate additional mechanistic details and the potential utility of this compound. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. Inflammatory stress of pancreatic beta cells drives release of extracellular heat-shock protein 90α.

    Science.gov (United States)

    Ocaña, Gail J; Pérez, Liliana; Guindon, Lynette; Deffit, Sarah N; Evans-Molina, Carmella; Thurmond, Debbie C; Blum, Janice S

    2017-06-01

    A major obstacle in predicting and preventing the development of autoimmune type 1 diabetes (T1D) in at-risk individuals is the lack of well-established early biomarkers indicative of ongoing beta cell stress during the pre-clinical phase of disease. Recently, serum levels of the α cytoplasmic isoform of heat-shock protein 90 (hsp90) were shown to be elevated in individuals with new-onset T1D. We therefore hypothesized that hsp90α could be released from beta cells in response to cellular stress and inflammation associated with the earliest stages of T1D. Here, human beta cell lines and cadaveric islets released hsp90α in response to stress induced by treatment with a combination of pro-inflammatory cytokines including interleukin-1β, tumour necrosis factor-α and interferon-γ. Mechanistically, hsp90α release was found to be driven by cytokine-induced endoplasmic reticulum stress mediated by c-Jun N-terminal kinase (JNK), a pathway that can eventually lead to beta cell apoptosis. Cytokine-induced beta cell hsp90α release and JNK activation were significantly reduced by pre-treating cells with the endoplasmic reticulum stress-mitigating chemical chaperone tauroursodeoxycholic acid. The hsp90α release by cells may therefore be a sensitive indicator of stress during inflammation and a useful tool in assessing therapeutic mitigation of cytokine-induced cell damage linked to autoimmunity. © 2017 John Wiley & Sons Ltd.

  16. Ganetespib, an HSP90 inhibitor, kills Epstein-Barr virus (EBV)-infected B and T cells and reduces the percentage of EBV-infected cells in the blood.

    Science.gov (United States)

    Shatzer, Amber; Ali, Mir A; Chavez, Mayra; Dowdell, Kennichi; Lee, Min-Jung; Tomita, Yusuke; El-Hariry, Iman; Trepel, Jane B; Proia, David A; Cohen, Jeffrey I

    2017-04-01

    HSP90 inhibitors have been shown to kill Epstein-Barr virus (EBV)-infected cells by reducing the level of EBV EBNA-1 and/or LMP1. We treated virus-infected cells with ganetespib, an HSP90 inhibitor currently being evaluated in multiple clinical trials for cancer and found that the drug killed EBV-positive B and T cells and reduced the level of both EBV EBNA-1 and LMP1. Treatment of cells with ganetespib also reduced the level of pAkt. Ganetespib delayed the onset of EBV-positive lymphomas and prolonged survival in SCID mice inoculated with one EBV-transformed B-cell line, but not another B-cell line. The former cell line showed lower levels of EBNA-1 after treatment with ganetespib in vitro. Treatment of a patient with T-cell chronic active EBV with ganetespib reduced the percentage of EBV-positive cells in the peripheral blood. These data indicate that HSP90 inhibitors may have a role in the therapy of certain EBV-associated diseases.

  17. Orchestrating Lean Implementation

    DEFF Research Database (Denmark)

    Riis, Jens Ove; Mikkelsen, Hans; Andersen, Jesper Rank

    2008-01-01

    The notion of Lean Manufacturing is not merely confined to a set of well defined techniques, but represents a broad approach to managing a company. Working with lean entails many aspects, such as production planning and control, production engineering, product development, supply chain......, and organizational issues. To become effective, many functional areas and departments must be involved. At the same time companies are embedded in a dynamic environment. The aim of the paper is to propose a comprehensive approach to better implementation of lean initiatives, based on two empirical studies. The paper...... will discuss how a concerted effort can be staged taking into account the interdependencies among individual improvement initiatives. The notion of orchestration will be introduced, and several means for orchestration will be presented. Critical behavioral issues for lean implementation will be discussed....

  18. Hsp90 inhibitor 17-AAG reduces ErbB2 levels and inhibits proliferation of the trastuzumab resistant breast tumor cell line JIMT-1.

    Science.gov (United States)

    Zsebik, Barbara; Citri, Ami; Isola, Jorma; Yarden, Yosef; Szöllosi, János; Vereb, György

    2006-04-15

    ErbB2, a member of the EGF receptor family of tyrosine kinases is overexpressed on many tumor cells of epithelial origin and is the molecular target of trastuzumab (Herceptin), the first humanized antibody used in the therapy of solid tumors. Trastuzumab, which is thought to act, at least in part, by downregulating ErbB2 expression is only effective in approximately 30-40% of ErbB2 positive breast tumors. Geldanamycin and its derivative 17-AAG are potential antitumor agents capable of downregulating client proteins of Hsp90, including ErbB2. To investigate the ability of 17-AAG to downregulate ErbB2 in trastuzumab resistant breast cancer cells and the possibility of 17-AAG and trastuzumab potentiating each other's effect, the recently established trastuzumab resistant breast cancer cell line, JIMT-1 was compared to the known trastuzumab sensitive SKBR-3 line. Baseline and stimulus-evoked dimerization and activation levels of ErbB2, and the effects of trastuzumab and 17-AAG alone and in combination on cell proliferation and apoptosis, as well as on ErbB2 expression and phosphorylation have been measured. Baseline activation and amenability to activation and downregulation by trastuzumab was much lower in the resistant line. However, 17-AAG enhanced ErbB2 homodimerization after 5-10 min of treatment in both cell lines, and decreased proliferation with an IC50 of 70 nM for SKBR-3 and 10nM for JIMT-1. Thus, 17-AAG may be a useful drug in trastuzumab resistant ErbB2 overexpressing tumors. The antiproliferative effect of 17-AAG was positively correlated with phosphorylation and downregulation of ErbB2 and was dominated by apoptosis, although, especially at higher doses, necrosis was also present. Interestingly, IC50 values for ErbB2 downregulation and phosphorylation, in the 30-40 nM range, were not significantly different for the two cell lines. This observation and the negative correlation between resting ErbB2 levels and the antiproliferative effect of 17-AAG may

  19. Downregulation of androgen receptors by NaAsO2 via inhibition of AKT-NF-κB and HSP90 in castration resistant prostate cancer.

    Science.gov (United States)

    Kim, Yunlim; Park, Sang Eun; Moon, Jeong-Weon; Kim, Bong-Min; Kim, Ha-Gyeong; Jeong, In Gab; Yoo, Sangjun; Ahn, Jae Beom; You, Dalsan; Pak, Jhang Ho; Kim, Sujong; Hwang, Jung Jin; Kim, Choung-Soo

    2017-07-01

    Androgen and androgen receptor (AR) play essential roles in the development and maintenance of prostate cancer. The recently identified AR splice variants (AR-Vs) have been considered as a plausible mechanism for the primary resistance against androgen deprivation therapy (ADT) in castration-resistant prostate cancer (CRPC). Sodium meta-arsenite (NaAsO 2 ; KML001; Kominox), a trivalent arsenical, is an orally bioavailable and water soluble, which is currently in phase I/II clinical trials for the treatment of prostate cancer. It has a potent anti-cancer effect on prostate cancer cells and xenografts. The aim of this study was to examine the effect of NaAsO 2 on AR signaling in LNCaP and 22Rv1 CRPC cells. We used hormone-sensitive LNCaP cells, hormone-insensitive 22Rv1 cells, and CRPC patient-derived primary cells. We analyzed anti-cancer effect of NaAsO 2 using real-time quantitative reverse transcription-PCR, Western blotting, immunofluorescence staining and CellTiter Glo® luminescent assay. Statistical evaluation of the results was performed by one-way ANOVA. NaAsO 2 significantly reduced the translocation of AR and AR-Vs to the nucleus as well as their level in LNCaP and 22Rv1 cells. Besides, the level of the prostate-specific antigen (PSA), downstream target gene of AR, was also decreased. This compound was also an effective modulator of AKT-dependent NF-κB activation which regulates AR. NaAsO 2 significantly inhibited phosphorylation of AKT and expression and nuclear translocation of NF-κB. We then investigated the effect of NaAsO 2 on AR stabilization. NaAsO 2 promoted HSP90 acetylation by down-regulating HDAC6, which reduces the stability of AR in prostate cancer cells. Here, we show that NaAsO 2 disrupts AR signaling at multiple levels by affecting AR expression, stability, and degradation in primary tumor cell cultures from prostate cancer patients as well as CRPC cell lines. These results suggest that NaAsO 2 could be a novel therapeutics for prostate

  20. Windows Terminal Servers Orchestration

    Science.gov (United States)

    Bukowiec, Sebastian; Gaspar, Ricardo; Smith, Tim

    2017-10-01

    Windows Terminal Servers provide application gateways for various parts of the CERN accelerator complex, used by hundreds of CERN users every day. The combination of new tools such as Puppet, HAProxy and Microsoft System Center suite enable automation of provisioning workflows to provide a terminal server infrastructure that can scale up and down in an automated manner. The orchestration does not only reduce the time and effort necessary to deploy new instances, but also facilitates operations such as patching, analysis and recreation of compromised nodes as well as catering for workload peaks.

  1. Orchestrating intensities and rhythms

    DEFF Research Database (Denmark)

    Staunæs, Dorthe; Juelskjær, Malou

    2016-01-01

    environmentality and learning-centered governance standards has dramatic and performative effects for the production of (educational) subjectivities. This implies a shift from governing identities, categories and structures towards orchestrating affective intensities and rhythms. Finally, the article discusses...... and the making of subjects have held sway for many years; and it is also well known that schools have been some of the most regular purchasers of psychological methods, tests and classifications. Following but also elaborating upon governmentality studies, it is suggested that a current shift towards...

  2. MicroRNA-27b plays a role in pulmonary arterial hypertension by modulating peroxisome proliferator-activated receptor γ dependent Hsp90-eNOS signaling and nitric oxide production

    Energy Technology Data Exchange (ETDEWEB)

    Bi, Rui; Bao, Chunrong; Jiang, Lianyong; Liu, Hao; Yang, Yang; Mei, Ju; Ding, Fangbao, E-mail: dbcar126@126.com

    2015-05-01

    Pulmonary artery endothelial dysfunction is associated with pulmonary arterial hypertension (PAH). Based on recent studies showing that microRNA (miR)-27b is aberrantly expressed in PAH, we hypothesized that miR-27b may contribute to pulmonary endothelial dysfunction and vascular remodeling in PAH. The effect of miR-27b on pulmonary endothelial dysfunction and the underlying mechanism were investigated in human pulmonary artery endothelial cells (HPAECs) in vitro and in a monocrotaline (MCT)-induced model of PAH in vivo. miR-27b expression was upregulated in MCT-induced PAH and inversely correlated with the levels of peroxisome proliferator-activated receptor (PPAR)-γ, and miR-27b inhibition attenuated MCT-induced endothelial dysfunction and remodeling and prevented PAH associated right ventricular hypertrophy and systolic pressure in rats. PPARγ was confirmed as a direct target of miR-27b in HPAECs and shown to mediate the effect of miR-27b on the disruption of endothelial nitric oxide synthase (eNOS) coupling to Hsp90 and the suppression of NO production associated with the PAH phenotype. We showed that miR-27b plays a role endothelial function and NO release and elucidated a potential mechanism by which miR-27b regulates Hsp90-eNOS and NO signaling by modulating PPARγ expression, providing potential therapeutic targets for the treatment of PAH. - Highlights: • miR-27b plays a role in endothelial function and NO release. • miR-27b inhibition ameliorates MCT-induced endothelial dysfunction and PAH. • miR-27b targets PPARγ in HPAECs. • miR-27b regulates PPARγ dependent Hsp90-eNOS and NO signaling.

  3. Multi-faceted proteomic characterization of host protein complement of Rift Valley fever virus virions and identification of specific heat shock proteins, including HSP90, as important viral host factors.

    Science.gov (United States)

    Nuss, Jonathan E; Kehn-Hall, Kylene; Benedict, Ashwini; Costantino, Julie; Ward, Michael; Peyser, Brian D; Retterer, Cary J; Tressler, Lyal E; Wanner, Laura M; McGovern, Hugh F; Zaidi, Anum; Anthony, Scott M; Kota, Krishna P; Bavari, Sina; Hakami, Ramin M

    2014-01-01

    Rift Valley fever is a potentially fatal disease of humans and domestic animals caused by Rift Valley fever virus (RVFV). Infection with RVFV in ruminants can cause near 100% abortion rates and recent outbreaks in naïve human populations have suggested case fatality rates of greater than thirty percent. To elucidate the roles that host proteins play during RVFV infection, proteomic analysis of RVFV virions was conducted using complementary analytical approaches, followed by functional validation studies of select identified host factors. Coupling the more traditional Gel LC/MS/MS approach (SDS PAGE followed by liquid chromatography tandem mass spectrometry) with an alternative technique that preserves protein complexes allowed the protein complement of these viral particles to be thoroughly examined. In addition to viral proteins present within the virions and virion-associated host proteins, multiple macromolecular complexes were identified. Bioinformatic analysis showed that host chaperones were among over-represented protein families associated with virions, and functional experiments using siRNA gene silencing and small molecule inhibitors identified several of these heat shock proteins, including heat shock protein 90 (HSP90), as important viral host factors. Further analysis indicated that HSP inhibition effects occur during the replication/transcription phase of the virus life cycle, leading to significant lowering of viral titers without compromising the functional capacity of released virions. Overall, these studies provide much needed further insight into interactions between RVFV and host cells, increasing our understanding of the infection process and suggesting novel strategies for anti-viral development. In particular, considering that several HSP90 inhibitors have been advancing through clinical trials for cancer treatment, these results also highlight the exciting potential of repurposing HSP90 inhibitors to treat RVF.

  4. Multi-faceted proteomic characterization of host protein complement of Rift Valley fever virus virions and identification of specific heat shock proteins, including HSP90, as important viral host factors.

    Directory of Open Access Journals (Sweden)

    Jonathan E Nuss

    Full Text Available Rift Valley fever is a potentially fatal disease of humans and domestic animals caused by Rift Valley fever virus (RVFV. Infection with RVFV in ruminants can cause near 100% abortion rates and recent outbreaks in naïve human populations have suggested case fatality rates of greater than thirty percent. To elucidate the roles that host proteins play during RVFV infection, proteomic analysis of RVFV virions was conducted using complementary analytical approaches, followed by functional validation studies of select identified host factors. Coupling the more traditional Gel LC/MS/MS approach (SDS PAGE followed by liquid chromatography tandem mass spectrometry with an alternative technique that preserves protein complexes allowed the protein complement of these viral particles to be thoroughly examined. In addition to viral proteins present within the virions and virion-associated host proteins, multiple macromolecular complexes were identified. Bioinformatic analysis showed that host chaperones were among over-represented protein families associated with virions, and functional experiments using siRNA gene silencing and small molecule inhibitors identified several of these heat shock proteins, including heat shock protein 90 (HSP90, as important viral host factors. Further analysis indicated that HSP inhibition effects occur during the replication/transcription phase of the virus life cycle, leading to significant lowering of viral titers without compromising the functional capacity of released virions. Overall, these studies provide much needed further insight into interactions between RVFV and host cells, increasing our understanding of the infection process and suggesting novel strategies for anti-viral development. In particular, considering that several HSP90 inhibitors have been advancing through clinical trials for cancer treatment, these results also highlight the exciting potential of repurposing HSP90 inhibitors to treat RVF.

  5. HSP90 inhibition is effective in breast cancer: a phase II trial of tanespimycin (17-AAG) plus trastuzumab in patients with HER2-positive metastatic breast cancer progressing on trastuzumab.

    Science.gov (United States)

    Modi, Shanu; Stopeck, Alison; Linden, Hannah; Solit, David; Chandarlapaty, Sarat; Rosen, Neal; D'Andrea, Gabriella; Dickler, Maura; Moynahan, Mary E; Sugarman, Steven; Ma, Weining; Patil, Sujata; Norton, Larry; Hannah, Alison L; Hudis, Clifford

    2011-08-01

    HSP90 is a chaperone protein required for the stability of a variety of client proteins. 17-Demethoxygeldanamycin (17-AAG) is a natural product that binds to HSP90 and inhibits its activity, thereby inducing the degradation of these clients. In preclinical studies, HER2 is one of the most sensitive known client proteins of 17-AAG. On the basis of these data and activity in a phase I study, we conducted a phase II study of 17-AAG (tanespimycin) with trastuzumab in advanced trastuzumab-refractory HER2-positive breast cancer. We enrolled patients with metastatic HER2(+) breast cancer whose disease had previously progressed on trastuzumab. All patients received weekly treatment with tanespimycin at 450 mg/m(2) intravenously and trastuzumab at a conventional dose. Therapy was continued until disease progression. The primary endpoint was response rate by Response Evaluation Criteria in Solid Tumors (RECIST) criteria. Thirty-one patients were enrolled with a median age of 53 years and a median Karnofsky performance status (KPS) of 90%. The most common toxicities, largely grade 1, were diarrhea, fatigue, nausea, and headache. The overall response rate was 22%, the clinical benefit rate [complete response + partial response + stable disease] was 59%, the median progression-free survival was 6 months (95% CI: 4-9), and the median overall survival was 17 months (95% CI: 16-28). This is the first phase II study to definitively show RECIST-defined responses for 17-AAG in solid tumors. Tanespimycin plus trastuzumab has significant anticancer activity in patients with HER2-positive, metastatic breast cancer previously progressing on trastuzumab. Further research exploring this therapeutic interaction and the activity of HSP90 inhibitors is clearly warranted. ©2011 AACR.

  6. The HSP90 inhibitor 17-N-allylamino-17-demethoxy geldanamycin (17-AAG) synergizes with cisplatin and induces apoptosis in cisplatin-resistant esophageal squamous cell carcinoma cell lines via the Akt/XIAP pathway.

    Science.gov (United States)

    Ui, Takashi; Morishima, Kazue; Saito, Shin; Sakuma, Yuji; Fujii, Hirofumi; Hosoya, Yoshinori; Ishikawa, Shumpei; Aburatani, Hiroyuki; Fukayama, Masashi; Niki, Toshiro; Yasuda, Yoshikazu

    2014-02-01

    Although cisplatin (CDDP) is a key drug in the treatment of esophageal squamous cell carcinoma (ESCC), acquired chemoresistance remains a major problem. Combination therapy may represent one strategy to overcome this resistance. Heat shock protein 90 (HSP90) is known to be overexpressed in several types of cancer cells, and its inhibition by small molecules, either alone or in combination, has shown promise in the treatment of solid malignancies. In the present study, we evaluated the synergistic effects of combining CDDP with the HSP90 inhibitor 17-N-allylamino-17-demethoxy geldanamycin (17-AAG) on two CDDP-resistant human esophageal squamous cancer cell lines, KYSE30 and KYSE150. The results obtained demonstrated the synergistic inhibitory effects of CDDP and 17-AAG on the growth of KYSE30 and KYSE150 cells. Cell growth and cell number were more effectively reduced by the combined treatment with CDDP and 17-AAG than by the treatment with either CDDP or 17-AAG alone. Western blotting revealed that the combined action of CDDP and 17-AAG cleaved poly (ADP-ribose) polymerase (PARP) and caspase-3, which demonstrated that the reduction in both cell growth and cell number was mediated by apoptosis. Time-course experiments showed that reduction in X-linked inhibitor of apoptosis protein (XIAP) and phosphorylated Akt were concomitant with apoptosis. The results of the present study demonstrate that 17-AAG synergizes with CDDP and induces apoptosis in CDDP-resistant ESCC cell lines, and also that modulation of the Akt/XIAP pathway may underlie this synergistic effect. Combination therapy with CDDP and an HSP90 inhibitor may represent a promising strategy to overcome CDDP resistance in ESCC.

  7. The Hsp72 and Hsp90α mRNA Responses to Hot Downhill Running Are Reduced Following a Prior Bout of Hot Downhill Running, and Occur Concurrently within Leukocytes and the Vastus Lateralis

    Directory of Open Access Journals (Sweden)

    James A. Tuttle

    2017-07-01

    Full Text Available The leukocyte heat shock response (HSR is used to determine individual's thermotolerance. The HSR and thermotolerance are enhanced following interventions such as preconditioning and/or acclimation/acclimatization. However, it is unclear whether the leukocyte HSR is an appropriate surrogate for the HSR in other tissues implicated within the pathophysiology of exertional heat illnesses (e.g., skeletal muscle, and whether an acute preconditioning strategy (e.g., downhill running can improve subsequent thermotolerance. Physically active, non-heat acclimated participants were split into two groups to investigate the benefits of hot downhill running as preconditioning strategy. A hot preconditioning group (HPC; n = 6 completed two trials (HPC1HOTDOWN and HPC2HOTDOWN of 30 min running at lactate threshold (LT on −10% gradient in 30°C and 50% relative humidity (RH separated by 7 d. A temperate preconditioning group (TPC; n = 5 completed 30 min running at LT on a −1% gradient in 20°C and 50% (TPC1TEMPFLAT and 7 d later completed 30 min running at LT on −10% gradient in 30°C and 50% RH (TPC2HOTDOWN. Venous blood samples and muscle biopsies (vastus lateralis; VL were obtained before, immediately after, 3, 24, and 48 h after each trial. Leukocyte and VL Hsp72, Hsp90α, and Grp78 mRNA relative expression was determined via RT-QPCR. Attenuated leukocyte and VL Hsp72 (2.8 to 1.8 fold and 5.9 to 2.4 fold; p < 0.05 and Hsp90α mRNA (2.9 to 2.4 fold and 5.2 to 2.4 fold; p < 0.05 responses accompanied reductions (p < 0.05 in physiological strain [exercising rectal temperature (−0.3°C and perceived muscle soreness (~ −14%] during HPC2HOTDOWN compared to HPC1HOTDOWN (i.e., a preconditioning effect. Both VL and leukocyte Hsp72 and Hsp90α mRNA increased (p < 0.05 simultaneously following downhill runs and demonstrated a strong relationship (p < 0.01 of similar magnitudes with one another. Hot downhill running is an effective preconditioning strategy

  8. Effects of HSP90 inhibitor 17-allylamino-17-demethoxygeldanamycin (17-AAG) on NEU/HER2 overexpressing mammary tumours in MMTV-NEU-NT mice monitored by Magnetic Resonance Spectroscopy.

    Science.gov (United States)

    Rodrigues, Loreta M; Chung, Yuen-Li; Al Saffar, Nada M S; Sharp, Swee Y; Jackson, Laura E; Banerji, Udai; Stubbs, Marion; Leach, Martin O; Griffiths, John R; Workman, Paul

    2012-05-23

    The importance of ERBB2/NEU/HER2 in the response of breast tumours to the heat shock protein 90 (HSP90) inhibitor 17-allylamino-17-demethoxygeldanamycin (17-AAG; tanespimycin) has been demonstrated in the clinic. ERBB2 is an oncoprotein client that is highly dependent on HSP90. This and other oncogenic client proteins (e.g. B-RAF, C-RAF, ALK and CDK4) are depleted by 17-AAG in both animal tumours and patients. Here we investigate by Magnetic Resonance Spectroscopy (MRS) the metabolic response of 17-AAG in spontaneous, NEU/HER2 driven mammary tumours in transgenic MMTV-NEU-NT mice and in cells isolated and cultured from these tumours. Mammary tumours were monitored by 31P MRS in vivo and in tumour extracts, comparing control and 17-AAG treated mice. A cell line derived from NEU/HER2 mammary tumours was also cultured and the effect of 17-AAG was measured by 31P MRS in cell extracts. Molecular biomarkers were assessed by immunoblotting in extracts from cells and tumours. For comparison of tumour volume, metabolite concentrations and Western blot band intensities, two-tailed unpaired t-tests were used. The NEU/HER2 mammary tumours were very sensitive to 17-AAG and responded in a dose-dependent manner to 3 daily doses of 20, 40 and 80mg/kg of 17-AAG, all of which caused significant regression. At the higher doses, 31P MRS of tumour extracts showed significant decreases in phosphocholine (PC) and phosphoethanolamine (PE) whereas no significant changes were seen at the 20mg/kg dose. Extracts of isolated cells cultured from the mammary carcinomas showed a significant decrease in viable cell number and total PME after 17-AAG treatment. Western blots confirmed the expected action of 17-AAG in inducing HSP72 and significantly depleting HSP90 client proteins, including NEU/HER2 both in tumours and in isolated cells. The data demonstrate the high degree of sensitivity of this clinically relevant NEU/HER2-driven tumour model to HSP90 inhibition by 17-AAG, consistent with the

  9. ER stress stimulates production of the key antimicrobial peptide, cathelicidin, by forming a previously unidentified intracellular S1P signaling complex.

    Science.gov (United States)

    Park, Kyungho; Ikushiro, Hiroko; Seo, Ho Seong; Shin, Kyong-Oh; Kim, Young Il; Kim, Jong Youl; Lee, Yong-Moon; Yano, Takato; Holleran, Walter M; Elias, Peter; Uchida, Yoshikazu

    2016-03-08

    We recently identified a previously unidentified sphingosine-1-phosphate (S1P) signaling mechanism that stimulates production of a key innate immune element, cathelicidin antimicrobial peptide (CAMP), in mammalian cells exposed to external perturbations, such as UVB irradiation and other oxidative stressors that provoke subapoptotic levels of endoplasmic reticulum (ER) stress, independent of the well-known vitamin D receptor-dependent mechanism. ER stress increases cellular ceramide and one of its distal metabolites, S1P, which activates NF-κB followed by C/EBPα activation, leading to CAMP production, but in a S1P receptor-independent fashion. We now show that S1P activates NF-κB through formation of a previously unidentified signaling complex, consisting of S1P, TRAF2, and RIP1 that further associates with three stress-responsive proteins; i.e., heat shock proteins (GRP94 and HSP90α) and IRE1α. S1P specifically interacts with the N-terminal domain of heat shock proteins. Because this ER stress-initiated mechanism is operative in both epithelial cells and macrophages, it appears to be a universal, highly conserved response, broadly protective against diverse external perturbations that lead to increased ER stress. Finally, these studies further illuminate how ER stress and S1P orchestrate critical stress-specific signals that regulate production of one protective response by stimulating production of the key innate immune element, CAMP.

  10. Increased extracellular heat shock protein 90α in severe sepsis and SIRS associated with multiple organ failure and related to acute inflammatory-metabolic stress response in children.

    Science.gov (United States)

    Fitrolaki, Michaela-Diana; Dimitriou, Helen; Venihaki, Maria; Katrinaki, Marianna; Ilia, Stavroula; Briassoulis, George

    2016-08-01

    Mammalian heat-shock-protein (HSP) 90α rapidly responses to environmental insults. We examined the hypothesis that not only serum HSP72 but also HSP90α is increased in the systemic inflammatory response syndrome (SIRS), severe-sepsis (SS), and/or sepsis (S) compared to healthy children (H); we assessed HSP90α relation to (a) multiple organ system failure (MOSF) and (b) inflammatory-metabolic response and severity of illness.A total of 65 children with S, SS, or SIRS and 25 H were included. ELISA was used to evaluate extracellular HSP90α and HSP72, chemiluminescence interleukins (ILs), flow-cytometry neutrophil-CD64 (nCD64)-expression.HSP90α, along with HSP72, were dramatically increased among MOSF patients. Patients in septic groups and SIRS had elevated HSP90α compared to H (P stress, fever, outcome endpoints, and predicted mortality and inversely related to the low-LDL/low-HDL stress metabolic pattern.

  11. Caracterización in silico de las proteínas del choque térmico Hsp70 y Hsp90 deBemisia tabaci (Hemiptera: Aleyrodidae y su posible actividad adaptativa

    Directory of Open Access Journals (Sweden)

    Eneida Torres Cabra

    2014-06-01

    Full Text Available La mosca blanca, Bemisia tabaci (Hemiptera: Aleyrodidae es una de las plagas más destructivas e invasivas en el mundo, ataca una gran cantidad de cultivos. Se adapta fácilmente a plantas hospederas y a nuevas regiones geográficas, lo que sugiere el desarrollo de mecanismos de control a daños producidos por factores estresantes. Las proteínas Hsp se expresanen los organismos como mecanismo de defensa, actúan como chaperonas en el correcto ensamblaje de las proteínas. En este estudio se realizó una caracterizaciónin silico de las proteínas Hsp70 y Hsp90 de B. tabaci, secuencias obtenidas de NCBI. La determinaciónde los perfiles de hidrofobicidad, polaridad, accesibilidady flexibilidad se obtuvieron con “ProScale” de ExPASy, el perfil de antigenicidad con JaMBW. La secuencia aminoacídica se analizó con GOR IV y SOPMA y la composición de aminoácidos con ProtParam. Para analizar el peso molecular, índice deinestabilidad, índice alifático y gradiente hidropático,con GRAVY. La estructura terciaria se obtuvo con HHpred, y ESyPred3D. Para validar las estructuras 3D se utilizó Procheck, What_check y errat. Hsp70 y Hsp90 de B. tabaci presentan valores bajos de hidrofobicidady altos de polaridad, flexibilidad y accesibilidad, características que le permiten a las proteínas extender su capacidad como chaperonas. La Hsp70tiene una estructura secundaria compuesta por 41-45% alfa hélices, 30-43% coil y menos del 6% en hoja plegada y la Hsp90 por 52 y 53% hélices, 26-34% coily 6% hoja plegada. Las Hsp juegan un rol importante en los insectos debido a su tamaño y corto ciclo de vida, pues la temperatura influye en su distribución y abundancia.

  12. Identification of a heat shock protein 90 gene involved in resistance to temperature stress in two wing-morphs of Nilaparvata lugens (Stål).

    Science.gov (United States)

    Lu, Kai; Chen, Xia; Liu, Wenting; Zhou, Qiang

    2016-07-01

    The brown planthopper, Nilaparvata lugens, is one of the most destructive pests damaging rice in Asia and exhibits wing dimorphism, with brachypters possessing severely reduced wings and macropters bearing fully developed wings. Previous studies have shown that macropters are more heat resistant than brachypters. To understand the molecular mechanism underlying the differential thermotolerance abilities of these two morphs, a full-length Hsp gene, NlHsp90 was cloned from N. lugen. Our results showed that the relative expression levels of NlHsp90 in N. lugens females increased with the rise of temperature. Interestingly, NlHsp90 in macropters females could be induced at lower temperature (32°C) than that in brachypters (34°C), and the NlHsp90 mRNA levels in macropters were significantly higher than those in brachypters from 34 to 40°C. In addition, the maximum expression levels of NlHsp90 were achieved much earlier in macropters, and NlHsp90 mRNA levels in macropters were significantly higher than those in brachypters from 1 to 6h of recovery after temperature stress. Furthermore, knockdown of NlHsp90 by dsRNA injection reduced survival in both morphs with a greater reduction in the macropters relative to that of the brachyters. These results indicated that NlHsp90 plays an important role for thermotolerance in N. lugens, and there is difference on induction between two morphs. Copyright © 2016 Elsevier Inc. All rights reserved.

  13. An improved route to 19-substituted geldanamycins as novel Hsp90 inhibitors ? potential therapeutics in cancer and neurodegeneration? ?Electronic supplementary information (ESI) available. See DOI: 10.1039/c3cc43457e Click here for additional data file. Click here for additional data file.

    OpenAIRE

    Kitson, Russell R. A.; Moody, Christopher J.

    2013-01-01

    19-Substituted geldanamycin derivatives are efficient Hsp90 inhibitors, without the toxicity associated with the other benzoquinone ansamycins, thus giving them potential for use as molecular therapeutics in cancer and neurodegeneration. Here a new method of synthesising these important compounds is reported, eliminating the need for toxic metals and metalloids.

  14. Molecular phylogeny of diplomonads and enteromonads based on SSU rRNA, alpha-tubulin and HSP90 genes: Implications for the evolutionary history of the double karyomastigont of diplomonads

    Directory of Open Access Journals (Sweden)

    Roger Andrew J

    2008-07-01

    Full Text Available Abstract Background Fornicata is a relatively recently established group of protists that includes the diplokaryotic diplomonads (which have two similar nuclei per cell, and the monokaryotic enteromonads, retortamonads and Carpediemonas, with the more typical one nucleus per cell. The monophyly of the group was confirmed by molecular phylogenetic studies, but neither the internal phylogeny nor its position on the eukaryotic tree has been clearly resolved. Results Here we have introduced data for three genes (SSU rRNA, α-tubulin and HSP90 with a wide taxonomic sampling of Fornicata, including ten isolates of enteromonads, representing the genera Trimitus and Enteromonas, and a new undescribed enteromonad genus. The diplomonad sequences formed two main clades in individual gene and combined gene analyses, with Giardia (and Octomitus on one side of the basal divergence and Spironucleus, Hexamita and Trepomonas on the other. Contrary to earlier evolutionary scenarios, none of the studied enteromonads appeared basal to diplokaryotic diplomonads. Instead, the enteromonad isolates were all robustly situated within the second of the two diplomonad clades. Furthermore, our analyses suggested that enteromonads do not constitute a monophyletic group, and enteromonad monophyly was statistically rejected in 'approximately unbiased' tests of the combined gene data. Conclusion We suggest that all higher taxa intended to unite multiple enteromonad genera be abandoned, that Trimitus and Enteromonas be considered as part of Hexamitinae, and that the term 'enteromonads' be used in a strictly utilitarian sense. Our result suggests either that the diplokaryotic condition characteristic of diplomonads arose several times independently, or that the monokaryotic cell of enteromonads originated several times independently by secondary reduction from the diplokaryotic state. Both scenarios are evolutionarily complex. More comparative data on the similarity of the

  15. Functional Genomic Screening Reveals Core Modulators of Echinocandin Stress Responses in Candida albicans

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    Tavia Caplan

    2018-05-01

    Full Text Available Summary: Candida albicans is a leading cause of death due to fungal infection. Treatment of systemic candidiasis often relies on echinocandins, which disrupt cell wall synthesis. Resistance is readily acquired via mutations in the drug target gene, FKS1. Both basal tolerance and resistance to echinocandins require cellular stress responses. We performed a systematic analysis of 3,030 C. albicans mutants to define circuitry governing cellular responses to echinocandins. We identified 16 genes for which deletion or transcriptional repression enhanced echinocandin susceptibility, including components of the Pkc1-MAPK signaling cascade. We discovered that the molecular chaperone Hsp90 is required for the stability of Pkc1 and Bck1, establishing key mechanisms through which Hsp90 mediates echinocandin resistance. We also discovered that perturbation of the CCT chaperonin complex causes enhanced echinocandin sensitivity, altered cell wall architecture, and aberrant septin localization. Thus, we provide insights into the mechanisms by which cellular chaperones enable crucial responses to echinocandin-induced stress. : Caplan et al. screen 3,030 Candida albicans mutants to define circuitry governing cellular responses to echinocandins, the first-line therapy for systemic candidiasis. They reveal that the molecular chaperone Hsp90 is required for stability of Pkc1 and Bck1 and that the CCT chaperonin complex is a key modulator of echinocandin susceptibility. Keywords: fungal pathogen, Candida albicans, echinocandins, Hsp90, Pkc1, CCT complex, client protein, stress response, functional genomic screen, drug resistance

  16. Orchestration in work environment policy programs

    DEFF Research Database (Denmark)

    Hasle, Peter; Limborg, Hans Jørgen; Grøn, Sisse

    2017-01-01

    In spite of many years’ efforts, it is difficult to prove substantial improvements of the work environment and policymakers are continuously searching for new efficient strategies. This paper examines the concept of orchestration of work environment programs, based on an empirical analysis...... of recent Danish policy. Orchestration is a strategy where different stakeholders and activities are integrated into a unified program aimed at a specific target group. The analysis includes three policy cases, supplemented with two company case studies. The research shows a move toward a more governance...... type of regulation, which is not only emerging in network but also includes more explicitly orchestrated policy programs. The stakeholders participate in the network with different interests and the orchestration of work environment policies is therefore built on a platform of regulation...

  17. Araloside C Prevents Hypoxia/Reoxygenation-Induced Endoplasmic Reticulum Stress via Increasing Heat Shock Protein 90 in H9c2 Cardiomyocytes

    Directory of Open Access Journals (Sweden)

    Yuyang Du

    2018-04-01

    Full Text Available Araloside C (AsC is a cardioprotective triterpenoid compound that is mainly isolated from Aralia elata. This study aims to determine the effects of AsC on hypoxia-reoxygenation (H/R-induced apoptosis in H9c2 cardiomyocytes and its underlying mechanisms. Results demonstrated that pretreatment with AsC (12.5 μM for 12 h significantly suppressed the H/R injury in H9c2 cardiomyocytes, including improving cell viability, attenuating the LDH leakage and preventing cardiomyocyte apoptosis. AsC also inhibited H/R-induced ER stress by reducing the activation of ER stress pathways (PERK/eIF2α and ATF6, and decreasing the expression of ER stress-related apoptotic proteins (CHOP and caspase-12. Moreover, AsC greatly improved the expression level of HSP90 compared with that in the H/R group. The use of HSP90 inhibitor 17-AAG and HSP90 siRNA blocked the above suppression effect of AsC on ER stress-related apoptosis caused by H/R. Taken together, AsC could reduce H/R-induced apoptosis possibly because it attenuates ER stress-dependent apoptotic pathways by increasing HSP90 expression.

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

    Science.gov (United States)

    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.

  19. Metric-Aware Secure Service Orchestration

    Directory of Open Access Journals (Sweden)

    Gabriele Costa

    2012-12-01

    Full Text Available Secure orchestration is an important concern in the internet of service. Next to providing the required functionality the composite services must also provide a reasonable level of security in order to protect sensitive data. Thus, the orchestrator has a need to check whether the complex service is able to satisfy certain properties. Some properties are expressed with metrics for precise definition of requirements. Thus, the problem is to analyse the values of metrics for a complex business process. In this paper we extend our previous work on analysis of secure orchestration with quantifiable properties. We show how to define, verify and enforce quantitative security requirements in one framework with other security properties. The proposed approach should help to select the most suitable service architecture and guarantee fulfilment of the declared security requirements.

  20. Russian orchestral works. Torgny Sporsen / Ivan March

    Index Scriptorium Estoniae

    March, Ivan

    1991-01-01

    Uuest heliplaadist "Russian orchestral works. Torgny Sporsen (bass). Gothenburg Symphony Brass Band. Gothenburg Symphony Chorus and Orchestra / Neeme Järvi" D6 MC 429 984 - 4 GH; CD 429 984 - 26H (76 minutes). Borodin: In Central Asia. Prince Igor - Polovtsian Dances

  1. Building a public for orchestral music

    OpenAIRE

    Pasler, Jann

    2017-01-01

    In spring 1873, two years after France’s devastating defeat in the Franco-Prussian war, Georges Hartmann hired Edouard Colonne to conduct eight “Concerts Nationals”. With them, he aimed to promote French music, especially that which he published. The concerts attracted a large, enthusiastic crowd, but lost money. A year later Colonne founded his own orchestra, subsequently known as the Concerts Colonne. Their challenge was to attract a mass public for serious orchestral music, successfully co...

  2. Microsoft System Center 2012 Orchestrator cookbook

    CERN Document Server

    Erskine, Samuel

    2013-01-01

    This book is written in a practical, Cookbook style with numerous chapters and recipes focusing on creating runbooks to automate mission critical and everyday administration tasks.System Center 2012 Orchestrator is for administrators who wish to simplify the process of automating systems administration tasks. This book assumes that you have a basic knowledge of Windows Server 2008 Administration, Active Directory, Network Systems, and Microsoft System Center technologies.

  3. Cocaine withdrawal causes delayed dysregulation of stress genes in the hippocampus.

    Directory of Open Access Journals (Sweden)

    M Julia García-Fuster

    Full Text Available Relapse, even following an extended period of withdrawal, is a major challenge in substance abuse management. Delayed neurobiological effects of the drug during prolonged withdrawal likely contribute to sustained vulnerability to relapse. Stress is a major trigger of relapse, and the hippocampus regulates the magnitude and duration of stress responses. Recent work has implicated hippocampal plasticity in various aspects of substance abuse. We asked whether changes in stress regulatory mechanisms in the hippocampus may participate in the neuroadaptations that occur during prolonged withdrawal. We therefore examined changes in the rat stress system during the course of withdrawal from extended daily access (5-hours of cocaine self-administration, an animal model of addiction. Tissue was collected at 1, 14 and 28 days of withdrawal. Plasma corticosterone levels were determined and corticosteroid receptors (GR, MR, MR/GR mRNA ratios and expression of other stress-related molecules (HSP90AA1 and HSP90AB1 mRNA were measured in hippocampal subfields using in situ hybridization. Results showed a delayed emergence of dysregulation of stress genes in the posterior hippocampus following 28 days of cocaine withdrawal. This included increased GR mRNA in DG and CA3, increased MR and HSP90AA1 mRNA in DG, and decreased MR/GR mRNA ratio in DG and CA1. Corticosterone levels progressively decreased during the course of withdrawal, were normalized following 28 days of withdrawal, and were correlated negatively with GR and positively with MR/GR mRNA ratio in DG. These results suggest a role for the posterior hippocampus in the neuroadaptations that occur during prolonged withdrawal, and point to a signaling partner of GR, HSP90AA1, as a novel dysregulated target during cocaine withdrawal. These delayed neurobiological effects of extended cocaine exposure likely contribute to sustained vulnerability to relapse.

  4. Resveratrol induces antioxidant and heat shock protein mRNA expression in response to heat stress in black-boned chickens.

    Science.gov (United States)

    Liu, L L; He, J H; Xie, H B; Yang, Y S; Li, J C; Zou, Y

    2014-01-01

    This study investigated the effects of dietary resveratrol at 0, 200, 400, or 600 mg/kg of diet on the performance, immune organ growth index, serum parameters, and expression levels of heat shock protein (Hsp) 27, Hsp70, and Hsp90 mRNA in the bursa of Fabricius, thymus, and spleen of 42-d-old female black-boned chickens exposed to heat stress at 37 ± 2°C for 15 d. The results showed that heat stress reduced daily feed intake and BW gain; decreased serum glutathione (GSH), growth hormone, and insulin-like growth factor-1 levels; and inhibited GSH peroxidase (GSH-Px), superoxide dismutase (SOD), and catalase (CAT) activities compared with birds subjected to thermo-neutral circumstances. Chickens that were fed diets supplemented with resveratrol exhibited a linear increase in feed intake and BW gain (P stress. In contrast, serum malonaldehyde concentrations were decreased (P stress also reduced (P stress and coincided with an increase in supplemental resveratrol levels. The expression of Hsp27, Hsp70, and Hsp90 mRNA in the bursa of Fabricius and spleen were increased (P stress compared with no heat stress. Resveratrol attenuated the heat stress-induced overexpression of Hsp27, Hsp70, and Hsp90 mRNA in the bursa of Fabricius and spleen and increased the low expression of Hsp27 and Hsp90 mRNA in thymus upon heat stress. The results suggest that supplemental resveratrol improves growth performance and reduces oxidative stress in heat-stressed black-boned chickens by increasing serum growth hormone concentrations and modulating the expression of heat shock genes in organs of the immune system.

  5. Hochu-ekki-to Treatment Improves Reproductive and Immune Modulation in the Stress-Induced Rat Model of Polycystic Ovarian Syndrome.

    Science.gov (United States)

    Park, Eunkuk; Choi, Chun Whan; Kim, Soo Jeong; Kim, Yong-In; Sin, Samkee; Chu, Jong-Phil; Heo, Jun Young

    2017-06-13

    The traditional herbal medicine, Hochu-ekki-to, has been shown to have preventive effects on viral infection and stress. This study aimed to evaluate the clinical effects of Hochu-ekki-to on two stress-related rat models of polycystic ovarian syndrome. Female Sprague-Dawley rats were divided into control and treatment groups, the latter of which were subjected to stress induced by exposure to adrenocorticotropic hormone (ACTH) or cold temperatures. After these stress inductions, rats were orally treated with dissolved Hochu-ekki-to once per day for 7 days. Rats subjected to the two different stressors exhibited upregulation of steroid hormone receptors (in ovaries) and reproductive hormones (in blood), and consequent stimulation of abnormal follicle development accompanied by elevation of Hsp 90 expression (in ovaries). Treatment with Hochu-ekki-to for 7 days after stress induction increased immune functions, reduced the stress-induced activation of Hsp 90, and normalized the levels of the tested steroid hormone receptors and reproductive hormones. Our findings suggest that stress stimulations may promote the activation of Hsp 90 via the dysregulation of steroid hormone receptors and reproductive hormones, but that post-stress treatment with Hochu-ekki-to improves reproductive and immune functions in the ovaries of stressed rats.

  6. Expression analysis of NOS family and HSP genes during thermal stress in goat ( Capra hircus)

    Science.gov (United States)

    Yadav, Vijay Pratap; Dangi, Satyaveer Singh; Chouhan, Vikrant Singh; Gupta, Mahesh; Dangi, Saroj K.; Singh, Gyanendra; Maurya, Vijay Prakash; Kumar, Puneet; Sarkar, Mihir

    2016-03-01

    Approximately 50 genes other than heat shock protein (HSP) expression changes during thermal stress. These genes like nitric oxide synthase (NOS) need proper attention and investigation to find out their possible role in the adaptation to thermal stress in animals. So, the present study was undertaken to demonstrate the expressions of inducible form type II NOS (iNOS), endothelial type III NOS (eNOS), constitutively expressed enzyme NOS (cNOS), HSP70, and HSP90 in peripheral blood mononuclear cells (PBMCs) during different seasons in Barbari goats. Real-time polymerase chain reaction, western blot, and immunocytochemistry were applied to investigate messenger RNA (mRNA) expression, protein expression, and immunolocalization of examined factors. The mRNA and protein expressions of iNOS, eNOS, cNOS, HSP70, and HSP90 were significantly higher ( P goats.

  7. Meteorological phenomena in Western classical orchestral music

    Science.gov (United States)

    Williams, P. D.; Aplin, K. L.

    2012-12-01

    The creative output of composers, writers, and artists is often influenced by their surroundings. To give a literary example, it has been claimed recently that some of the characters in Oliver Twist and A Christmas Carol were based on real-life people who lived near Charles Dickens in London. Of course, an important part of what we see and hear is not only the people with whom we interact, but also our geophysical surroundings. Of all the geophysical phenomena to influence us, the weather is arguably the most significant, because we are exposed to it directly and daily. The weather was a great source of inspiration for Monet, Constable, and Turner, who are known for their scientifically accurate paintings of the skies. But to what extent does weather inspire composers? The authors of this presentation, who are atmospheric scientists by day but amateur classical musicians by night, have been contemplating this question. We have built a systematic musical database, which has allowed us to catalogue and analyze the frequencies with which weather is depicted in a sample of classical orchestral music. The depictions vary from explicit mimicry using traditional and specialized orchestral instruments, through to subtle suggestions. We have found that composers are generally influenced by their own environment in the type of weather they choose to represent. As befits the national stereotype, British composers seem disproportionately keen to depict the UK's variable weather patterns and stormy coastline. Reference: Aplin KL and Williams PD (2011) Meteorological phenomena in Western classical orchestral music. Weather, 66(11), pp 300-306. doi:10.1002/wea.765

  8. From Orchestration to Choreography through Contract Automata

    Directory of Open Access Journals (Sweden)

    Davide Basile

    2014-10-01

    Full Text Available We study the relations between a contract automata and an interaction model. In the former model, distributed services are abstracted away as automata - oblivious of their partners - that coordinate with each other through an orchestrator. The interaction model relies on channel-based asynchronous communication and choreography to coordinate distributed services. We define a notion of strong agreement on the contract model, exhibit a natural mapping from the contract model to the interaction model, and give conditions to ensure that strong agreement corresponds to well-formed choreography.

  9. Orchestrating Transnational Environmental Governance in Maritime Shipping

    DEFF Research Database (Denmark)

    Lister, Jane; Taudal Poulsen, René; Ponte, Stefano

    2015-01-01

    Maritime shipping is the transmission belt of the global economy. It is also a major contributor to global environmental change through its under-regulated air, water and land impacts. It is puzzling that shipping is a lagging sector as it has a well-established global regulatory body—the Interna......Maritime shipping is the transmission belt of the global economy. It is also a major contributor to global environmental change through its under-regulated air, water and land impacts. It is puzzling that shipping is a lagging sector as it has a well-established global regulatory body......—the International Maritime Organization. Drawing on original empirical evidence and archival data, we introduce a four-factor framework to investigate two main questions: why is shipping lagging in its environmental governance; and what is the potential for the International Maritime Organization to orchestrate......, and growing regulatory fragmentation and uncertainty. The paper concludes with pragmatic recommendations for the International Maritime Organization to acknowledge the regulatory difficulties and seize the opportunity to orchestrate environmental progress....

  10. Orchestration of Social Modes in e-Learning

    DEFF Research Database (Denmark)

    Weinberger, Armin; Papadopoulos, Pantelis M.

    2016-01-01

    of tools offering possibilities to the teachers, orchestration refers to the purposeful mixture of different aspects of the learning experience, serving a particular set of learning goals. In this paper, we present the current dialogue on e-learning orchestration, identifying the questions and open issues...

  11. Service Orchestration on the Internet of Things

    Directory of Open Access Journals (Sweden)

    Jordán Pascual Espada

    2012-12-01

    Full Text Available On July 27, 2010, Jordán Pascual Espada defended his Master’s thesis at Oviedo University (Spain, titled: “Service Orchestration on the internet of things”. This Master’s thesis is the final part of the Web Engineering Official Research Master belonging to the European Higher Education Area. Jordán Pascual Espada defended his dissertation in a publicly open presentation held in the School of Computer Engineering at Oviedo University, and was able to comment on every question raised by his committee and the audience. The master’s thesis was supervised by his advisors, Juan Manuel Cueva Lovelle and Oscar Sanjuán Martínez. The thesis has been read and approved by his thesis committee, receiving the highest rating.

  12. Orchestration of Globally Distributed Knowledge for Innovation in Multinational Companies

    DEFF Research Database (Denmark)

    Sajadirad, Solmaz; Lassen, Astrid Heidemann

    Conducting a multiple-case study in five companies from Danish industry, this paper explores how multinational companies orchestrate knowledge from their globally distributed subsidiaries for innovation. Comparisons of knowledge orchestration within headquarter and subsidiaries for improvement...... and innovation show that a combination of the dynamic use of inter-firm objects and a well-established knowledge orchestration process underlies knowledge orchestration for innovation in multinational companies, as it advances headquarters’ abilities to effectively acquire, evaluate, disseminate, and utilize...... globally distributed knowledge. This study contributes to the understanding of knowledge orchestration between headquarter and distributed subsidiaries in multinational companies and how it is related to innovation. Specifically, this paper has important implications regarding the use of inter-firm objects...

  13. Intracellular proteins produced by mammalian cells in response to environmental stress

    Science.gov (United States)

    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.

  14. Molecular Stress-inducing Compounds Increase Osteoclast Formation in a Heat Shock Factor 1 Protein-dependent Manner*

    Science.gov (United States)

    Chai, Ryan C.; Kouspou, Michelle M.; Lang, Benjamin J.; Nguyen, Chau H.; van der Kraan, A. Gabrielle J.; Vieusseux, Jessica L.; Lim, Reece C.; Gillespie, Matthew T.; Benjamin, Ivor J.; Quinn, Julian M. W.; Price, John T.

    2014-01-01

    Many anticancer therapeutic agents cause bone loss, which increases the risk of fractures that severely reduce quality of life. Thus, in drug development, it is critical to identify and understand such effects. Anticancer therapeutic and HSP90 inhibitor 17-(allylamino)-17-demethoxygeldanamycin (17-AAG) causes bone loss by increasing osteoclast formation, but the mechanism underlying this is not understood. 17-AAG activates heat shock factor 1 (Hsf1), the master transcriptional regulator of heat shock/cell stress responses, which may be involved in this negative action of 17-AAG upon bone. Using mouse bone marrow and RAW264.7 osteoclast differentiation models we found that HSP90 inhibitors that induced a heat shock response also enhanced osteoclast formation, whereas HSP90 inhibitors that did not (including coumermycin A1 and novobiocin) did not affect osteoclast formation. Pharmacological inhibition or shRNAmir knockdown of Hsf1 in RAW264.7 cells as well as the use of Hsf1 null mouse bone marrow cells demonstrated that 17-AAG-enhanced osteoclast formation was Hsf1-dependent. Moreover, ectopic overexpression of Hsf1 enhanced 17-AAG effects upon osteoclast formation. Consistent with these findings, protein levels of the essential osteoclast transcription factor microphthalmia-associated transcription factor were increased by 17-AAG in an Hsf1-dependent manner. In addition to HSP90 inhibitors, we also identified that other agents that induced cellular stress, such as ethanol, doxorubicin, and methotrexate, also directly increased osteoclast formation, potentially in an Hsf1-dependent manner. These results, therefore, indicate that cellular stress can enhance osteoclast differentiation via Hsf1-dependent mechanisms and may significantly contribute to pathological and therapeutic related bone loss. PMID:24692538

  15. Molecular stress-inducing compounds increase osteoclast formation in a heat shock factor 1 protein-dependent manner.

    Science.gov (United States)

    Chai, Ryan C; Kouspou, Michelle M; Lang, Benjamin J; Nguyen, Chau H; van der Kraan, A Gabrielle J; Vieusseux, Jessica L; Lim, Reece C; Gillespie, Matthew T; Benjamin, Ivor J; Quinn, Julian M W; Price, John T

    2014-05-09

    Many anticancer therapeutic agents cause bone loss, which increases the risk of fractures that severely reduce quality of life. Thus, in drug development, it is critical to identify and understand such effects. Anticancer therapeutic and HSP90 inhibitor 17-(allylamino)-17-demethoxygeldanamycin (17-AAG) causes bone loss by increasing osteoclast formation, but the mechanism underlying this is not understood. 17-AAG activates heat shock factor 1 (Hsf1), the master transcriptional regulator of heat shock/cell stress responses, which may be involved in this negative action of 17-AAG upon bone. Using mouse bone marrow and RAW264.7 osteoclast differentiation models we found that HSP90 inhibitors that induced a heat shock response also enhanced osteoclast formation, whereas HSP90 inhibitors that did not (including coumermycin A1 and novobiocin) did not affect osteoclast formation. Pharmacological inhibition or shRNAmir knockdown of Hsf1 in RAW264.7 cells as well as the use of Hsf1 null mouse bone marrow cells demonstrated that 17-AAG-enhanced osteoclast formation was Hsf1-dependent. Moreover, ectopic overexpression of Hsf1 enhanced 17-AAG effects upon osteoclast formation. Consistent with these findings, protein levels of the essential osteoclast transcription factor microphthalmia-associated transcription factor were increased by 17-AAG in an Hsf1-dependent manner. In addition to HSP90 inhibitors, we also identified that other agents that induced cellular stress, such as ethanol, doxorubicin, and methotrexate, also directly increased osteoclast formation, potentially in an Hsf1-dependent manner. These results, therefore, indicate that cellular stress can enhance osteoclast differentiation via Hsf1-dependent mechanisms and may significantly contribute to pathological and therapeutic related bone loss.

  16. Role of Hsp90 in Androgen-Refractory Prostate Cancer

    Science.gov (United States)

    2010-03-01

    designed siRNA sequence using Intergrated DNA Technologies RNAi online software tool (IDT, Coralville, IA). The sequence of siRNA specific for...proliferation and production of prostate-specific antigen in androgen-sensitive pros- tatic cancer cells, LNCaP, by dihydrotestosterone. Endocrinology 136

  17. Molecular mechanisms of canalization: Hsp90 and beyond

    Indian Academy of Sciences (India)

    Madhu Sudhan

    2007-03-26

    Mar 26, 2007 ... clients are essential nodes in signal transduction pathways and regulatory circuits, accounting for the .... respective contributions of genetics versus epigenetics ... authors succeeded in elucidating the molecular mechanism.

  18. Targeting SRC Family Kinases and HSP90 in Lung Cancer

    Science.gov (United States)

    2016-12-01

    accomplished under these goals? Because of the anticipated longer period of analysis, we first began with performance of Major Task 2, assessing the effect ...of NEDD9 genotype with dasatinib and/or ganetespib treatment for effects on tumor growth. Accrual of cohorts was initially slowed because of slow...inhalation of Adeno-Cre, followed by MRI imaging at regular intervals to detect tumor initiation and growth, followed by euthanasia and processing of

  19. Optimization of Orchestral Layouts Based on Instrument Directivity Patterns

    Science.gov (United States)

    Stroud, Nathan Paul

    The experience of hearing an exceptional symphony orchestra perform in an excel- lent concert hall can be profound and moving, causing a level of excitement not often reached for listeners. Romantic period style orchestral music, recognized for validating the use of intense emotion for aesthetic pleasure, was the last significant development in the history of the orchestra. In an age where orchestral popularity is waning, the possibil- ity of evolving the orchestral sound in our modern era exists through the combination of our current understanding of instrument directivity patterns and their interaction with architectural acoustics. With the aid of wave field synthesis (WFS), newly proposed variations on orchestral layouts are tested virtually using a 64-channel WFS array. Each layout is objectively and subjectively compared for determination of which layout could optimize the sound of the orchestra and revitalize the excitement of the performance.

  20. Tchaikovsky, P.: Orchestral Suite no. 3 op. 55 / Terry Williams

    Index Scriptorium Estoniae

    Williams, Terry

    1996-01-01

    Uuest heliplaadist "Tchaikovsky, P.: Orchestral Suite no. 3 op. 55. Francesca di Rimini op. 32. Detroit Symphony Orchestra, Neeme Järvi". Chandos CHAN 9 419, distribution Media 7 (CD: 160F). TT: 1h 09'20"

  1. Orchestration of Globally Distributed Knowledge within MNC Network

    DEFF Research Database (Denmark)

    Sajadirad, Solmaz; Lassen, Astrid Heidemann; Søberg, Peder Veng

    2017-01-01

    distributed knowledge in multinational companies. The findings also indicate that adoption of different knowledge orchestration approaches by companies can be influenced by headquarter-subsidiaries relationship, type of products, knowledge relevance between headquarter and subsidiaries, and level...... on knowledge orchestration in multinational companies. By introducing a dynamic view of the use of inter-firm objects, facilitating collaboration between headquarters and subsidiaries, our findings would help multinational companies improve headquarter-subsidiaries relationship in favor of the efficiency......This paper explores how the use of inter-firm objects within different knowledge orchestration processes affects collaboration between headquarter and distributed subsidiaries. The discussion focuses on different approaches to knowledge orchestration based on combinations of the use of inter...

  2. The orchestration of occupation: the dance of mothers.

    Science.gov (United States)

    Larson, E A

    2000-01-01

    This article describes the relationship of mothers' orchestration of daily occupations, the specialized maternal work of parenting a child with a disability, and the mother's subjective well-being. Mothers' daily occupations and subjective well-being were studied using multiple in-depth interviews, participant observation of a day's round of occupations, and scales of well-being. Data were treated to a recursive analysis, which included theoretical notes generated during transcriptions that identified important themes and additional points of inquiry, line-by-line coding of transcripts, and theoretical sorting of codes and regrouping, recoding. To account for patterns in the data, a relational analysis was conducted that included the generation of metaphors. Emergent findings of this analysis identified the mothers' guiding occupational motif and eight processes of orchestration in their daily routines. The occupational motif, the embrace of paradox, directed the mother's orchestration of daily occupations. The orchestration processes included planning, organizing, balancing, anticipating, interpreting, forecasting, perspective shifting, and meaning making. Examples illustrate the maternally driven and child-sensitive nature of these processes. In their daily rounds, the mothers studied were attentive to the manner and method with which they interacted with their children to produce child-contingent occupations commensurate with their values of being a good mother. Using these orchestration processes, mothers made sense of their past, designed their present, and planned for their future within their daily occupational rounds for themselves and family members.

  3. 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).

  4. 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)

  5. Stress, and pathogen response gene expression in modeled microgravity

    Science.gov (United States)

    Sundaresan, Alamelu; Pellis, Neal R.

    2006-01-01

    Purpose: Immune suppression in microgravity has been well documented. With the advent of human exploration and long-term space travel, the immune system of the astronaut must be optimally maintained. It is important to investigate the expression patterns of cytokine genes, because they are directly related to immune response. Heat shock proteins (HSPs), also called stress proteins, are a group of proteins that are present in the cells of every life form. These proteins are induced when a cell responds to stressors such as heat, cold and oxygen deprivation. Microgravity is another stressor that may regulate HSPs. Heat shock proteins trigger immune response through activities that occur both inside the cell (intracellular) and outside the cell (extracellular). Knowledge about these two gene groups could lead to establishment of a blueprint of the immune response and adaptation-related genes in the microgravity environment. Methods: Human peripheral blood cells were cultured in 1g (T flask) and modeled microgravity (MMG, rotating-wall vessel) for 24 and 72 hours. Cell samples were collected and subjected to gene array analysis using the Affymetrix HG_U95 array. Data was collected and subjected to a two-way analysis of variance. The genes related to immune and stress responses were analyzed. Results and Conclusions: HSP70 was up-regulated by more than two fold in microgravity culture, while HSP90 was significantly down-regulated. HSP70 is not typically expressed in all kinds of cells, but it is expressed at high levels in stress conditions. HSP70 participates in translation, protein translocation, proteolysis and protein folding, suppressing aggregation and reactivating denatured proteins. Increased serum HSP70 levels correlate with a better outcome for heat-stroke or severe trauma patients. At the same time, elevated serum levels of HSP70 have been detected in patients with peripheral or renal vascular disease. HSP90 has been identified in the cytosol, nucleus and

  6. Role of Heat Shock Protein 70 in Induction of Stress Fiber Formation in Rat Arterial Endothelial Cells in Response to Stretch Stress

    International Nuclear Information System (INIS)

    Luo, Shan-Shun; Sugimoto, Keiji; Fujii, Sachiko; Takemasa, Tohru; Fu, Song-Bin; Yamashita, Kazuo

    2007-01-01

    We investigated the mechanism by which endothelial cells (ECs) resist various forms of physical stress using an experimental system consisting of rat arterial EC sheets. Formation of actin stress fibers (SFs) and expression of endothelial heat-shock stress proteins (HSPs) in response to mechanical stretch stress were assessed by immunofluorescence microscopy. Stretch stimulation increased expression of HSPs 25 and 70, but not that of HSP 90. Treatment with SB203580, a p38 MAP kinase inhibitor that acts upstream of the HSP 25 activation cascade, or with geldanamycin, an inhibitor of HSP 90, had no effect on the SF formation response to mechanical stretch stress. In contrast, treatment with quercetin, an HSP 70 inhibitor, inhibited both upregulation of endothelial HSP 70 and formation of SFs in response to tensile stress. In addition, treatment of stretched ECs with cytochalasin D, which disrupts SF formation, did not adversely affect stretch-induced upregulation of endothelial HSP 70. Our data suggest that endothelial HSP 70 plays an important role in inducing SF formation in response to tensile stress

  7. Orchestration in Learning Technology Research: Evaluation of a Conceptual Framework

    Science.gov (United States)

    Prieto, Luis P.; Dimitriadis, Yannis; Asensio-Pérez, Juan I.; Looi, Chee-Kit

    2015-01-01

    The term "orchestrating learning" is being used increasingly often, referring to the coordination activities performed while applying learning technologies to authentic settings. However, there is little consensus about how this notion should be conceptualised, and what aspects it entails. In this paper, a conceptual framework for…

  8. Roles of Teachers in Orchestrating Learning in Elementary Science Classrooms

    Science.gov (United States)

    Zhai, Junqing; Tan, Aik-Ling

    2015-01-01

    This study delves into the different roles that elementary science teachers play in the classroom to orchestrate science learning opportunities for students. Examining the classroom practices of three elementary science teachers in Singapore, we found that teachers shuttle between four key roles in enabling student learning in science. Teachers…

  9. Pärt: Chamber and Orchestral Works / Robert Cowan

    Index Scriptorium Estoniae

    Cowan, Robert

    1994-01-01

    Uuest heliplaadist "Pärt: Chamber and Orchestral Works. Bournemouth Sinfonietta, Richard Studt. EMI Eminence CD CD-EMX 2221; Fratres; Cantus in memory of Benjamin Britten; Summa; Spiegel im Spiegel; Festina lente; Tabula Rasa; Fratres - selected comparison: Bachmann, Kibonoff (12/94)(CATA) 09026 61824-2. Kremer, Jarrett (ECM) 817 764-2

  10. Survivable resource orchestration for optically interconnected data center networks.

    Science.gov (United States)

    Zhang, Qiong; She, Qingya; Zhu, Yi; Wang, Xi; Palacharla, Paparao; Sekiya, Motoyoshi

    2014-01-13

    We propose resource orchestration schemes in overlay networks enabled by optical network virtualization. Based on the information from underlying optical networks, our proposed schemes provision the fewest data centers to guarantee K-connect survivability, thus maintaining resource availability for cloud applications under any failure.

  11. Shostakovich: The Orchestral Songs Vol. 2 / Michael Tanner

    Index Scriptorium Estoniae

    Tanner, Michael

    1996-01-01

    Uuest heliplaadist "Shostakovich: The Orchestral Songs Vol. 2: Six Romances on texts by Japanese poets, Op. 21. Six Poems on Marina Tsvetayeva, Op. 143. Suite on Verses of Michelangelo, Op. 145. Gothenburg Symphony Orchestra, Neeme Järvi". DG 447 085-2GH (71 minutes:DDD)

  12. Orchestration of Social Modes in e-Learning

    DEFF Research Database (Denmark)

    Weinberger, Armin; Papadopoulos, Pantelis M.

    2016-01-01

    The concept of orchestration has recently emerged as a useful metaphor in technology-enhanced learning research communities, because of its explanatory power and appeal in describing how different learning activities, tools, and arrangements could be combined to promote learning. More than a buff...

  13. Differential responses of the coral host and their algal symbiont to thermal stress.

    Directory of Open Access Journals (Sweden)

    William Leggat

    Full Text Available The success of any symbiosis under stress conditions is dependent upon the responses of both partners to that stress. The coral symbiosis is particularly susceptible to small increases of temperature above the long term summer maxima, which leads to the phenomenon known as coral bleaching, where the intracellular dinoflagellate symbionts are expelled. Here we for the first time used quantitative PCR to simultaneously examine the gene expression response of orthologs of the coral Acropora aspera and their dinoflagellate symbiont Symbiodinium. During an experimental bleaching event significant up-regulation of genes involved in stress response (HSP90 and HSP70 and carbon metabolism (glyceraldehyde-3-phosphate dehydrogenase, α-ketoglutarate dehydrogenase, glycogen synthase and glycogen phosphorylase from the coral host were observed. In contrast in the symbiont, HSP90 expression decreased, while HSP70 levels were increased on only one day, and only the α-ketoglutarate dehydrogenase expression levels were found to increase. In addition the changes seen in expression patterns of the coral host were much larger, up to 10.5 fold, compared to the symbiont response, which in all cases was less than 2-fold. This targeted study of the expression of key metabolic and stress genes demonstrates that the response of the coral and their symbiont vary significantly, also a response in the host transcriptome was observed prior to what has previously been thought to be the temperatures at which thermal stress events occur.

  14. SGX-Aware Container Orchestration for Heterogeneous Clusters

    OpenAIRE

    Vaucher, Sébastien; Pires, Rafael; Felber, Pascal; Pasin, Marcelo; Schiavoni, Valerio; Fetzer, Christof

    2018-01-01

    Containers are becoming the de facto standard to package and deploy applications and micro-services in the cloud. Several cloud providers (e.g., Amazon, Google, Microsoft) begin to offer native support on their infrastructure by integrating container orchestration tools within their cloud offering. At the same time, the security guarantees that containers offer to applications remain questionable. Customers still need to trust their cloud provider with respect to data and code integrity. The ...

  15. Orchestration Framework for Learning Activities in Augmented Reality Environments

    OpenAIRE

    Ibáñez, María Blanca; Delgado Kloos, Carlos; Di Serio, Angela

    2011-01-01

    Proceedings of: Across Spaces11 Workshop in conjunction with the EC-TEL2011, Palermo, Italy, September 21, 2011 In this paper we show how Augmented Reality (AR) technology restricted to the use of mobiles or PCs, can be used to develop learning activities with the minimun level of orchestation required by meaningful learning sequences. We use Popcode as programming language to deploy orchestrated learning activities specified with an AR framework. Publicado

  16. Decentralized Services Orchestration Using Intelligent Mobile Agents with Deadline Restrictions

    OpenAIRE

    Magalhães , Alex; Lung , Lau Cheuk; Rech , Luciana

    2010-01-01

    International audience; The necessity for better performance drives service orchestration towards decentralization. There is a recent approach where the integrator - that traditionally centralizes all corporative services and business logics - remains as a repository of interface services, but now lacks to know all business logics and business workflows. There are several techniques using this recent approach, including hybrid solutions, peer-to-peer solutions and trigger-based mechanisms. A ...

  17. Perception of orchestral musicians about work environment and conditions

    OpenAIRE

    Clarissa Stefani Teixeira; Fausto Kothe; Luis Felipe Dias Lopes; Érico Felden Pereira

    2014-01-01

    The objective of this study was to investigate the perception of 11 orchestral string (viola and violin) musicians of both genders with respect to their work environment and conditions. We applied a questionnaire with demographic information and the scale Profile of Work Environment and Working Conditions by Nahas et al. (2009), which analyzes the following components: physical environment, social environment, development and professional achievement, salary and benefits, and social relev...

  18. Blockchain Orchestration and Experimentation Framework: A Case Study of KYC

    OpenAIRE

    Shbair, Wazen; Steichen, Mathis Georges; François, Jérôme; State, Radu

    2018-01-01

    Conducting experiments to evaluate blockchain applications is a challenging task for developers, because there is a range of configuration parameters that control blockchain environments. Many public testnets (e.g. Rinkeby Ethereum) can be used for testing, however, we cannot adjust their parameters (e.g. Gas limit, Mining difficulty) to further the understanding of the application in question and of the employed blockchain. This paper proposes an easy to use orchestration framework over the ...

  19. Seasonal variations of cellular stress response of the gilthead sea bream (Sparus aurata).

    Science.gov (United States)

    Feidantsis, Konstantinos; Antonopoulou, Efthimia; Lazou, Antigone; Pörtner, Hans O; Michaelidis, Basile

    2013-07-01

    The present study aimed to investigate the seasonal cellular stress response in vital organs, like the heart, the liver, the whole blood and the skeletal (red and white) muscles of the Mediterranean fish Sparus aurata during a 1-year acclimatization period in the field, in two examined depths (0-2 m and 10-12 m). Processes studied included heat shock protein expression and protein kinase activation. Molecular responses were addressed through the expression of Hsp70 and Hsp90, the phosphorylation of stress-activated protein kinases and particularly p38 mitogen-activated protein kinase (p38 MAPK), the extracellular signal-regulated kinases (ERK-1/2) and c-Jun N-terminal kinases (JNK1/2/3). The induction of Hsp70 and Hsp90 and the phosphorylation of p38 MAPK, JNKs and ERKs in the examined five tissues of the gilthead sea bream indicated a cellular stress response under the prism of a seasonal pattern which was characterized by distinct tissue specificity. Specifically, Hsp induction and MAPK activation occurred before peak summer water temperatures, with no further increases in their levels despite increases in water temperatures. Moreover, although water temperature did not vary significantly with depth of immersion, significant effects of depth on cellular stress response were observed, probably caused by different light regime. The expression and the activation of these certain proteins can be used as tools to define the extreme thermal limits of the gilthead sea bream.

  20. MicroRNA-orchestrated pathophysiologic control in gut homeostasis and inflammation.

    Science.gov (United States)

    Lee, Juneyoung; Park, Eun Jeong; Kiyono, Hiroshi

    2016-05-01

    The intestine represents the largest and most elaborate immune system organ, in which dynamic and reciprocal interplay among numerous immune and epithelial cells, commensal microbiota, and external antigens contributes to establishing both homeostatic and pathologic conditions. The mechanisms that sustain gut homeostasis are pivotal in maintaining gut health in the harsh environment of the gut lumen. Intestinal epithelial cells are critical players in creating the mucosal platform for interplay between host immune cells and luminal stress inducers. Thus, knowledge of the epithelial interface between immune cells and the luminal environment is a prerequisite for a better understanding of gut homeostasis and pathophysiologies such as inflammation. In this review, we explore the importance of the epithelium in limiting or promoting gut inflammation (e.g., inflammatory bowel disease). We also introduce recent findings on how small RNAs such as microRNAs orchestrate pathophysiologic gene regulation. [BMB Reports 2016; 49(5): 263-269].

  1. Retrograde Signals: Integrators of Interorganellar Communication and Orchestrators of Plant Development.

    Science.gov (United States)

    de Souza, Amancio; Wang, Jin-Zheng; Dehesh, Katayoon

    2017-04-28

    Interorganellar cooperation maintained via exquisitely controlled retrograde-signaling pathways is an evolutionary necessity for maintenance of cellular homeostasis. This signaling feature has therefore attracted much research attention aimed at improving understanding of the nature of these communication signals, how the signals are sensed, and ultimately the mechanism by which they integrate targeted processes that collectively culminate in organellar cooperativity. The answers to these questions will provide insight into how retrograde-signal-mediated regulatory mechanisms are recruited and which biological processes are targeted, and will advance our understanding of how organisms balance metabolic investments in growth against adaptation to environmental stress. This review summarizes the present understanding of the nature and the functional complexity of retrograde signals as integrators of interorganellar communication and orchestrators of plant development, and offers a perspective on the future of this critical and dynamic area of research.

  2. Transport stress induces heart damage in newly hatched chicks via blocking the cytoprotective heat shock response and augmenting nitric oxide production.

    Science.gov (United States)

    Sun, F; Zuo, Y-Z; Ge, J; Xia, J; Li, X-N; Lin, J; Zhang, C; Xu, H-L; Li, J-L

    2018-04-20

    Transport stress affects the animal's metabolism and psychological state. As a pro-survival pathway, the heat shock response (HSR) protects healthy cells from stressors. However, it is unclear whether the HSR plays a role in transport stress-induced heart damage. To evaluate the effects of transport stress on heart damage and HSR protection, newly hatched chicks were treated with transport stress for 2 h, 4 h and 8 h. Transport stress caused decreases in body weight and increases in serum creatine kinase (CK) activity, nitric oxide (NO) content in heart tissue, cardiac nitric oxide syntheses (NOS) activity and NOS isoforms transcription. The mRNA expression of heat shock factors (HSFs, including HSF1-3) and heat shock proteins (HSPs, including HSP25, HSP40, HSP47, HSP60, HSP70, HSP90 and HSP110) in the heart of 2 h transport-treated chicks was upregulated. After 8 h of transport stress in chicks, the transcription levels of the same HSPs and HSF2 were reduced in the heart. It was also found that the changes in the HSP60, HSP70 and HSP90 protein levels had similar tendencies. These results suggested that transport stress augmented NO generation through enhancing the activity of NOS and the transcription of NOS isoforms. Therefore, this study provides new evidence that transport stress induces heart damage in the newly hatched chicks by blocking the cytoprotective HSR and augmenting NO production.

  3. Effects of Aging on Musical Performance in Professional Orchestral Musicians.

    Science.gov (United States)

    Kenny, Dianna T; Driscoll, Tim; Ackermann, Bronwen J

    2018-03-01

    The physical and psychological demands of playing a musical instrument are likely to be affected by age-related decline in function, including physical, cognitive, psychological, and organ-related changes. However, the complex neurophysiological demands of playing a musical instrument may delay many normal aging-related changes. This study compared professional classical musicians of different ages, using a range of physical and psychological measures, to discover how increasing age might affect work performance and to identify possible risk and protective factors for physical and psychological health as the musicians age. 377 professional orchestral musicians from eight Australian orchestras (70% response rate), ages 18 to 68 yrs (mean 42.1). Multiple standardized physical and psychological tools were used to evaluate the impact of age on a range of physical and mental health variables. Age was not statistically associated with frequency or severity of performance-related musculoskeletal disorders, ratings of perceived exertion, QuickDASH scores, use of beta-blockers, workplace satisfaction, and most psychological tests. Differences were observed on SPIN (social anxiety) scores, with lowest scores in the oldest age group (10.66 in 55+ yrs vs 17.83 in 18-30 yrs, p=0.016). Older musicians had higher BMIs and fewer practice sessions per day than younger musicians and also were more likely to consume alcohol on 5+ days/wk (44% vs 9%, p=0.003). Advancing age does not appear to exert undue negative impacts on physical and psychological health or performance capacity of professional orchestral musicians. However, dwindling numbers in the older age groups may suggest a "survivor" effect, whereby those who develop significant age-related decrements may cease professional performance at earlier ages. Longitudinal studies on the professional trajectories of professional orchestral musicians are needed to explore this question further.

  4. Orchestrating care: nursing practice with hospitalised older adults.

    Science.gov (United States)

    Dahlke, Sherry Ann; Phinney, Alison; Hall, Wendy Ann; Rodney, Patricia; Baumbusch, Jennifer

    2015-12-01

    The increased incidence of health challenges with aging means that nurses are increasingly caring for older adults, often in hospital settings. Research about the complexity of nursing practice with this population remains limited. To seek an explanation of nursing practice with hospitalised older adults. Design. A grounded theory study guided by symbolic interactionism was used to explore nursing practice with hospitalised older adults from a nursing perspective. Glaserian grounded theory methods were used to develop a mid-range theory after analysis of 375 hours of participant observation, 35 interviews with 24 participants and review of selected documents. The theory of orchestrating care was developed to explain how nurses are continuously trying to manage their work environments by understanding the status of the patients, their unit, mobilising the assistance of others and stretching available resources to resolve their problem of providing their older patients with what they perceived as 'good care' while sustaining themselves as 'good' nurses. They described their practice environments as hard and under-resourced. Orchestrating care is comprised of two subprocesses: building synergy and minimising strain. These two processes both facilitated and constrained each other and nurses' abilities to orchestrate care. Although system issues presented serious constraints to nursing practice, the ways in which nurses were making meaning of their work environment both aided them in managing their challenges and constrained their agency. Nurses need to be encouraged to share their important perspective about older adult care. Administrators have a role to play in giving nurses voice in workplace committees and in forums. Further research is needed to better understand how multidisciplinary teams influence care of hospitalized older adults. © 2014 John Wiley & Sons Ltd.

  5. Hexavalent chromium, a lung carcinogen, confers resistance to thermal stress and interferes with heat shock protein expression in human bronchial epithelial cells.

    Science.gov (United States)

    Abreu, Patrícia L; Cunha-Oliveira, Teresa; Ferreira, Leonardo M R; Urbano, Ana M

    2018-03-16

    Exposure to hexavalent chromium [Cr(VI)], a lung carcinogen, triggers several types of cellular stresses, namely oxidative, genotoxic and proteotoxic stresses. Given the evolutionary character of carcinogenesis, it is tempting to speculate that cells that survive the stresses produced by this carcinogen become more resistant to subsequent stresses, namely those encountered during neoplastic transformation. To test this hypothesis, we determined whether pre-incubation with Cr(VI) increased the resistance of human bronchial epithelial cells (BEAS-2B cells) to the antiproliferative action of acute thermal shock, used here as a model for stress. In line with the proposed hypothesis, it was observed that, at mildly cytotoxic concentrations, Cr(VI) attenuated the antiproliferative effects of both cold and heat shock. Mechanistically, Cr(VI) interfered with the expression of two components of the stress response pathway: heat shock proteins Hsp72 and Hsp90α. Specifically, Cr(VI) significantly depleted the mRNA levels of the former and the protein levels of the latter. Significantly, these two proteins are members of heat shock protein (Hsp) families (Hsp70 and Hsp90, respectively) that have been implicated in carcinogenesis. Thus, our results confirm and extend previous studies showing the capacity of Cr(VI) to interfere with the expression of stress response components.

  6. Public Orchestration, Social Networks, and Transnational Environmental Governance

    DEFF Research Database (Denmark)

    Henriksen, Lasse Folke; Ponte, Stefano

    2018-01-01

    This article contributes to current debates on the potential and limitations of transnational environmental governance, addressing in particular the issue of how private and public regulation compete and/or reinforce each other - and with what results. One of the most influential approaches...... that a social network analytical perspective on orchestration can improve our understanding of how governments and international organizations can shape transnational environmental governance. Through a case study of aviation, we provide two contributions to these debates: first, we propose four analytical...... and institutions that provide the infrastructure of governance....

  7. Natural Killer Cells in the Orchestration of Chronic Inflammatory Diseases

    Directory of Open Access Journals (Sweden)

    Luca Parisi

    2017-01-01

    Full Text Available Inflammation, altered immune cell phenotype, and functions are key features shared by diverse chronic diseases, including cardiovascular, neurodegenerative diseases, diabetes, metabolic syndrome, and cancer. Natural killer cells are innate lymphoid cells primarily involved in the immune system response to non-self-components but their plasticity is largely influenced by the pathological microenvironment. Altered NK phenotype and function have been reported in several pathological conditions, basically related to impaired or enhanced toxicity. Here we reviewed and discussed the role of NKs in selected, different, and “distant” chronic diseases, cancer, diabetes, periodontitis, and atherosclerosis, placing NK cells as crucial orchestrator of these pathologic conditions.

  8. Visual Complaints and Eye Problems in Orchestral Musicians.

    Science.gov (United States)

    Beckers, Henny Jm; van Kooten-Noordzij, Marina Aw; de Crom, Ronald Mpc; Schouten, Jan Sag; Webers, Carroll Ab

    2016-09-01

    To study visual complaints and eye diseases among professional and amateur orchestral musicians in the Netherlands. In this observational study, members from professional and amateur symphony or wind orchestras were asked to complete a questionnaire collecting demographic data, musical, medical, and family history, and data on present visual complaints and/or eye diseases. Questions about playing in the orchestra were also asked. Data from 70 professionals and 48 amateurs showed that most musicians needed glasses or contact lenses for playing in the orchestra (61% of the professionals, 63% of the amateurs). A majority (66% of professionals, 71% of amateurs) had visited an ophthalmologist at least once during their lifetime, and 10% of the professionals and 23% of the amateurs were currently under treatment of an ophthalmologist. Visual complaints while playing in the orchestra were quite common and included poor lighting conditions, problems with reading small notes, blurred vision, tired eyes, and itching or burning eyes. Professional musicians especially reported adverse effects of eye complaints encountered in the orchestra for daily life; 35% got tired earlier and 33% felt that they could not adequately perform their tasks in the orchestra. The results show that visual complaints and eye problems probably are quite common among orchestral musicians and therefore warrant further interest and research.

  9. TRIO: Burst Buffer Based I/O Orchestration

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Teng [Auburn University; Oral, H Sarp [ORNL; Pritchard, Michael [Auburn University; Wang, Bin [Auburn University; Yu, Weikuan [Auburn University

    2015-01-01

    The growing computing power on leadership HPC systems is often accompanied by ever-escalating failure rates. Checkpointing is a common defensive mechanism used by scientific applications for failure recovery. However, directly writing the large and bursty checkpointing dataset to parallel filesystem can incur significant I/O contention on storage servers. Such contention in turn degrades the raw bandwidth utilization of storage servers and prolongs the average job I/O time of concurrent applications. Recently burst buffer has been proposed as an intermediate layer to absorb the bursty I/O traffic from compute nodes to storage backend. But an I/O orchestration mechanism is still desired to efficiently move checkpointing data from bursty buffers to storage backend. In this paper, we propose a burst buffer based I/O orchestration framework, named TRIO, to intercept and reshape the bursty writes for better sequential write traffic to storage severs. Meanwhile, TRIO coordinates the flushing orders among concurrent burst buffers to alleviate the contention on storage server bandwidth. Our experimental results reveal that TRIO can deliver 30.5% higher bandwidth and reduce the average job I/O time by 37% on average for data-intensive applications in various checkpointing scenarios.

  10. Effects of different rearing temperatures on muscle development and stress response in the early larval stages of Acipenser baerii

    Directory of Open Access Journals (Sweden)

    Lucia Aidos

    2017-11-01

    Full Text Available The present study aims at investigating muscle development and stress response in early stages of Siberian sturgeon when subjected to different rearing temperatures, by analysing growth and development of the muscle and by assessing the stress response of yolk-sac larvae. Siberian sturgeon larvae were reared at 16°C, 19°C and 22°C until the yolk-sac was completely absorbed. Sampling timepoints were: hatching, schooling and complete yolk-sac absorption stage. Histometrical, histochemical and immunohistochemical analyses were performed in order to characterize muscle growth (total muscle area, TMA; slow muscle area, SMA; fast muscle area, FMA, development (anti-proliferating cell nuclear antigen -PCNA or anticaspase as well as stress conditions by specific stress biomarkers (heat shock protein 70 or 90, HSP70 or HSP90. Larvae subjected to the highest water temperature showed a faster yolk-sac absorption. Histometry revealed that both TMA and FMA were larger in the schooling stage at 19°C while no differences were observed in the SMA at any of the tested rearing temperatures. PCNA quantification revealed a significantly higher number of proliferating cells in the yolk-sac absorption phase at 22°C than at 16°C. HSP90 immunopositivity seems to be particularly evident at 19°C. HPS70 immunopositivity was never observed in the developing lateral muscle.

  11. Molecular Cloning and mRNA Expression of Heat Shock Protein Genes and Their Response to Cadmium Stress in the Grasshopper Oxya chinensis.

    Directory of Open Access Journals (Sweden)

    Yuping Zhang

    Full Text Available Heat shock proteins (Hsps are highly conserved molecular chaperones that are synthesized in response to stress. In this study, we cloned the full-length sequences of the Grp78 (glucose-regulated protein 78, Hsp70, Hsp90, and Hsp40 genes from the Chinese rice grasshopper Oxya chinensis. The full-length cDNA sequences of OcGrp78, OcHsp70, OcHsp90, and OcHsp40 contain open reading frames of 1947, 1920, 2172, and 1042 bp that encode proteins of 649, 640, 724, and 347 amino acids, respectively. Fluorescent real-time quantitative PCR (RT-qPCR was performed to quantify the relative transcript levels of these Hsp genes in different tissues and developmental stages. The mRNAs encoding these four Hsp genes were present at all developmental stages and in all tissues examined but were expressed at varying levels. Additionally, we investigated the mRNA expression profiles of these four Hsps in O. chinensis subjected to Cadmium (Cd stress. OcGrp78, OcHsp70, OcHsp90, and OcHsp40 mRNA expression was induced under acute Cd stress; the levels reached a maximum within a short time (6 h, were reduced significantly at 12 h, and were lowered to or below control levels by 48 h. Regarding induction efficiency, OcHsp70 was the most sensitive gene to acute Cd stress. Chronic Cd exposure showed that dietary Cd treatment induced increased OcGrp78, OcHsp90, and OcHsp40 expression. However, dietary Cd induced a significant reduction of OcHsp70 expression. In the period tested, no significant difference in the mortality of the grasshoppers was observed. Our results suggest that these four Hsps genes, especially OcHsp70, are sensitive to acute Cd stress and could be used as molecular markers for toxicology studies. However, our results also indicate that OcHsp70 is not suitable for use as a molecular marker of chronic Cd contamination.

  12. Molecular Cloning and mRNA Expression of Heat Shock Protein Genes and Their Response to Cadmium Stress in the Grasshopper Oxya chinensis.

    Science.gov (United States)

    Zhang, Yuping; Liu, Yaoming; Zhang, Jianzhen; Guo, Yaping; Ma, Enbo

    2015-01-01

    Heat shock proteins (Hsps) are highly conserved molecular chaperones that are synthesized in response to stress. In this study, we cloned the full-length sequences of the Grp78 (glucose-regulated protein 78), Hsp70, Hsp90, and Hsp40 genes from the Chinese rice grasshopper Oxya chinensis. The full-length cDNA sequences of OcGrp78, OcHsp70, OcHsp90, and OcHsp40 contain open reading frames of 1947, 1920, 2172, and 1042 bp that encode proteins of 649, 640, 724, and 347 amino acids, respectively. Fluorescent real-time quantitative PCR (RT-qPCR) was performed to quantify the relative transcript levels of these Hsp genes in different tissues and developmental stages. The mRNAs encoding these four Hsp genes were present at all developmental stages and in all tissues examined but were expressed at varying levels. Additionally, we investigated the mRNA expression profiles of these four Hsps in O. chinensis subjected to Cadmium (Cd) stress. OcGrp78, OcHsp70, OcHsp90, and OcHsp40 mRNA expression was induced under acute Cd stress; the levels reached a maximum within a short time (6 h), were reduced significantly at 12 h, and were lowered to or below control levels by 48 h. Regarding induction efficiency, OcHsp70 was the most sensitive gene to acute Cd stress. Chronic Cd exposure showed that dietary Cd treatment induced increased OcGrp78, OcHsp90, and OcHsp40 expression. However, dietary Cd induced a significant reduction of OcHsp70 expression. In the period tested, no significant difference in the mortality of the grasshoppers was observed. Our results suggest that these four Hsps genes, especially OcHsp70, are sensitive to acute Cd stress and could be used as molecular markers for toxicology studies. However, our results also indicate that OcHsp70 is not suitable for use as a molecular marker of chronic Cd contamination.

  13. Technology-Supported Orchestration Matters: Outperforming Paper-Based Scripting in a Jigsaw Classroom

    Science.gov (United States)

    Balestrini, Mara; Hernandez-Leo, Davinia; Nieves, Raul; Blat, Josep

    2014-01-01

    Under the umbrella of ubiquitous technologies, many computational artifacts have been designed to enhance the learning experience in physical settings such as classrooms or playgrounds, but few of them focus on aiding orchestration. This paper presents a systematic evaluation of the signal orchestration system (SOS) used by students for a jigsaw…

  14. Effect of Majority Consensus on Preferences for Recorded Orchestral and Popular Music.

    Science.gov (United States)

    Furman, Charles E.; Duke, Robert A.

    1988-01-01

    Examines group influences regarding music preferences to determine the effect that conformity has on the decision-making process. The study tested participants selections of popular and orchestral excerpts which had altered pitch and/or tempo. Concludes that preferences of music majors regarding orchestral music are not significantly affected by…

  15. Engaging with Mathematics in the Kindergarten. Orchestrating a Fairy Tale through Questioning and Use of Tools

    Science.gov (United States)

    Carlsen, Martin

    2013-01-01

    The aim of this study is to analyse how a kindergarten teacher orchestrated a mathematical activity involving a fairy tale. Taking a sociocultural perspective on learning and development, naturally occurring talk-in-interaction has been analysed in order to scrutinise the subtleties of the orchestration. The fairy tale "Goldilocks and the…

  16. A Study of Master's Degrees in Orchestral Conducting in the United States

    Science.gov (United States)

    St. John, Brian Allen

    2010-01-01

    In order to learn to be an orchestra conductor in the United States of America, students often begins their formal education by seeking to earn a master's degree in orchestral conducting. This project compiled a listing of American universities which offer a master's degree in orchestral conducting and categorized the component parts of their…

  17. Seasonal variations of cellular stress response in the heart and gastrocnemius muscle of the water frog (Pelophylax ridibundus).

    Science.gov (United States)

    Feidantsis, Konstantinos; Anestis, Andreas; Vasara, Eleni; Kyriakopoulou-Sklavounou, Pasqualina; Michaelidis, Basile

    2012-08-01

    The present study aimed to investigate the seasonal cellular stress response in the heart and the gastrocnemius muscle of the amphibian Pelophylax ridibundus (former name Rana ridibunda) during an 8 month acclimatization period in the field. Processes studied included heat shock protein expression and protein kinase activation. The cellular stress response was addressed through the expression of Hsp70 and Hsp90 and the phosphorylation of stress-activated protein kinases and particularly p38 mitogen-activated protein kinase (p38 MAPK), the extracellular signal-regulated kinases (ERK-1/2) and c-Jun N-terminal kinases (JNK1/2/3). Due to a general metabolic depression during winter hibernation, the induction of Hsp70 and Hsp90 and the phosphorylation of p38 MAPK, JNKs and ERKs are retained at low levels of expression in the examined tissues of P. ridibundus. Recovery from hibernation induces increased levels of the specific proteins, probably providing stamina to the animals during their arousal. Copyright © 2012 Elsevier Inc. All rights reserved.

  18. Expression of HSPs: an adaptive mechanism during long-term heat stress in goats ( Capra hircus)

    Science.gov (United States)

    Dangi, Satyaveer Singh; Gupta, Mahesh; Dangi, Saroj K.; Chouhan, Vikrant Singh; Maurya, V. P.; Kumar, Puneet; Singh, Gyanendra; Sarkar, Mihir

    2015-08-01

    Menacing global rise in surface temperature compelled more focus of research over understanding heat stress response mechanism of animals and mitigation of heat stress. Twenty-four goats divided into four groups ( n = 6) such as NHS (non-heat-stressed), HS (heat-stressed), HS + VC (heat-stressed administered with vitamin C), and HS + VE + Se (heat-stressed administered with vitamin E and selenium). Except NHS group, other groups were exposed to repeated heat stress (42 °C) for 6 h on 16 consecutive days. Blood samples were collected at the end of heat exposure on days 1, 6, 11, and 16. When groups compared between days, expression of all heat shock proteins (HSPs) showed a similar pattern as first peak on day 1, reached to basal level on the sixth day, and followed by second peak on day 16. The relative messenger RNA (mRNA) and protein expression of HSP 60, HSP70, and HSP90 was observed highest ( P < 0.05) in HS group, followed by antioxidant-administered group on days 1 and 16, which signifies that antioxidants have dampening effect on HSP expression. HSP105/110 expression was highest ( P < 0.05) on day 16. We conclude that HSP expression pattern is at least two-peak phenomenon, i.e., primary window of HSP protection on the first day followed by second window of protection on day 16. HSP60, HSP70, and HSP90 play an important role during the initial phase of heat stress acclimation whereas HSP105/110 joins this cascade at later phase. Antioxidants may possibly attenuate the HSP expression by reducing the oxidative stress.

  19. The effects of acute waterborne exposure to sublethal concentrations of molybdenum on the stress response in rainbow trout, Oncorhynchus mykiss.

    Directory of Open Access Journals (Sweden)

    Chelsea D Ricketts

    Full Text Available To determine if molybdenum (Mo is a chemical stressor, fingerling and juvenile rainbow trout (Oncorhynchus mykiss were exposed to waterborne sodium molybdate (0, 2, 20, or 1,000 mg l-1 of Mo and components of the physiological (plasma cortisol, blood glucose, and hematocrit and cellular (heat shock protein [hsp] 72, hsp73, and hsp90 in the liver, gills, heart, and erythrocytes and metallothionein [MT] in the liver and gills stress responses were measured prior to initiation of exposure and at 8, 24, and 96 h. During the acute exposure, plasma cortisol, blood glucose, and hematocrit levels remained unchanged in all treatments. Heat shock protein 72 was not induced as a result of exposure and there were no detectable changes in total hsp70 (72 and 73, hsp90, and MT levels in any of the tissues relative to controls. Both fingerling and juvenile fish responded with similar lack of apparent sensitivity to Mo exposure. These experiments demonstrate that exposure to waterborne Mo of up to 1,000 mg l(-1 did not activate a physiological or cellular stress response in fish. Information from this study suggests that Mo water quality guidelines for the protection of aquatic life are highly protective of freshwater fish, namely rainbow trout.

  20. The Effects of Acute Waterborne Exposure to Sublethal Concentrations of Molybdenum on the Stress Response in Rainbow Trout, Oncorhynchus mykiss

    Science.gov (United States)

    Ricketts, Chelsea D.; Bates, William R.; Reid, Scott D.

    2015-01-01

    To determine if molybdenum (Mo) is a chemical stressor, fingerling and juvenile rainbow trout (Oncorhynchus mykiss) were exposed to waterborne sodium molybdate (0, 2, 20, or 1,000 mg l-1 of Mo) and components of the physiological (plasma cortisol, blood glucose, and hematocrit) and cellular (heat shock protein [hsp] 72, hsp73, and hsp90 in the liver, gills, heart, and erythrocytes and metallothionein [MT] in the liver and gills) stress responses were measured prior to initiation of exposure and at 8, 24, and 96 h. During the acute exposure, plasma cortisol, blood glucose, and hematocrit levels remained unchanged in all treatments. Heat shock protein 72 was not induced as a result of exposure and there were no detectable changes in total hsp70 (72 and 73), hsp90, and MT levels in any of the tissues relative to controls. Both fingerling and juvenile fish responded with similar lack of apparent sensitivity to Mo exposure. These experiments demonstrate that exposure to waterborne Mo of up to 1,000 mg l-1 did not activate a physiological or cellular stress response in fish. Information from this study suggests that Mo water quality guidelines for the protection of aquatic life are highly protective of freshwater fish, namely rainbow trout. PMID:25629693

  1. Effects of elevated temperature and cadmium exposure on stress protein response in eastern oysters Crassostrea virginica (Gmelin)

    Energy Technology Data Exchange (ETDEWEB)

    Ivanina, A.V. [Department of Biology, University of North Carolina at Charlotte, 9201 University City Blvd., Charlotte, NC 28223 (United States); Taylor, C. [Department of Biology, University of North Carolina at Charlotte, 9201 University City Blvd., Charlotte, NC 28223 (United States); Johnson C. Smith University, 100 Beatties Ford Rd., Charlotte, NC 28216 (United States); Sokolova, I.M. [Department of Biology, University of North Carolina at Charlotte, 9201 University City Blvd., Charlotte, NC 28223 (United States)], E-mail: isokolov@uncc.edu

    2009-02-19

    Stress proteins such as heat shock proteins (HSPs) and metallothioneins (MTs) play a key role in cellular protection against environmental stress. Marine ectotherms such as eastern oysters Crassostrea virginica are commonly exposed to multiple stressors including temperature and pollution by metals such as cadmium (Cd) in estuaries and coastal zones; however, the combined effects of these stressors on their cellular protection mechanisms are poorly understood. We acclimated C. virginica from populations adapted to different thermal regimes (Washington, North Carolina and Texas) at a common temperature of 12 deg. C, and analyzed their expression of MTs and HSPs (cytosolic HSP69, HSC72-77, HSP90 and mitochondrial HSP60) in response to the combined acute temperature stress and long-term Cd exposure. Overall, HSP and MT induction patterns were similar in oysters from the three studied geographically distant populations. HSP69 and MTs were significantly up-regulated by Cd and temperature stress implying their important role in cellular stress protection. In contrast, HSC72-77, HSP60 and HSP90 were not consistently induced by either acute heat or Cd exposure. The induction temperature for MTs was higher than for HSP69 (>28 deg. C vs. 20 deg. C, respectively), and MTs were more strongly induced by Cd than by temperature stress (to up to 38-94-fold compared by 3.5-7.5-fold, respectively) consistent with their predominant role in metal detoxification. Notably, heat stress did not result in an additional increase in metallothionein expression in Cd-exposed oysters suggesting a capacity limitation during the combined exposure to Cd and temperature stress. Levels of HSP69 and in some cases, HSC72-77 and HSP90 were lower in Cd-exposed oysters as compared to their control counterparts during heat stress indicating that simultaneous exposure to these two stressors may have partially suppressed the cytoprotective upregulation of molecular chaperones. These limitations of stress

  2. Differences in Susceptibility to Heat Stress along the Chicken Intestine and the Protective Effects of Galacto-Oligosaccharides.

    Directory of Open Access Journals (Sweden)

    Soheil Varasteh

    Full Text Available High ambient temperatures negatively affect the human well-being as well as animal welfare and production. The gastrointestinal tract is predominantly responsive to heat stress. The currently available information about the multifaceted response to heat stress within different parts of the intestine is limited, especially in avian species. Hence, this study aims to evaluate the heat stress-induced sequence of events in the intestines of chickens. Furthermore, the gut health-promoting effect of dietary galacto-oligosaccharides (GOS was investigated in these heat stress-exposed chickens. Chickens were fed a control diet or diet supplemented with 1% or 2.5% GOS (6 days prior to and during a temperature challenge for 5 days (38-39°C, 8h per day. The parameters measured in different parts of the intestines included the genes (qPCR HSF1, HSF3, HSP70, HSP90, E-cadherin, claudin-1, claudin-5, ZO-1, occludin, TLR-2, TLR-4, IL-6, IL-8, HO-1, HIF-1α and their associated proteins HSP70, HSP90 and pan-cadherin (western blots. In addition, IL-6 and IL-8 plasma concentrations were measured by ELISA. In the jejunum, HSF3, HSP70, HSP90, E-cadherin, claudin-5, ZO-1, TLR-4, IL-6 and IL-8 mRNA expression and HSP70 protein expression were increased after heat stress exposure and a more pronounced increase in gene expression was observed in ileum after heat stress exposure, and in addition HSF1, claudin-1 and HIF-1α mRNA levels were upregulated. Furthermore, the IL-8 plasma levels were decreased in chickens exposed to heat stress. Interestingly, the heat stress-related effects in the jejunum were prevented in chickens fed a GOS diet, while dietary GOS did not alter these effects in ileum. In conclusion, our results demonstrate the differences in susceptibility to heat stress along the intestine, where the most obvious modification in gene expression is observed in ileum, while dietary GOS only prevent the heat stress-related changes in jejunum.

  3. The Live Access Server Scientific Product Generation Through Workflow Orchestration

    Science.gov (United States)

    Hankin, S.; Calahan, J.; Li, J.; Manke, A.; O'Brien, K.; Schweitzer, R.

    2006-12-01

    The Live Access Server (LAS) is a well-established Web-application for display and analysis of geo-science data sets. The software, which can be downloaded and installed by anyone, gives data providers an easy way to establish services for their on-line data holdings, so their users can make plots; create and download data sub-sets; compare (difference) fields; and perform simple analyses. Now at version 7.0, LAS has been in operation since 1994. The current "Armstrong" release of LAS V7 consists of three components in a tiered architecture: user interface, workflow orchestration and Web Services. The LAS user interface (UI) communicates with the LAS Product Server via an XML protocol embedded in an HTTP "get" URL. Libraries (APIs) have been developed in Java, JavaScript and perl that can readily generate this URL. As a result of this flexibility it is common to find LAS user interfaces of radically different character, tailored to the nature of specific datasets or the mindset of specific users. When a request is received by the LAS Product Server (LPS -- the workflow orchestration component), business logic converts this request into a series of Web Service requests invoked via SOAP. These "back- end" Web services perform data access and generate products (visualizations, data subsets, analyses, etc.). LPS then packages these outputs into final products (typically HTML pages) via Jakarta Velocity templates for delivery to the end user. "Fine grained" data access is performed by back-end services that may utilize JDBC for data base access; the OPeNDAP "DAPPER" protocol; or (in principle) the OGC WFS protocol. Back-end visualization services are commonly legacy science applications wrapped in Java or Python (or perl) classes and deployed as Web Services accessible via SOAP. Ferret is the default visualization application used by LAS, though other applications such as Matlab, CDAT, and GrADS can also be used. Other back-end services may include generation of Google

  4. Semantic orchestration of image processing services for environmental analysis

    Science.gov (United States)

    Ranisavljević, Élisabeth; Devin, Florent; Laffly, Dominique; Le Nir, Yannick

    2013-09-01

    In order to analyze environmental dynamics, a major process is the classification of the different phenomena of the site (e.g. ice and snow for a glacier). When using in situ pictures, this classification requires data pre-processing. Not all the pictures need the same sequence of processes depending on the disturbances. Until now, these sequences have been done manually, which restricts the processing of large amount of data. In this paper, we present how to realize a semantic orchestration to automate the sequencing for the analysis. It combines two advantages: solving the problem of the amount of processing, and diversifying the possibilities in the data processing. We define a BPEL description to express the sequences. This BPEL uses some web services to run the data processing. Each web service is semantically annotated using an ontology of image processing. The dynamic modification of the BPEL is done using SPARQL queries on these annotated web services. The results obtained by a prototype implementing this method validate the construction of the different workflows that can be applied to a large number of pictures.

  5. Perception of orchestral musicians about work environment and conditions

    Directory of Open Access Journals (Sweden)

    Clarissa Stefani Teixeira

    2014-04-01

    Full Text Available The objective of this study was to investigate the perception of 11 orchestral string (viola and violin musicians of both genders with respect to their work environment and conditions. We applied a questionnaire with demographic information and the scale Profile of Work Environment and Working Conditions by Nahas et al. (2009, which analyzes the following components: physical environment, social environment, development and professional achievement, salary and benefits, and social relevance. The social environment component presented the highest score - 8.00 (1.50 points, followed by professional achievement - 7.11 (1.96 points, and physical environment - 6.89 (0.93 points. The salary and benefits provided by the orchestra presented the lowest score - 6.78 (1.56 points. In general, the musicians showed positive perceptions of the components related to work environment and working conditions. However, remuneration and social relevance are work bases that could contribute to improve the working conditions of these professionals.

  6. Orchestrated structure evolution: modeling growth-regulated nanomanufacturing

    Energy Technology Data Exchange (ETDEWEB)

    Abbasi, Shaghayegh; Boehringer, Karl F [Department of Electrical Engineering, University of Washington, Seattle, WA 98195-2500 (United States); Kitayaporn, Sathana; Schwartz, Daniel T, E-mail: karlb@washington.edu [Department of Chemical Engineering, University of Washington, Seattle, WA 98195-2500 (United States)

    2011-04-22

    Orchestrated structure evolution (OSE) is a scalable manufacturing method that combines the advantages of top-down (tool-directed) and bottom-up (self-propagating) approaches. The method consists of a seed patterning step that defines where material nucleates, followed by a growth step that merges seeded islands into the final patterned thin film. We develop a model to predict the completed pattern based on a computationally efficient approximate Green's function solution of the diffusion equation plus a Voronoi diagram based approach that defines the final grain boundary structure. Experimental results rely on electron beam lithography to pattern the seeds, followed by the mass transfer limited growth of copper via electrodeposition. The seed growth model is compared with experimental results to quantify nearest neighbor seed-to-seed interactions as well as how seeds interact with the pattern boundary to impact the local growth rate. Seed-to-seed and seed-to-pattern interactions are shown to result in overgrowth of seeds on edges and corners of the shape, where seeds have fewer neighbors. We explore how local changes to the seed location can be used to improve the patterning quality without increasing the manufacturing cost. OSE is shown to enable a unique set of trade-offs between the cost, time, and quality of thin film patterning.

  7. Predicting protein amidation sites by orchestrating amino acid sequence features

    Science.gov (United States)

    Zhao, Shuqiu; Yu, Hua; Gong, Xiujun

    2017-08-01

    Amidation is the fourth major category of post-translational modifications, which plays an important role in physiological and pathological processes. Identifying amidation sites can help us understanding the amidation and recognizing the original reason of many kinds of diseases. But the traditional experimental methods for predicting amidation sites are often time-consuming and expensive. In this study, we propose a computational method for predicting amidation sites by orchestrating amino acid sequence features. Three kinds of feature extraction methods are used to build a feature vector enabling to capture not only the physicochemical properties but also position related information of the amino acids. An extremely randomized trees algorithm is applied to choose the optimal features to remove redundancy and dependence among components of the feature vector by a supervised fashion. Finally the support vector machine classifier is used to label the amidation sites. When tested on an independent data set, it shows that the proposed method performs better than all the previous ones with the prediction accuracy of 0.962 at the Matthew's correlation coefficient of 0.89 and area under curve of 0.964.

  8. Automation Hooks Architecture Trade Study for Flexible Test Orchestration

    Science.gov (United States)

    Lansdowne, Chatwin A.; Maclean, John R.; Graffagnino, Frank J.; McCartney, Patrick A.

    2010-01-01

    We describe the conclusions of a technology and communities survey supported by concurrent and follow-on proof-of-concept prototyping to evaluate feasibility of defining a durable, versatile, reliable, visible software interface to support strategic modularization of test software development. The objective is that test sets and support software with diverse origins, ages, and abilities can be reliably integrated into test configurations that assemble and tear down and reassemble with scalable complexity in order to conduct both parametric tests and monitored trial runs. The resulting approach is based on integration of three recognized technologies that are currently gaining acceptance within the test industry and when combined provide a simple, open and scalable test orchestration architecture that addresses the objectives of the Automation Hooks task. The technologies are automated discovery using multicast DNS Zero Configuration Networking (zeroconf), commanding and data retrieval using resource-oriented Restful Web Services, and XML data transfer formats based on Automatic Test Markup Language (ATML). This open-source standards-based approach provides direct integration with existing commercial off-the-shelf (COTS) analysis software tools.

  9. The nucleosome: orchestrating DNA damage signaling and repair within chromatin.

    Science.gov (United States)

    Agarwal, Poonam; Miller, Kyle M

    2016-10-01

    DNA damage occurs within the chromatin environment, which ultimately participates in regulating DNA damage response (DDR) pathways and repair of the lesion. DNA damage activates a cascade of signaling events that extensively modulates chromatin structure and organization to coordinate DDR factor recruitment to the break and repair, whilst also promoting the maintenance of normal chromatin functions within the damaged region. For example, DDR pathways must avoid conflicts between other DNA-based processes that function within the context of chromatin, including transcription and replication. The molecular mechanisms governing the recognition, target specificity, and recruitment of DDR factors and enzymes to the fundamental repeating unit of chromatin, i.e., the nucleosome, are poorly understood. Here we present our current view of how chromatin recognition by DDR factors is achieved at the level of the nucleosome. Emerging evidence suggests that the nucleosome surface, including the nucleosome acidic patch, promotes the binding and activity of several DNA damage factors on chromatin. Thus, in addition to interactions with damaged DNA and histone modifications, nucleosome recognition by DDR factors plays a key role in orchestrating the requisite chromatin response to maintain both genome and epigenome integrity.

  10. Slow oscillations orchestrating fast oscillations and memory consolidation.

    Science.gov (United States)

    Mölle, Matthias; Born, Jan

    2011-01-01

    Slow-wave sleep (SWS) facilitates the consolidation of hippocampus-dependent declarative memory. Based on the standard two-stage memory model, we propose that memory consolidation during SWS represents a process of system consolidation which is orchestrated by the neocortical memory. The slow oscillations temporally group neuronal activity into up-states of strongly enhanced neuronal activity and down-states of neuronal silence. In a feed-forward efferent action, this grouping is induced not only in the neocortex but also in other structures relevant to consolidation, namely the thalamus generating 10-15Hz spindles, and the hippocampus generating sharp wave-ripples, with the latter well known to accompany a replay of newly encoded memories taking place in hippocampal circuitries. The feed-forward synchronizing effect of the slow oscillation enables the formation of spindle-ripple events where ripples and accompanying reactivated hippocampal memory information become nested into the single troughs of spindles. Spindle-ripple events thus enable reactivated memory-related hippocampal information to be fed back to neocortical networks in the excitable slow oscillation up-state where they can induce enduring plastic synaptic changes underlying the effective formation of long-term memories. Copyright © 2011 Elsevier B.V. All rights reserved.

  11. Stress

    Science.gov (United States)

    ... can be life-saving. But chronic stress can cause both physical and mental harm. There are at least three different types of stress: Routine stress related to the pressures of work, family, and other daily responsibilities Stress brought about ...

  12. Differential gene expressions in testes of L2 strain Taiwan country chicken in response to acute heat stress.

    Science.gov (United States)

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

    2013-01-15

    Acute heat stress affects genes involved in spermatogenesis in mammals. However, there is apparently no elaborate research on the effects of acute heat stress on gene expression in avian testes. The purpose of this study was to investigate global gene expression in testes of the L2 strain of Taiwan country chicken after acute heat stress. Twelve roosters, 45 weeks old, 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-hour recovery, and with 6-hour recovery, respectively. Testis samples were collected for RNA isolation and microarray analysis. Based on gene expression profiles, 169 genes were upregulated and 140 genes were downregulated after heat stress using a cutoff value of twofold or greater change. Based on gene ontology analysis, differentially expressed genes were mainly related to response to stress, transport, signal transduction, and metabolism. A functional network analysis displayed that heat shock protein genes and related chaperones were the major upregulated groups in chicken testes after acute heat stress. A quantitative real-time polymerase chain reaction analysis of mRNA expressions of HSP70, HSP90AA1, BAG3, SERPINB2, HSP25, DNAJA4, CYP3A80, CIRBP, and TAGLN confirmed the results of the microarray analysis. Because the HSP genes (HSP25, HSP70, and HSP90AA1) and the antiapoptotic BAG3 gene were dramatically altered in heat-stressed chicken testes, we concluded that these genes were important factors in the avian testes under acute heat stress. Whether these genes could be candidate genes for thermotolerance in roosters requires further investigation. Copyright © 2013 Elsevier Inc. All rights reserved.

  13. Formal Model of Web Service Composition: An Actor-Based Approach to Unifying Orchestration and Choreography

    OpenAIRE

    Wang, Yong

    2013-01-01

    Web Service Composition creates new composite Web Services from the collection of existing ones to be composed further and embodies the added values and potential usages of Web Services. Web Service Composition includes two aspects: Web Service orchestration denoting a workflow-like composition pattern and Web Service choreography which represents an aggregate composition pattern. There were only a few works which give orchestration and choreography a relationship. In this paper, we introduce...

  14. Joining Jolie to Docker - Orchestration of Microservices on a Containers-as-a-Service Layer

    OpenAIRE

    Giaretta, Alberto; Dragoni, Nicola; Mazzara, Manuel

    2017-01-01

    Cloud computing is steadily growing and, as IaaS vendors have started to offer pay-as-you-go billing policies, it is fundamental to achieve as much elasticity as possible, avoiding over-provisioning that would imply higher costs. In this paper, we briefly analyse the orchestration characteristics of PaaSSOA, a proposed architecture already implemented for Jolie microservices, and Kubernetes, one of the various orchestration plugins for Docker; then, we outline similarities and differences of ...

  15. Orchestrating Shots for the National Ignition Facility (NIF)

    International Nuclear Information System (INIS)

    Mathisen, D G; Bettenhausen, R C; Beeler, R G; Bowers, G A; Carey, R W; Casavant, D D; Cline, B D; Demaret, R D; Domyancic, D M; Elko, S D; Fisher, J M; Krammen, J E; Lagin, L J; Ludwigsen, A P; Patterson, R W; Sanchez, R J; Stout, E A

    2005-01-01

    The National Ignition Facility (NIF), currently under construction at the Lawrence Livermore National Laboratory, is a stadium-sized facility containing a 192-beam, 1.8 Megajoule, 500-Terawatt, ultra-violet laser system together with a 10-meter diameter target chamber with room for nearly 100 experimental diagnostics. When completed, NIF will be the world's largest and most energetic laser experimental system, providing an international center to study inertial confinement fusion and physics of matter at extreme densities and pressures. The NIF is operated by the Integrated Computer Control System (ICCS), which is a layered architecture of over 700 lower-level front-end processors attached to nearly 60,000 control points and coordinated by higher-level supervisory subsystems in the main control room. A shot automation framework has been developed and deployed during the past year to orchestrate and automate shots performed at the NIF using the ICCS. The Shot Automation framework is designed to automate 4-8 hour shot sequences, that includes deriving shot goals from an experiment definition, set up of the laser and diagnostics, automatic alignment of laser beams, and a countdown to charge and fire the lasers. These sequences consist of set of preparatory verification shots, leading to amplified system shots followed by post-shot analysis and archiving. The framework provides for a flexible, model-based work-flow execution, driven by scripted automation called macro steps. The shot director software is the orchestrating component of a very flexible automation layer which allows us to define, coordinate and reuse simpler automation sequences. This software provides a restricted set of shot life cycle state transitions to 26 collaboration supervisors that automate 8-laser beams (bundle) and a common set of shared resources. Each collaboration supervisor commands approximately 10 subsystem shot supervisors that perform automated control and status verification

  16. Reactive oxygen species (ROS) and the heat stress response of Daphnia pulex: ROS-mediated activation of hypoxia-inducible factor 1 (HIF-1) and heat shock factor 1 (HSF-1) and the clustered expression of stress genes.

    Science.gov (United States)

    Klumpen, Eva; Hoffschröer, Nadine; Zeis, Bettina; Gigengack, Ulrike; Dohmen, Elias; Paul, Rüdiger J

    2017-01-01

    Heat stress in ectotherms involves direct (e.g. protein damage) and/or indirect effects (temperature-induced hypoxia and ROS formation), which cause activation of the transcription factors (TF) heat shock factor 1 (HSF-1) and/or hypoxia-inducible factor 1 (HIF-1). The present study focused on the links between stress (ROS) signals, nuclear (n) and cytoplasmic (c) HSF-1/HIF-1 levels, and stress gene expression on mRNA and protein levels (e.g. heat-shock protein 90, HSP90) upon acute heat and ROS (H 2 O 2 ) stress. Acute heat stress (30°C) evoked fluctuations in ROS level. Different feeding regimens, which affected the glutathione (GSH) level, allowed altering the frequency of ROS fluctuations. Other data showed fluctuation frequency to depend also on ROS production rate. The heat-induced slow or fast ROS fluctuations (at high or low GSH levels) evoked slow or fast fluctuations in the levels of nHIF-1α, nHSF-1 and gene products (mRNAs and protein), albeit after different time delays. Time delays to ROS fluctuations were, for example,shorter for nHIF-1α than for nHSF-1 fluctuations, and nHIF-1α fluctuations preceded and nHSF-1 fluctuations followed fluctuations in HSP90 mRNA level. Cytoplasmic TF levels either changed little (cHIF-1α) or showed a steady increase (cHSF-1). Applying acute H 2 O 2 stress (at 20°C) revealed effects on nHIF-1α and mRNA levels, but no significant effects on nHSF-1 level. Transcriptome data additionally showed coordinated fluctuations of mRNA levels upon acute heat stress, involving mRNAs for HSPs and other stress proteins, with all corresponding genes carrying DNA binding motifs for HIF-1 and HSF-1. This study provided evidence for promoting effects of ROS and HIF-1 on early haemoglobin, HIF-1α and HSP90 mRNA expressions upon heat or ROS stress. The increasing cHSF-1 level likely affected nHSF-1 level and later HSP90 mRNA expression. Heat stress evoked ROS fluctuations, with this stress signal forwarded via nHIF-1 and nHSF-1

  17. Design of Service Net based Correctness Verification Approach for Multimedia Conferencing Service Orchestration

    Directory of Open Access Journals (Sweden)

    Cheng Bo

    2012-02-01

    Full Text Available Multimedia conferencing is increasingly becoming a very important and popular application over Internet. Due to the complexity of asynchronous communications and handle large and dynamically concurrent processes for multimedia conferencing, which confront relevant challenge to achieve sufficient correctness guarantees, and supporting the effective verification methods for multimedia conferencing services orchestration is an extremely difficult and challenging problem. In this paper, we firstly present the Business Process Execution Language (BPEL based conferencing service orchestration, and mainly focus on the service net based correction verification approach for multimedia conferencing services orchestration, which can automatically translated the BPEL based service orchestration into a corresponding Petri net model with the Petri Net Markup Language (PNML, and also present the BPEL service net reduction rules and multimedia conferencing service orchestration correction verification algorithms. We perform the correctness analysis and verification using the service net properties as safeness, reachability and deadlocks, and also provide an automated support tool for the formal analysis and soundness verification for the multimedia conferencing services orchestration scenarios. Finally, we give the comparison and evaluations.

  18. Antarctic krill 454 pyrosequencing reveals chaperone and stress transcriptome.

    Directory of Open Access Journals (Sweden)

    Melody S Clark

    Full Text Available BACKGROUND: The Antarctic krill Euphausia superba is a keystone species in the Antarctic food chain. Not only is it a significant grazer of phytoplankton, but it is also a major food item for charismatic megafauna such as whales and seals and an important Southern Ocean fisheries crop. Ecological data suggest that this species is being affected by climate change and this will have considerable consequences for the balance of the Southern Ocean ecosystem. Hence, understanding how this organism functions is a priority area and will provide fundamental data for life history studies, energy budget calculations and food web models. METHODOLOGY/PRINCIPAL FINDINGS: The assembly of the 454 transcriptome of E. superba resulted in 22,177 contigs with an average size of 492bp (ranging between 137 and 8515bp. In depth analysis of the data revealed an extensive catalogue of the cellular chaperone systems and the major antioxidant proteins. Full length sequences were characterised for the chaperones HSP70, HSP90 and the super-oxide dismutase antioxidants, with the discovery of potentially novel duplications of these genes. The sequence data contained 41,470 microsatellites and 17,776 Single Nucleotide Polymorphisms (SNPs/INDELS, providing a resource for population and also gene function studies. CONCLUSIONS: This paper details the first 454 generated data for a pelagic Antarctic species or any pelagic crustacean globally. The classical "stress proteins", such as HSP70, HSP90, ferritin and GST were all highly expressed. These genes were shown to be over expressed in the transcriptomes of Antarctic notothenioid fish and hypothesized as adaptations to living in the cold, with the associated problems of decreased protein folding efficiency and increased vulnerability to damage by reactive oxygen species. Hence, these data will provide a major resource for future physiological work on krill, but in particular a suite of "stress" genes for studies understanding

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

    Science.gov (United States)

    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.

  20. Injury and the orchestral environment: part II. Organisational culture, behavioural norms, and attitudes to injury.

    Science.gov (United States)

    Rickert, Dale Ll; Barrett, Margaret S; Ackermann, Bronwen J

    2014-06-01

    The organisational culture, behavioural norms, and attitudes of a workplace have a profound influence on levels of injury and illness amongst its workers. While this is well established in Work Health and Safety literature, very little research has attempted to understand the influence of organisational culture on injury risk in the orchestral profession. To address this, the current study aimed to investigate the influence of organisational culture on injury outcomes for orchestral musicians. Using a qualitative case study methodology, in-depth semi-structured interviews were undertaken with 10 professional orchestral cellists (2 freelance and 8 fulltime members) from a single Australian orchestra. After initial data analysis, further interviews were undertaken with a set of 5 orchestral management staff as a means of data triangulation. All data were analysed using a themes-based "analysis of narrative" approach. The findings indicate that an orchestral culture exists in which musicians see injury as a sign of weakness, failure, and poor musicianship. Such negative perceptions of injury influence musicians to play through considerable levels of pain and continue performing with injuries. Because of perceived judgment from the orchestral group, musicians were found to conceal injuries from colleagues and management staff. Freelance musicians felt that disclosing injuries may lead to decreased work opportunities, and both full-time and casual musicians felt that "opening up" about injury may subject them to group judgment about their technique or musicianship. The study suggests education measures which may be effective at influencing individual behaviours and attitudes as well as cultural change initiatives which could lead to long-term positive health outcomes in the orchestral workplace.

  1. Understanding how discrete populations of hypothalamic neurons orchestrate complicated behavioral states

    Directory of Open Access Journals (Sweden)

    Allison eGraebner

    2015-08-01

    Full Text Available A major question in systems neuroscience is how a single population of neurons can interact with the rest of the brain to orchestrate complex behavioral states. The hypothalamus contains many such discrete neuronal populations that individually regulate arousal, feeding, and drinking. For example, hypothalamic neurons that express hypocretin (Hcrt neuropeptides can sense homeostatic and metabolic factors affecting wakefulness and orchestrate organismal arousal. Neurons that express agouti-related protein (AgRP can sense the metabolic needs of the body and orchestrate a state of hunger. The organum vasculosum of the lamina terminalis (OVLT can detect the hypertonicity of blood and orchestrate a state of thirst. Each hypothalamic population is sufficient to generate complicated behavioral states through the combined efforts of distinct efferent projections. The principal challenge to understanding these brain systems is therefore to determine the individual roles of each downstream projection for each behavioral state. In recent years, the development and application of temporally precise, genetically encoded tools have greatly improved our understanding of the structure and function of these neural systems. This review will survey recent advances in our understanding of how these individual hypothalamic populations can orchestrate complicated behavioral states due to the combined efforts of individual downstream projections.

  2. Sound Practice– Improving occupational health and safety for professional orchestral musicians in Australia

    Directory of Open Access Journals (Sweden)

    Bronwen Jane Ackermann

    2014-09-01

    Full Text Available The Sound Practice Project is a five-year study involving baseline evaluation, development and implementation of musician-specific work health and safety initiatives. A cross-sectional population physical and psychological survey and physical assessment were conducted at the same time, with an auditory health assessment conducted later. The results were used to guide the development of a series of targeted interventions, encompassing physical, psychological and auditory health components. This paper provides an overview of the project but focuses on the health findings arising from the cross-sectional survey.377 musicians from the eight professional symphony orchestras in Australia took part in the cross-sectional study (about 70% of eligible musicians. Eighty-four percent (84% of musicians reported past performance-related musculoskeletal disorder (PRMD episodes; 50% were suffering a current PRMD. Of the 63% who returned hearing surveys, 43% believed they had hearing loss, and 64% used earplugs at least intermittently. Noise exposure was found to be high in private practice, although awareness of risk and earplug use in this environment was lower than in orchestral settings. Improved strategic approaches, acoustic screens and recently developed active earplugs were found to provide effective new options for hearing protection. With respect to psychosocial screening, female musicians reported significantly more trait anxiety, music performance anxiety, social anxiety, and other forms of anxiety and depression than male musicians. The youngest musicians were significantly more anxious compared with the oldest musicians. Thirty-three percent (33% of musicians may meet criteria for a diagnosis of social phobia; 32% returned a positive depression screen and 22% for post-traumatic stress disorder. PRMDs and trigger point discomfort levels were strongly associated with increasing severity of psychological issues such as depression and music

  3. Stress

    Science.gov (United States)

    ... taking care of an aging parent. With mental stress, the body pumps out hormones to no avail. Neither fighting ... with type 1 diabetes. This difference makes sense. Stress blocks the body from releasing insulin in people with type 2 ...

  4. Effect of patchouli alcohol on the regulation of heat shock-induced oxidative stress in IEC-6 cells.

    Science.gov (United States)

    Liu, Xiaoxi; Jiang, Linshu; Liu, Fenghua; Chen, Yuping; Xu, Lei; Li, Deyin; Ma, Yunfei; Li, Huanrong; Xu, Jianqin

    2016-08-01

    Purpose Patchouli alcohol (PA) is used to treat gastrointestinal dysfunction. The purpose of this study was to ascertain the function of PA in the regulated process of oxidative stress in rat intestinal epithelial cells (IEC-6). Materials and methods Oxidative stress was stimulated by exposing IEC-6 cells to heat shock (42 °C for 3 h). IEC-6 cells in treatment groups were pretreated with various concentrations of PA (10, 40, and 80 ng/mL) for 3 h before heat shock. Results Heat shock caused damage to the morphology of IEC-6 cells, and increased reactive oxygen species (ROS) level and malondialdehyde (MDA) content. Moreover, mRNA and protein expression by target genes related to oxidative stress in heat shock were also altered. Specifically, the mRNA expression by HSP70, HSP90, GSH-px, NRF2 nd HO-1were all increased, and Nrf2 and Keap1 protein expression were increased after heat shock. However, pretreatment with PA weakened the level of damage to the cellular morphology, and decreased the MDA content caused by heat shock, indicating PA had cytoprotective activities. Pretreatment with PA at high dose significantly increased generation of intracellular ROS. Compared with the heat shock group alone, PA pretreatment significantly decreased the mRNA expression by HSP70, HSP90, SOD, CAT, GSH-px, KEAP1 and HO-1. Furthermore, the high dose of PA significantly increased Nrf2 protein expression, while both the intermediate and high dose of PA significantly increased HO-1 protein expression. Conclusion Heat-shock-induced oxidative stress in IEC-6 cells, and PA could alleviate the Nrf2-Keap1 cellular oxidative stress responses.

  5. Local Inhibition of HSP90 to Prevent Intimal Hyperplasia after Balloon Injury

    Science.gov (United States)

    2017-10-01

    IACUC documents. Cell culture and viability testing, optimizing proliferation and migration assays, perfecting microRNA extraction, perfecting cDNA ...4) qrtPCR: cDNA was prepared using the High Capacity cDNA Revers Transcription kit (Applied Biosystems. Grand Island, NY). qPCR was performed...and actions or plans to resolve them  We have had some difficulty in optimizing the protocol for creating cDNA from MicroRNA. We have tried three

  6. STTEPping in the Right Direction? Western Classical Music in an Orchestral Programme for Disadvantaged African Youth

    Science.gov (United States)

    van Niekerk, Caroline; Salminen, Sanna

    2008-01-01

    This article looks at STTEP, an outreach project currently housed at the University of Pretoria, which concentrates on the teaching of western orchestral instruments, plus background areas such as music theory, to disadvantaged children and youth from a variety of townships around Pretoria, South Africa. STTEP's direction can well be described as…

  7. Kindergarten Teachers' Orchestration of Mathematical Activities Afforded by Technology: Agency and Mediation

    Science.gov (United States)

    Carlsen, Martin; Erfjord, Ingvald; Hundeland, Per Sigurd; Monaghan, John

    2016-01-01

    This paper focuses on kindergarten teachers' interactions with young children during mathematical learning activities involving the use of digital tools. We aim to characterise the teachers' roles and actions in these activities and extend considerations of teachers' orchestrations current in the research literature with regard to agency and…

  8. Multimodal Teaching Analytics: Automated Extraction of Orchestration Graphs from Wearable Sensor Data

    Science.gov (United States)

    Prieto, L. P.; Sharma, K.; Kidzinski, L.; Rodríguez-Triana, M. J.; Dillenbourg, P.

    2018-01-01

    The pedagogical modelling of everyday classroom practice is an interesting kind of evidence, both for educational research and teachers' own professional development. This paper explores the usage of wearable sensors and machine learning techniques to automatically extract orchestration graphs (teaching activities and their social plane over time)…

  9. Orchestrating innovation networks: The case of innovation brokers in the agri-food sector

    NARCIS (Netherlands)

    Batterink, M.H.; Wubben, E.F.M.; Klerkx, L.W.A.; Omta, S.W.F.

    2010-01-01

    This explorative study of network orchestration processes conducted by innovation brokers addresses new issues in bridging small and medium-sized enterprises (SMEs) and research institutes in innovation networks. The study includes four in-depth case studies in the agri-food sector from different

  10. Orchestrating the management of patients with high-output stomas.

    Science.gov (United States)

    McDonald, Alison

    Working in isolation, managing high-output stomas can be stressful and difficult, with patient outcomes varying significantly. For the stoma care clinical nurse specialist, managing the choice of stoma appliance is only a small part of the care provided. To standardise and improve outcomes for patients with high-output stomas, team working is required. After contacting other stoma care services and using guidance from the High Impact Actions for Stoma Care document ( Coloplast, 2010 ), it was evident that the team should put together an algorithm/flow chart to guide both specialists and ward nursing staff in the evidence-based and standardised management of patients with high-output stomas. This article presents the flowchart that was produced and uses case studies to demonstrate improvements.

  11. Cloning, purification and characterization of a 90kDa heat shock protein from Citrus sinensis (sweet orange).

    Science.gov (United States)

    Mendonça, Yuri A; Ramos, Carlos H I

    2012-01-01

    Protein misfolding is stimulated by stress, such as heat, and heat shock proteins (Hsps) are the first line of defense against these undesirable situations. Plants, which are naturally sessile, are perhaps more exposed to stress factors than some other organisms, and consequently, the role of Hsps is crucial to maintain homeostasis. Hsp90, because of its key role in infection and other stresses, is targeted in therapies that improve plant production by increasing resistance to both biotic and abiotic stress. In addition, Hsp90 is a primary factor in the maintenance of homeostasis in plants. Therefore, we cloned and purified Hsp90 from Citrus sinensis (sweet orange). Recombinant C. sinensis Hsp90 (rCsHsp90) was produced and measured by circular dichroism (CD), intrinsic fluorescence spectroscopy and dynamic light scattering. rCsHsp90 formed a dimer in solution with a Stokes radius of approximately 62Å. In addition, it was resistant to thermal unfolding, was able to protect citrate synthase from aggregation, and Western blot analysis demonstrated that CsHsp90 was constitutively expressed in C. sinensis cells. Our analysis indicated that CsHsp90 is conformationally similar to that of yeast Hsp90, for which structural information is available. Therefore, we showed that C. sinensis expresses an Hsp90 chaperone that has a conformation and function similar to other Hsp90s. Copyright © 2011 Elsevier Masson SAS. All rights reserved.

  12. Stress

    DEFF Research Database (Denmark)

    Keller, Hanne Dauer

    2015-01-01

    Kapitlet handler om stress som følelse, og det trækker primært på de få kvalitative undersøgelser, der er lavet af stressforløb.......Kapitlet handler om stress som følelse, og det trækker primært på de få kvalitative undersøgelser, der er lavet af stressforløb....

  13. Stress !!!

    OpenAIRE

    Fledderus, M.

    2012-01-01

    Twee op de vijf UT-studenten hebben last van ernstige studiestress, zo erg zelfs dat het ze in hun privéleven belemmert. Die cijfers komen overeen met het landelijk beeld van stress onder studenten. Samen met 14 andere universiteits- en hogeschoolbladen enquêteerde UT Nieuws bijna 5500 studenten. Opvallend is dat mannelijke studenten uit Twente zich veel minder druk lijken te maken over hun studie. Onder vrouwen ligt de stress juist erg hoog ten opzichte van het landelijk gemiddelde.

  14. Ameliorative effect of vitamin E on hepatic oxidative stress and hypoimmunity induced by high-fat diet in turbot (Scophthalmus maximus).

    Science.gov (United States)

    Jia, Yudong; Jing, Qiqi; Niu, Huaxin; Huang, Bin

    2017-08-01

    This study was conducted to examine the effects of vitamin E on growth performance, oxidative stress and non-specific immunity of turbot (Scophthalmus maximus) fed with high-fat diet. Results showed that high-fat diet significantly increased hepatosomatic index, viscerosomatic index, hepatic malondialdehyde level and decreased catalase and superoxide dismutase activities, whereas final weight, specific growth rate and survival rate remained unchanged. Meanwhile, nitro blue tetrazolium positive leucocytes of head kidney, respiratory burst activity in head-kidney macrophage, phagocytic index and serum lysozyme activity were significantly reduced after feeding with high-fat diet. Furthermore, fish fed with high-fat diet promoted higher expression of heat shock protein (hsp70, hsp90), and inhibited expression of complement component 3 (c3) in the liver and tumor necrosis factor-α (tnf-α), interleukine 1β (il-1β), toll like receptor 22 (tlr-22) in the spleen and head-kidney, respectively. However, simultaneous supplementation with 480 mg kg -1 vitamin E protected turbot against high-fat diet-induced hepatic oxidative stress, hypoimmunity through attenuating lipid peroxidation, renewing antioxidant enzymes activities and nonspecific immune responses, and modulating the expression of stress protein (hsp70, hsp90) and immune-related genes (c3, tnf-α, il-1β, tlr-22). In conclusion, the obtained results indicate the vitamin E as a wildly used functional feed additive contributes potentially to alleviate high-fat diet-induced hepatic oxidative stress and hypoimmunity, maintain the health, and improve the broodstock management for turbot. Copyright © 2017 Elsevier Ltd. All rights reserved.

  15. DECENTRALIZED ORCHESTRATION OF COMPOSITE OGC WEB PROCESSING SERVICES IN THE CLOUD

    Directory of Open Access Journals (Sweden)

    F. Xiao

    2016-09-01

    Full Text Available Current web-based GIS or RS applications generally rely on centralized structure, which has inherent drawbacks such as single points of failure, network congestion, and data inconsistency, etc. The inherent disadvantages of traditional GISs need to be solved for new applications on Internet or Web. Decentralized orchestration offers performance improvements in terms of increased throughput and scalability and lower response time. This paper investigates build time and runtime issues related to decentralized orchestration of composite geospatial processing services based on OGC WPS standard specification. A case study of dust storm detection was demonstrated to evaluate the proposed method and the experimental results indicate that the method proposed in this study is effective for its ability to produce the high quality solution at a low cost of communications for geospatial processing service composition problem.

  16. Bio-jETI: a service integration, design, and provisioning platform for orchestrated bioinformatics processes.

    Science.gov (United States)

    Margaria, Tiziana; Kubczak, Christian; Steffen, Bernhard

    2008-04-25

    With Bio-jETI, we introduce a service platform for interdisciplinary work on biological application domains and illustrate its use in a concrete application concerning statistical data processing in R and xcms for an LC/MS analysis of FAAH gene knockout. Bio-jETI uses the jABC environment for service-oriented modeling and design as a graphical process modeling tool and the jETI service integration technology for remote tool execution. As a service definition and provisioning platform, Bio-jETI has the potential to become a core technology in interdisciplinary service orchestration and technology transfer. Domain experts, like biologists not trained in computer science, directly define complex service orchestrations as process models and use efficient and complex bioinformatics tools in a simple and intuitive way.

  17. A Dual-Promoter Gene Orchestrates the Sucrose-Coordinated Synthesis of Starch and Fructan in Barley

    Energy Technology Data Exchange (ETDEWEB)

    Jin, Yunkai; Fei, Mingliang; Rosenquist, Sara; Jin, Lu; Gohil, Suresh; Sandström, Corine; Olsson, Helena; Persson, Cecilia; Höglund, Anna-Stina; Fransson, Gunnel; Ruan, Ying; Åman, Per; Jansson, Christer; Liu, Chunlin; Andersson, Roger; Sun, Chuanxin

    2017-12-01

    Starch and fructan are two important carbohydrates in many flowering plants and in human diets. Understanding how plants allocate photosynthates and how they prioritize synthesis of different carbohydrates during development is essential in efforts to improve cereals for increased stress tolerance and for desirable carbohydrate compositions in food and feed. We report the coordinated synthesis of starch and fructan in barley, orchestrated by two functionally opposing transcription factors encoded from two alternative promoters, one intronic/exonic, harbored on a single gene. . This dual-transcription factor system employs an autoregulatory, antagonsitic mechanism in sensing sucrose at one promoter, potentially via sucrose/glucose/fructose/trehalose 6-phosphate signaling, and conduct a coordinated synthesis of starch and fructan synthesis by competitive transcription factor binding to the second promoter The finding of an intron/exon-spanning promoter in a hosting gene, resulting in proteins with distinct functions, contributes to our appreciation of the complexity of the plant genome As a case in point for the physiological role of the antagonistic transcription factor system, we have demonstrated that it can be exploited in breeding barley with tailored amounts of fructan for production of specialty food ingredients.

  18. Software Defined Resource Orchestration System for Multitask Application in Heterogeneous Mobile Cloud Computing

    Directory of Open Access Journals (Sweden)

    Qi Qi

    2016-01-01

    Full Text Available The mobile cloud computing (MCC that combines mobile computing and cloud concept takes wireless access network as the transmission medium and uses mobile devices as the client. When offloading the complicated multitask application to the MCC environment, each task executes individually in terms of its own computation, storage, and bandwidth requirement. Due to user’s mobility, the provided resources contain different performance metrics that may affect the destination choice. Nevertheless, these heterogeneous MCC resources lack integrated management and can hardly cooperate with each other. Thus, how to choose the appropriate offload destination and orchestrate the resources for multitask is a challenge problem. This paper realizes a programming resource provision for heterogeneous energy-constrained computing environments, where a software defined controller is responsible for resource orchestration, offload, and migration. The resource orchestration is formulated as multiobjective optimal problem that contains the metrics of energy consumption, cost, and availability. Finally, a particle swarm algorithm is used to obtain the approximate optimal solutions. Simulation results show that the solutions for all of our studied cases almost can hit Pareto optimum and surpass the comparative algorithm in approximation, coverage, and execution time.

  19. Architectural Principles for Orchestration of Cross-Organizational Service Delivery: Case Studies from the Netherlands

    Science.gov (United States)

    van Veenstra, Anne Fleur; Janssen, Marijn

    One of the main challenges for e-government is to create coherent services for citizens and businesses. Realizing Integrated Service Delivery (ISD) requires government agencies to collaborate across their organizational boundaries. The coordination of processes across multiple organizations to realize ISD is called orchestration. One way of achieving orchestration is to formalize processes using architecture. In this chapter we identify architectural principles for orchestration by looking at three case studies of cross-organizational service delivery chain formation in the Netherlands. In total, six generic principles were formulated and subsequently validated in two workshops with experts. These principles are: (i) build an intelligent front office, (ii) give processes a clear starting point and end, (iii) build a central workflow application keeping track of the process, (iv) differentiate between simple and complex processes, (v) ensure that the decision-making responsibility and the overview of the process are not performed by the same process role, and (vi) create a central point where risk profiles are maintained. Further research should focus on how organizations can adapt these principles to their own situation.

  20. An Orchestrating Evaluation of Complex Educational Technologies: a Case Study of a CSCL System

    Directory of Open Access Journals (Sweden)

    Luis P. Prieto

    2014-06-01

    Full Text Available As digital technologies permeate every aspect of our lives, the complexity of the educational settings, and of the technological support we use within them, unceasingly rises. This increased complexity, along with the need for educational practitioners to apply such technologies within multi-constraint authentic settings, has given rise to the notion of technology-enhanced learning practice as “orchestration of learning”. However, at the same time, the complexity involved in evaluating the benefits of such educational technologies has also increased, prompting questions about the way evaluators can cope with the different places, technologies, informants and issues involved in their evaluation activity. By proposing the notion of “orchestrating evaluation”, this paper tries to reconcile the often disparate “front office accounts” of research publications and the “shop floor practice” of evaluation of educational technology, through the case study of evaluating a system to help teachers in coordinating computer-supported collaborative learning (CSCL scenarios. We reuse an internationally-evaluated conceptual framework of “orchestration aspects” (design, management, adaptation, pragmatism, etc. to structure the case‟s narrative, showing how the original evaluation questions and methods were modulated in the face of the multiple (authentic evaluation setting constraints.

  1. Stress !!!

    NARCIS (Netherlands)

    Fledderus, M.

    2012-01-01

    Twee op de vijf UT-studenten hebben last van ernstige studiestress, zo erg zelfs dat het ze in hun privéleven belemmert. Die cijfers komen overeen met het landelijk beeld van stress onder studenten. Samen met 14 andere universiteits- en hogeschoolbladen enquêteerde UT Nieuws bijna 5500 studenten.

  2. SDN/NFV orchestration for dynamic deployment of virtual SDN controllers as VNF for multi-tenant optical networks

    OpenAIRE

    Muñoz, Raül; Vilalta, Ricard; Casellas, Ramon; Martínez, Ricardo; Szyrkowiec, T.; Autenrieth, A.; López, Víctor; López, D.

    2015-01-01

    We propose to virtualize the SDN control functions and move them to the cloud. We experimentally evaluate the first SDN/NFV orchestration architecture to dynamically deploy independent SDN controller instances for each deployed virtual optical network.

  3. Assessing Orchestrated Simulation Through Modeling to Quantify the Benefits of Unmanned-Teaming in a Tactical ASW Scenario

    Science.gov (United States)

    2018-03-01

    be used by the model to create pseudo- random behavior for agents. This process is a critical aspect of the simulation because it allows for the...ORCHESTRATED SIMULATION THROUGH MODELING TO QUANTIFY THE BENEFITS OF UNMANNED–MANNED TEAMING IN A TACTICAL ASW SCENARIO by Preston T. Tilus March...Leave blank) 2. REPORT DATE March 2018 3. REPORT TYPE AND DATES COVERED Master’s thesis 4. TITLE AND SUBTITLE ASSESSING ORCHESTRATED SIMULATION

  4. Neutrophil gelatinase-associated lipocalin (NGAL) and matrix metalloproteinases as novel stress markers in children and young adults on chronic dialysis.

    Science.gov (United States)

    Musiał, Kinga; Zwolińska, Danuta

    2011-03-01

    Phenomena related to chronic kidney disease, such as atherosclerosis, aggravate with the introduction of dialysis. Matrix metalloproteinases (MMP) and factors modifying their activity, such as their tissue inhibitors (TIMP) or neutrophil gelatinase-associated lipocalin (NGAL), take part in the matrix turnover and the endothelial damage characteristic for atherogenesis. However, there are no data on the associations between these parameters and other known pro-atherogenic factors, or on the impact of various dialysis modalities on them. The aim of our study was to assess the serum concentrations of NGAL, MMP-7, MMP-9, and TIMP-1, as well as their correlations with human heat shock proteins (Hsp90α, anti-Hsp60), endothelial dysfunction (sE-selectin), and inflammation (hsCRP) in pediatric patients chronically dialyzed. Twenty-two children on automated peritoneal dialysis (APD), 17 patients on hemodialysis (HD) and 24 controls were examined. The serum concentrations of NGAL, MMP-7, MMP-9, TIMP-1, Hsp90α, anti-Hsp60, and sE-selectin were assessed by enzyme-linked immunosorbent assay (ELISA). The median values of NGAL, MMP-7, MMP-9, TIMP-1, and MMP-9/NGAL ratio were significantly elevated in all dialyzed children vs. controls and were higher in HD than in APD. The values of MMP-9/TIMP-1 and MMP-7/TIMP-1 ratios in the HD subjects were lower than those in the APD children. Hsp90α and anti-Hsp60 predicted the values of NGAL, MMPs, and TIMP-1. Additionally, sE-selectin was a predictor of NGAL levels, whereas NGAL predicted the MMP and TIMP-1 concentrations. The increased concentrations of examined parameters indicate the dysfunction of MMP/TIMP/NGAL system in the dialyzed children, more pronounced on hemodialysis. The discrepancies between dialysis modalities and correlations with heat shock proteins (HSPs) suggest that NGAL may be considered a novel stress protein, whereas MMP-7, MMP-9, and TIMP-1 may be regarded as indicators of stress response in the pediatric

  5. Differential effects of metal contamination on the transcript expression of immune- and stress-response genes in the Sydney Rock oyster, Saccostrea glomerata

    International Nuclear Information System (INIS)

    Taylor, Daisy A.; Thompson, Emma L.; Nair, Sham V.; Raftos, David A.

    2013-01-01

    Environmental contamination by metals is a serious threat to the biological sustainability of coastal ecosystems. Our current understanding of the potential biological effects of metals in these ecosystems is limited. This study tested the transcriptional expression of immune- and stress-response genes in Sydney Rock oysters (Saccostrea glomerata). Oysters were exposed to four metals (cadmium, copper, lead and zinc) commonly associated with anthropogenic pollution in coastal waterways. Seven target genes (superoxide dismutase, ferritin, ficolin, defensin, HSP70, HSP90 and metallothionein) were selected. Quantitative (real-time) PCR analyses of the transcript expression of these genes showed that each of the different metals elicited unique transcriptional profiles. Significant changes in transcription were found for 18 of the 28 combinations tested (4 metals × 7 genes). Of these, 16 reflected down-regulation of gene transcription. HSP90 was the only gene significantly up-regulated by metal contamination (cadmium and zinc only), while defensin expression was significantly down-regulated by exposure to all four metals. This inhibition could have a significant negative effect on the oyster immune system, promoting susceptibility to opportunistic infections and disease. -- Highlights: ► Oysters were exposed to Cd, Cu, Pb or Zn, all commonly associated with coastal pollution. ► qPCR identified significant down-regulation in stress- and immune-response genes in oysters exposed to these metals. ► qPCR showed that each of the different metals elicited unique transcriptional profiles. ► The genes identified have the potential to lead to increased disease susceptibility in oysters. -- qPCR identified significant down-regulation in stress- and immune-response genes in oysters exposed to metals, which could lead to increased disease susceptibility

  6. Stress.

    Science.gov (United States)

    Chambers, David W

    2008-01-01

    We all experience stress as a regular, and sometimes damaging and sometimes useful, part of our daily lives. In our normal ups and downs, we have our share of exhaustion, despondency, and outrage--matched with their corresponding positive moods. But burnout and workaholism are different. They are chronic, dysfunctional, self-reinforcing, life-shortening habits. Dentists, nurses, teachers, ministers, social workers, and entertainers are especially susceptible to burnout; not because they are hard-working professionals (they tend to be), but because they are caring perfectionists who share control for the success of what they do with others and perform under the scrutiny of their colleagues (they tend to). Workaholics are also trapped in self-sealing cycles, but the elements are ever-receding visions of control and using constant activity as a barrier against facing reality. This essay explores the symptoms, mechanisms, causes, and successful coping strategies for burnout and workaholism. It also takes a look at the general stress response on the physiological level and at some of the damage American society inflicts on itself.

  7. Characterization of novel heat-responsive transcription factor (TaHSFA6e) gene involved in regulation of heat shock proteins (HSPs) - A key member of heat stress-tolerance network of wheat.

    Science.gov (United States)

    Kumar, Ranjeet R; Goswami, Suneha; Singh, Khushboo; Dubey, Kavita; Rai, Gyanendra K; Singh, Bhupinder; Singh, Shivdhar; Grover, Monendra; Mishra, Dwijesh; Kumar, Sanjeev; Bakshi, Suman; Rai, Anil; Pathak, Himanshu; Chinnusamy, Viswanathan; Praveen, Shelly

    2018-08-10

    Heat stress has an adverse effect on the quality and quantity of agriculturally important crops, especially wheat. The tolerance mechanism has not been explored much in wheat and very few genes/ TFs responsive to heat stress is available on public domain. Here, we identified, cloned and characterized a putative TaHSFA6e TF gene of 1.3 kb from wheat cv. HD2985. We observed an ORF of 368 aa with Hsf DNA binding signature domain in the amino acid sequence. Single copy number of TaHSFA6e was observed integrated in the genome of wheat. Expression analysis of TaHSFA6e under differential HS showed maximum transcripts in wheat cv. Halna (thermotolerant) in response to 38 °C for 2 h during pollination and grain-filling stages, as compared to PBW343, HD2329 and HD2985. Putative target genes of TaHSFA6e (HSP17, HSP70 and HSP90) showed upregulation in response to differential HS (30 & 38 °C, 2 h) during pollination and grain-filling stages. Small HSP17 was observed most triggered in Halna under HS. We observed increase in the catalase, guaiacol peroxidase, total antioxidant capacity (TAC), and decrease in the lipid peroxidation in thermotolerant cvs. (Halna, HD2985), as compared to thermosusceptible (PBW343, HD2329) under differential HS. Multiple stresses (heat - 38 °C, 2 h, and drought - 100 mL of 20% polyethylene Glycol 6000) during seedling stage of wheat showed positive correlation between the expression of TaHSFA6e, putative targets (HSP70, HSP90, HSP17) and TAC. Halna (thermotolerant) performed better, as compared to other contrasting cvs. TaHSFA6e TF can be used as promising candidate gene for manipulating the heat stress-tolerance network. Copyright © 2018 Elsevier B.V. All rights reserved.

  8. Response of heat shock protein genes of the oriental fruit moth under diapause and thermal stress reveals multiple patterns dependent on the nature of stress exposure.

    Science.gov (United States)

    Zhang, Bo; Peng, Yu; Zheng, Jincheng; Liang, Lina; Hoffmann, Ary A; Ma, Chun-Sen

    2016-07-01

    Heat shock protein gene (Hsp) families are thought to be important in thermal adaptation, but their expression patterns under various thermal stresses have still been poorly characterized outside of model systems. We have therefore characterized Hsp genes and their stress responses in the oriental fruit moth (OFM), Grapholita molesta, a widespread global orchard pest, and compared patterns of expression in this species to that of other insects. Genes from four Hsp families showed variable expression levels among tissues and developmental stages. Members of the Hsp40, 70, and 90 families were highly expressed under short exposures to heat and cold. Expression of Hsp40, 70, and Hsc70 family members increased in OFM undergoing diapause, while Hsp90 was downregulated. We found that there was strong sequence conservation of members of large Hsp families (Hsp40, Hsp60, Hsp70, Hsc70) across taxa, but this was not always matched by conservation of expression patterns. When the large Hsps as well as small Hsps from OFM were compared under acute and ramping heat stress, two groups of sHsps expression patterns were apparent, depending on whether expression increased or decreased immediately after stress exposure. These results highlight potential differences in conservation of function as opposed to sequence in this gene family and also point to Hsp genes potentially useful as bioindicators of diapause and thermal stress in OFM.

  9. Tomato yellow leaf curl virus infection mitigates the heat stress response of plants grown at high temperatures

    Science.gov (United States)

    Ghandi, Anfoka; Adi, Moshe; Lilia, Fridman; Linoy, Amrani; Or, Rotem; Mikhail, Kolot; Mouhammad, Zeidan; Henryk, Czosnek; Rena, Gorovits

    2016-01-01

    Cultured tomatoes are often exposed to a combination of extreme heat and infection with Tomato yellow leaf curl virus (TYLCV). This stress combination leads to intense disease symptoms and yield losses. The response of TYLCV-susceptible and resistant tomatoes to heat stress together with viral infection was compared. The plant heat-stress response was undermined in TYLCV infected plants. The decline correlated with the down-regulation of heat shock transcription factors (HSFs) HSFA2 and HSFB1, and consequently, of HSF-regulated genes Hsp17, Apx1, Apx2 and Hsp90. We proposed that the weakened heat stress response was due to the decreased capacity of HSFA2 to translocate into the nuclei of infected cells. All the six TYLCV proteins were able to interact with tomato HSFA2 in vitro, moreover, coat protein developed complexes with HSFA2 in nuclei. Capturing of HSFA2 by viral proteins could suppress the transcriptional activation of heat stress response genes. Application of both heat and TYLCV stresses was accompanied by the development of intracellular large protein aggregates containing TYLCV proteins and DNA. The maintenance of cellular chaperones in the aggregated state, even after recovery from heat stress, prevents the circulation of free soluble chaperones, causing an additional decrease in stress response efficiency. PMID:26792235

  10. The Role of Mast Cells in Tuberculosis: Orchestrating Innate Immune Crosstalk?

    Directory of Open Access Journals (Sweden)

    Karen M. Garcia-Rodriguez

    2017-10-01

    Full Text Available Tuberculosis causes more annual deaths globally than any other infectious disease. However, progress in developing novel vaccines, diagnostics, and therapies has been hampered by an incomplete understanding of the immune response to Mycobacterium tuberculosis (Mtb. While the role of many immune cells has been extensively explored, mast cells (MCs have been relatively ignored. MCs are tissue resident cells involved in defense against bacterial infections playing an important role mediating immune cell crosstalk. This review discusses specific interactions between MCs and Mtb, their contribution to both immunity and disease pathogenesis, and explores their role in orchestrating other immune cells against infections.

  11. Automation Hooks Architecture for Flexible Test Orchestration - Concept Development and Validation

    Science.gov (United States)

    Lansdowne, C. A.; Maclean, John R.; Winton, Chris; McCartney, Pat

    2011-01-01

    The Automation Hooks Architecture Trade Study for Flexible Test Orchestration sought a standardized data-driven alternative to conventional automated test programming interfaces. The study recommended composing the interface using multicast DNS (mDNS/SD) service discovery, Representational State Transfer (Restful) Web Services, and Automatic Test Markup Language (ATML). We describe additional efforts to rapidly mature the Automation Hooks Architecture candidate interface definition by validating it in a broad spectrum of applications. These activities have allowed us to further refine our concepts and provide observations directed toward objectives of economy, scalability, versatility, performance, severability, maintainability, scriptability and others.

  12. Theta oscillations orchestrate medial temporal lobe and neocortex in remembering autobiographical memories.

    Science.gov (United States)

    Fuentemilla, L; Barnes, G R; Düzel, E; Levine, B

    2014-01-15

    Remembering autobiographical events can be associated with detailed visual imagery. The medial temporal lobe (MTL), precuneus and prefrontal cortex are held to jointly enable such vivid retrieval, but how these regions are orchestrated remains unclear. An influential prediction from animal physiology is that neural oscillations in theta frequency may be important. In this experiment, participants prospectively collected audio recordings describing personal autobiographical episodes or semantic knowledge over 2 to 7 months. These were replayed as memory retrieval cues while recording brain activity with magnetoencephalography (MEG). We identified a peak of theta power within a left MTL region of interest during both autobiographical and General Semantic retrieval. This MTL region was selectively phase-synchronized with theta oscillations in precuneus and medial prefrontal cortex, and this synchrony was higher during autobiographical as compared to General Semantic knowledge retrieval. Higher synchrony also predicted more detailed visual imagery during retrieval. Thus, theta phase-synchrony orchestrates in humans the MTL with a distributed neocortical memory network when vividly remembering autobiographical experiences. © 2013.

  13. Monoaminergic orchestration of motor programs in a complex C. elegans behavior.

    Directory of Open Access Journals (Sweden)

    Jamie L Donnelly

    Full Text Available Monoamines provide chemical codes of behavioral states. However, the neural mechanisms of monoaminergic orchestration of behavior are poorly understood. Touch elicits an escape response in Caenorhabditis elegans where the animal moves backward and turns to change its direction of locomotion. We show that the tyramine receptor SER-2 acts through a Gαo pathway to inhibit neurotransmitter release from GABAergic motor neurons that synapse onto ventral body wall muscles. Extrasynaptic activation of SER-2 facilitates ventral body wall muscle contraction, contributing to the tight ventral turn that allows the animal to navigate away from a threatening stimulus. Tyramine temporally coordinates the different phases of the escape response through the synaptic activation of the fast-acting ionotropic receptor, LGC-55, and extrasynaptic activation of the slow-acting metabotropic receptor, SER-2. Our studies show, at the level of single cells, how a sensory input recruits the action of a monoamine to change neural circuit properties and orchestrate a compound motor sequence.

  14. Transcriptional response of stress genes to metal exposure in zebra mussel larvae and adults

    International Nuclear Information System (INIS)

    Navarro, Anna; Faria, Melissa; Barata, Carlos; Pina, Benjamin

    2011-01-01

    Development of stress markers for the invader freshwater zebra mussel (Dreissena polymorpha) is of great interest for both conservation and biomonitoring purposes. Gene expression profiles of several putative or already established gene expression stress markers (Metallothionein, Superoxide dismutase, Catalase, Glutathione S transferase, Glutathione peroxidase, Cytochrome c oxidase, the multixenobiotic resistance P-gp1, and heat shock proteins HSP70 and HSP90) were analyzed by quantitative Real-Time PCR in adults and pediveliger larvae after exposure to metals (Hg, Cu, Cd). A defined pattern of coordinated responses to metal exposure and, presumably, to oxidative stress was observed in gills and digestive gland from adults. A similar, albeit partial response was observed in larvae, indicating an early development of stress-related gene responses in zebra mussel. The tools developed in this study may be useful both for future control strategies and for the use of zebra mussel as sentinel species in water courses with stable populations. - Coordinated expression of stress genes in zebra mussel.

  15. Transcriptional response of stress genes to metal exposure in zebra mussel larvae and adults

    Energy Technology Data Exchange (ETDEWEB)

    Navarro, Anna; Faria, Melissa; Barata, Carlos [Institute of Environmental Assessment and Water Research (IDAEA-CSIC), Jordi Girona 18, 08034 Barcelona (Spain); Pina, Benjamin, E-mail: bpcbmc@cid.csic.e [Institute of Environmental Assessment and Water Research (IDAEA-CSIC), Jordi Girona 18, 08034 Barcelona (Spain)

    2011-01-15

    Development of stress markers for the invader freshwater zebra mussel (Dreissena polymorpha) is of great interest for both conservation and biomonitoring purposes. Gene expression profiles of several putative or already established gene expression stress markers (Metallothionein, Superoxide dismutase, Catalase, Glutathione S transferase, Glutathione peroxidase, Cytochrome c oxidase, the multixenobiotic resistance P-gp1, and heat shock proteins HSP70 and HSP90) were analyzed by quantitative Real-Time PCR in adults and pediveliger larvae after exposure to metals (Hg, Cu, Cd). A defined pattern of coordinated responses to metal exposure and, presumably, to oxidative stress was observed in gills and digestive gland from adults. A similar, albeit partial response was observed in larvae, indicating an early development of stress-related gene responses in zebra mussel. The tools developed in this study may be useful both for future control strategies and for the use of zebra mussel as sentinel species in water courses with stable populations. - Coordinated expression of stress genes in zebra mussel.

  16. 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.

  17. Genome-wide analysis of heat shock proteins in C4 model, foxtail millet identifies potential candidates for crop improvement under abiotic stress.

    Science.gov (United States)

    Singh, Roshan Kumar; Jaishankar, Jananee; Muthamilarasan, Mehanathan; Shweta, Shweta; Dangi, Anand; Prasad, Manoj

    2016-09-02

    Heat shock proteins (HSPs) perform significant roles in conferring abiotic stress tolerance to crop plants. In view of this, HSPs and their encoding genes were extensively characterized in several plant species; however, understanding their structure, organization, evolution and expression profiling in a naturally stress tolerant crop is necessary to delineate their precise roles in stress-responsive molecular machinery. In this context, the present study has been performed in C4 panicoid model, foxtail millet, which resulted in identification of 20, 9, 27, 20 and 37 genes belonging to SiHSP100, SiHSP90, SiHSP70, SiHSP60 and SisHSP families, respectively. Comprehensive in silico characterization of these genes followed by their expression profiling in response to dehydration, heat, salinity and cold stresses in foxtail millet cultivars contrastingly differing in stress tolerance revealed significant upregulation of several genes in tolerant cultivar. SisHSP-27 showed substantial higher expression in response to heat stress in tolerant cultivar, and its over-expression in yeast system conferred tolerance to several abiotic stresses. Methylation analysis of SiHSP genes suggested that, in susceptible cultivar, higher levels of methylation might be the reason for reduced expression of these genes during stress. Altogether, the study provides novel clues on the role of HSPs in conferring stress tolerance.

  18. Acute exposure to offshore produced water has an effect on stress- and secondary stress responses in three-spined stickleback Gasterosteus aculeatus.

    Science.gov (United States)

    Knag, Anne Christine; Taugbøl, Annette

    2013-09-01

    Pollution is one of today's greatest problems, and the release of contaminants into the environment can cause adverse changes in vitally important biological pathways. In this study, we exposed three-spined stickleback Gasterosteus aculeatus to produced water (PW), i.e. wastewater from offshore petroleum production. PW contains substances such as alkylphenols (APs) and aromatic hydrocarbons (PAHs) known to induce toxicant stress and endocrine disruption in a variety of organisms. Following exposure to PW, a standardized confinement treatment was applied as a second stressor (PW-stress), testing how fish already under stress from the pollutant would respond to an additional stressor. The endpoint for analysis was a combination of blood levels of cortisol and glucose, in addition to transcribed levels of a set of genes related to toxicant stress, endocrine disruption and general stress. The findings of this study indicate that low doses of PW do not induce vitellogenin in immature female stickleback, but do cause an upregulation of cytochrome (CYP1A) and UDP-glucuronsyltransferase (UDP-GT), two biomarkers related to toxicant stress. However, when the second stressor was applied, both genes were downregulated, indicating that the confinement exposure had a suppressive effect on the expression of toxicant biomarkers (CYP1A and UDP-GT). Further, two of the stress related genes, heat shock protein 90 (HSP90) and stress-induced phosphoprotein (STIP), were upregulated in both PW- and PW-stress-treatment, but not in the water control confinement treatment, indicating that PW posed as a larger stress-factor than confinement for these genes. The confinement stressor caused an increased level of glucose in both control and PW-treated fish, indicating hyperglycemia, a commonly reported stress response in fish. © 2013.

  19. Management of natural crises with choreography and orchestration of federated warning-systems

    Science.gov (United States)

    Haener, Rainer; Waechter, Joachim; Hammitzsch, Martin

    2013-04-01

    The project Collaborative, Complex and Critical Decision-Support in Evolving Crises (TRIDEC), co-funded by the European Commission in its Seventh Framework Programme focuses on real-time intelligent information management in earth management. The addressed challenges include the design and implementation of a robust and scalable service infrastructure supporting the integration of existing resources, components and systems. Key challenge for TRIDEC is establishing a network of independent systems, cooperatively interacting as a collective in a system-of-systems (SoS). For this purpose TRIDEC adopts enhancements of service-oriented architecture (SOA) principles in terms of an event-driven architecture (EDA) design (SOA 2.0). In this way TRIDEC establishes large-scale concurrent and intelligent information management of a manifold of crisis types by focusing on the integration of autonomous, task-oriented and geographically distributed systems. To this end TRIDEC adapts both ways SOA 2.0 offers: orchestration and choreography. In orchestration, a central knowledge-based processing framework takes control over the involved services and coordinates their execution. Choreography on the other hand avoids central coordination. Rather, each system involved in the SoS follows a global scenario without a single point of control but specifically defined (enacted, agreed upon) trigger conditions. More than orchestration choreography allows collaborative business processes of various heterogeneous sub-systems (e.g. cooperative decision making) by concurrent Complex Event Processing (CEP) and asynchronous communication. These types of interaction adapt the concept of decoupled relationships between information producers (e.g. sensors and sensor systems) and information consumers (e.g. warning systems and warning dissemination systems). Asynchronous communication is useful if a participant wants to trigger specific actions by delegating the responsibility (separation of concerns

  20. Search and Orchestration of Data and Processes in a Federated Environment

    Science.gov (United States)

    Siao Him Fa, J.; Reed, T. W.; Tan, C.; West, G.; McMeekin, D. A.; Moncrieff, S.; Cox, S.

    2015-06-01

    This paper describes on-going research on streamlining the access and use of spatial data and processes in Australia. Spatial data in Australia is available on-line at many levels of government from local authorities, state and territories (jurisdictions), and nationally from the Commonwealth and other sources. Much of this data is available via Open Geospatial Consortium and World Wide Web Consortium standard web services. This abstract discusses three related research topics that have been identified by a wide range of stakeholders through a comprehensive consultation process. These are search and discovery, federation and orchestration of data and processes. The commonality across the three research topics is that they all require Semantic Web and Artificial Intelligence methods and embrace the various standards, and if needed, propose modifications to such standards.

  1. Multimodal Teaching Analytics: Automated Extraction of Orchestration Graphs from Wearable Sensor Data.

    Science.gov (United States)

    Prieto, Luis P; Sharma, Kshitij; Kidzinski, Łukasz; Rodríguez-Triana, María Jesús; Dillenbourg, Pierre

    2018-04-01

    The pedagogical modelling of everyday classroom practice is an interesting kind of evidence, both for educational research and teachers' own professional development. This paper explores the usage of wearable sensors and machine learning techniques to automatically extract orchestration graphs (teaching activities and their social plane over time), on a dataset of 12 classroom sessions enacted by two different teachers in different classroom settings. The dataset included mobile eye-tracking as well as audiovisual and accelerometry data from sensors worn by the teacher. We evaluated both time-independent and time-aware models, achieving median F1 scores of about 0.7-0.8 on leave-one-session-out k-fold cross-validation. Although these results show the feasibility of this approach, they also highlight the need for larger datasets, recorded in a wider variety of classroom settings, to provide automated tagging of classroom practice that can be used in everyday practice across multiple teachers.

  2. Differentially activated macrophages orchestrate myogenic precursor cell fate during human skeletal muscle regeneration

    DEFF Research Database (Denmark)

    Saclier, Marielle; Yacoub-Youssef, Houda; Mackey, Abigail

    2013-01-01

    , we explored both in vitro and in vivo, in human, the interactions of differentially activated MPs with myogenic precursor cells (MPCs) during adult myogenesis and skeletal muscle regeneration. We showed in vitro that through the differential secretion of cytokines and growth factors, proinflammatory...... anti-inflammatory markers. These data demonstrate for the first time in human that MPs sequentially orchestrate adult myogenesis during regeneration of damaged skeletal muscle. These results support the emerging concept that inflammation, through MP activation, controls stem cell fate and coordinates......Macrophages (MPs) exert either beneficial or deleterious effects on tissue repair, depending on their activation/polarization state. They are crucial for adult skeletal muscle repair, notably by acting on myogenic precursor cells. However, these interactions have not been fully characterized. Here...

  3. A study of an adaptive replication framework for orchestrated composite web services.

    Science.gov (United States)

    Mohamed, Marwa F; Elyamany, Hany F; Nassar, Hamed M

    2013-01-01

    Replication is considered one of the most important techniques to improve the Quality of Services (QoS) of published Web Services. It has achieved impressive success in managing resource sharing and usage in order to moderate the energy consumed in IT environments. For a robust and successful replication process, attention should be paid to suitable time as well as the constraints and capabilities in which the process runs. The replication process is time-consuming since outsourcing some new replicas into other hosts is lengthy. Furthermore, nowadays, most of the business processes that might be implemented over the Web are composed of multiple Web services working together in two main styles: Orchestration and Choreography. Accomplishing a replication over such business processes is another challenge due to the complexity and flexibility involved. In this paper, we present an adaptive replication framework for regular and orchestrated composite Web services. The suggested framework includes a number of components for detecting unexpected and unhappy events that might occur when consuming the original published web services including failure or overloading. It also includes a specific replication controller to manage the replication process and select the best host that would encapsulate a new replica. In addition, it includes a component for predicting the incoming load in order to decrease the time needed for outsourcing new replicas, enhancing the performance greatly. A simulation environment has been created to measure the performance of the suggested framework. The results indicate that adaptive replication with prediction scenario is the best option for enhancing the performance of the replication process in an online business environment.

  4. FAST: A fully asynchronous and status-tracking pattern for geoprocessing services orchestration

    Science.gov (United States)

    Wu, Huayi; You, Lan; Gui, Zhipeng; Gao, Shuang; Li, Zhenqiang; Yu, Jingmin

    2014-09-01

    Geoprocessing service orchestration (GSO) provides a unified and flexible way to implement cross-application, long-lived, and multi-step geoprocessing service workflows by coordinating geoprocessing services collaboratively. Usually, geoprocessing services and geoprocessing service workflows are data and/or computing intensive. The intensity feature may make the execution process of a workflow time-consuming. Since it initials an execution request without blocking other interactions on the client side, an asynchronous mechanism is especially appropriate for GSO workflows. Many critical problems remain to be solved in existing asynchronous patterns for GSO including difficulties in improving performance, status tracking, and clarifying the workflow structure. These problems are a challenge when orchestrating performance efficiency, making statuses instantly available, and constructing clearly structured GSO workflows. A Fully Asynchronous and Status-Tracking (FAST) pattern that adopts asynchronous interactions throughout the whole communication tier of a workflow is proposed for GSO. The proposed FAST pattern includes a mechanism that actively pushes the latest status to clients instantly and economically. An independent proxy was designed to isolate the status tracking logic from the geoprocessing business logic, which assists the formation of a clear GSO workflow structure. A workflow was implemented in the FAST pattern to simulate the flooding process in the Poyang Lake region. Experimental results show that the proposed FAST pattern can efficiently tackle data/computing intensive geoprocessing tasks. The performance of all collaborative partners was improved due to the asynchronous mechanism throughout communication tier. A status-tracking mechanism helps users retrieve the latest running status of a GSO workflow in an efficient and instant way. The clear structure of the GSO workflow lowers the barriers for geospatial domain experts and model designers to

  5. Alteration of Transcripts of Stress-Protective Genes and Transcriptional Factors by γ-Aminobutyric Acid (GABA Associated with Improved Heat and Drought Tolerance in Creeping Bentgrass (Agrostis stolonifera

    Directory of Open Access Journals (Sweden)

    Zhou Li

    2018-05-01

    Full Text Available Gamma-aminobutyric acid (GABA may play a positive role in regulating plant tolerance to drought or heat stress. The objectives of this study were to investigate the physiological effects of GABA on tolerance of creeping bentgrass (Agrostis stolonifera to heat and drought stress and to determine whether enhanced heat and drought tolerance due to GABA treatment was associated with the up-regulation of selected genes and transcriptional factors involved in stress protection. Creeping bentgrass (cultivar “Penncross” plants were treated with 0.5 mM GABA or water (untreated control as a foliar spray and were subsequently exposed to heat stress (35/30 °C, day/night, drought stress by withholding irrigation, or non-stress conditions in controlled-environment growth chambers. Exogenous application of GABA significantly improved plant tolerance to heat and drought stress, as reflected by increased leaf water content, cell membrane stability, and chlorophyll content. The analysis of gene transcript level revealed that exogenous GABA up-regulated the expression of ABF3, POD, APX, HSP90, DHN3, and MT1 during heat stress and the expression of CDPK26, MAPK1, ABF3, WRKY75, MYB13, HSP70, MT1, 14-3-3, and genes (SOD, CAT, POD, APX, MDHAR, DHAR, and GR encoding antioxidant enzymes during drought stress. The up-regulation of the aforementioned stress-protective genes and transcriptional factors could contribute to improved heat and drought tolerance in creeping bentgrass.

  6. Alteration of Transcripts of Stress-Protective Genes and Transcriptional Factors by γ-Aminobutyric Acid (GABA) Associated with Improved Heat and Drought Tolerance in Creeping Bentgrass (Agrostis stolonifera).

    Science.gov (United States)

    Li, Zhou; Peng, Yan; Huang, Bingru

    2018-05-31

    Gamma-aminobutyric acid (GABA) may play a positive role in regulating plant tolerance to drought or heat stress. The objectives of this study were to investigate the physiological effects of GABA on tolerance of creeping bentgrass ( Agrostis stolonifera ) to heat and drought stress and to determine whether enhanced heat and drought tolerance due to GABA treatment was associated with the up-regulation of selected genes and transcriptional factors involved in stress protection. Creeping bentgrass (cultivar "Penncross") plants were treated with 0.5 mM GABA or water (untreated control) as a foliar spray and were subsequently exposed to heat stress (35/30 °C, day/night), drought stress by withholding irrigation, or non-stress conditions in controlled-environment growth chambers. Exogenous application of GABA significantly improved plant tolerance to heat and drought stress, as reflected by increased leaf water content, cell membrane stability, and chlorophyll content. The analysis of gene transcript level revealed that exogenous GABA up-regulated the expression of ABF3 , POD , APX , HSP90 , DHN3 , and MT1 during heat stress and the expression of CDPK26 , MAPK1 , ABF3 , WRKY75 , MYB13 , HSP70 , MT1 , 14-3-3 , and genes ( SOD , CAT , POD , APX , MDHAR , DHAR , and GR ) encoding antioxidant enzymes during drought stress. The up-regulation of the aforementioned stress-protective genes and transcriptional factors could contribute to improved heat and drought tolerance in creeping bentgrass.

  7. Siim Nestor soovitab : Bell Orchestre. DJ Deep ja TRL. Akrobatik ja MC Mr.Lif. Black Dice / Siim Nestor

    Index Scriptorium Estoniae

    Nestor, Siim, 1974-

    2008-01-01

    Muusikaüritustest: Kanada post-rock ansambel Bell Orchestre12. sept. Tallinnas klubis Juuksur, DJ Deep 12. sept. Tallinna Linnahalli ruumides Plektrumi klubiööl (vt. www.plektrumfestival.ee), Akrobatik ja Mr.Lif 12. sept. Tartus klubis Illusion, New Yorgi kollektiiv Black Dice 17. sept. Tallinnas Von Krahlis

  8. Plant Nucleolar Stress Response, a New Face in the NAC-Dependent Cellular Stress Responses

    OpenAIRE

    Iwai Ohbayashi; Munetaka Sugiyama

    2018-01-01

    The nucleolus is the most prominent nuclear domain, where the core processes of ribosome biogenesis occur vigorously. All these processes are finely orchestrated by many nucleolar factors to build precisely ribosome particles. In animal cells, perturbations of ribosome biogenesis, mostly accompanied by structural disorders of the nucleolus, cause a kind of cellular stress to induce cell cycle arrest, senescence, or apoptosis, which is called nucleolar stress response. The best-characterized p...

  9. Investigating the underlying mechanism of Saccharomyces cerevisiae in response to ethanol stress employing RNA-seq analysis.

    Science.gov (United States)

    Li, Ruoyun; Xiong, Guotong; Yuan, Shukun; Wu, Zufang; Miao, Yingjie; Weng, Peifang

    2017-11-03

    Saccharomyces cerevisiae has been widely used for wine fermentation and bio-fuels production. A S. cerevisiae strain Sc131 isolated from tropical fruit shows good fermentation properties and ethanol tolerance, exhibiting significant potential in Chinese bayberry wine fermentation. In this study, RNA-sequence and RT-qPCR was used to investigate the transcriptome profile of Sc131 in response to ethanol stress. Scanning Electron Microscopy were carried out to observe surface morphology of yeast cells. Totally, 937 genes were identified differential expressed, including 587 up-regulated and 350 down-regulated genes, after 4-h ethanol stress (10% v/v). Transcriptomic analysis revealed that, most genes involved in regulating filamentous growth or pseudohyphal growth were significantly up-regulated in response to ethanol stress. The complex protein quality control machineries, Hsp90/Hsp70 and Hsp104/Hsp70/Hsp40 based chaperone system combining with ubiquitin-proteasome proteolytic pathway were both activated to recognize and degrade misfolding proteins. Genes related to biosynthesis and metabolism of two well-known stress-responsive substances trehalose and ergosterol were generally up-regulated, while genes associated with amino acids biosynthesis and metabolism processes were differentially expressed. Moreover, thiamine was also important in response to ethanol stress. This research may promote the potential applications of Sc131 in the fermentation of Chinese bayberry wine.

  10. Prospects of engineering thermotolerance in crops through modulation of heat stress transcription factor and heat shock protein networks.

    Science.gov (United States)

    Fragkostefanakis, Sotirios; Röth, Sascha; Schleiff, Enrico; Scharf, Klaus-Dieter

    2015-09-01

    Cell survival under high temperature conditions involves the activation of heat stress response (HSR), which in principle is highly conserved among different organisms, but shows remarkable complexity and unique features in plant systems. The transcriptional reprogramming at higher temperatures is controlled by the activity of the heat stress transcription factors (Hsfs). Hsfs allow the transcriptional activation of HSR genes, among which heat shock proteins (Hsps) are best characterized. Hsps belong to multigene families encoding for molecular chaperones involved in various processes including maintenance of protein homeostasis as a requisite for optimal development and survival under stress conditions. Hsfs form complex networks to activate downstream responses, but are concomitantly subjected to cell-type-dependent feedback regulation through factor-specific physical and functional interactions with chaperones belonging to Hsp90, Hsp70 and small Hsp families. There is increasing evidence that the originally assumed specialized function of Hsf/chaperone networks in the HSR turns out to be a complex central stress response system that is involved in the regulation of a broad variety of other stress responses and may also have substantial impact on various developmental processes. Understanding in detail the function of such regulatory networks is prerequisite for sustained improvement of thermotolerance in important agricultural crops. © 2014 John Wiley & Sons Ltd.

  11. The FKBP51-Glucocorticoid Receptor Balance in Stress-Related Mental Disorders.

    Science.gov (United States)

    Fries, Gabriel R; Gassen, Nils C; Schmidt, Ulrike; Rein, Theo

    2015-01-01

    The immunophilin FK506 binding protein 51 (FKBP51) has emerged as one of the most intensely investigated proteins in stress-related mental disorders. It was originally characterized as Hsp90 cochaperone and part of the receptor-chaperone heterocomplex that governs the activity of steroid receptors. It turned out that the presence of FKBP51 in this heterocomplex leads to diminished activity of the corticosteroid receptors. In particular, based on its inhibitory action on the glucocorticoid receptor (GR), FKBP51 was included in a candidate gene approach to discover gene polymorphisms that might be relevant for the development and treatment of major depression. The discovery that polymorphisms in the gene coding for FKBP51 were linked to the treatment response of depressed patients intensified the research on the role of FKBP51 in stress-related diseases worldwide. It has become evident that FKBP51 is not only a regulator of GR action, but also a GR target. The function of this ultrashort intracellular feedback loop is critically important for cellular and physiological stress regulation as it does not only calibrate the function of GR, but also the levels of FKBP51. Given the pleiotropic functions of FKBP51, its levels might be equally important for mental disorders as GR function and hence for the development of potential FKBP51 drug targets.

  12. A Proteomic Screen Identified Stress-Induced Chaperone Proteins as Targets of Akt Phosphorylation in Mesangial Cells

    OpenAIRE

    Barati, Michelle T.; Rane, Madhavi J.; Klein, Jon B.; McLeish, Kenneth R.

    2006-01-01

    The serine-threonine kinase Akt regulates mesangial cell apoptosis, proliferation, and hypertrophy. To define Akt signaling pathways in mesangial cells, we performed a functional proteomic screen for rat mesangial cell proteins phosphorylated by Akt. A group of chaperone proteins, heat shock protein (Hsp) 70, Hsp90α, Hsp90β, Glucose-regulated protein (Grp) Grp78, Grp94, and protein disulfide isomerase (PDI) were identified as potential Akt substrates by two techniques: (a) in vitro phosphoryl...

  13. Development of Tethered Hsp90 Inhibitors Carrying Radioiodinated Probes to Specifically Discriminate and Kill Malignant Breast Tumor Cells

    Science.gov (United States)

    2017-05-01

    ligand localization . These molecules will be tested as imaging agents in mouse models of breast cancer. 15. SUBJECT TERMS Radiodination, tethered...annually leading to the diagnosis and surgical resection of breast cancer or breast carcinoma in situ in over 250,000 women respectively. Unfortunately

  14. Interactions of p60, a mediator of progesterone receptor assembly, with heat shock proteins hsp90 and hsp70

    DEFF Research Database (Denmark)

    Chen, S; Prapapanich, V; Rimerman, R A

    1996-01-01

    Previous studies on the assembly of progesterone receptor (PR) complexes in vitro have suggested that PR assembly is a dynamic, ordered process involving at least eight nonreceptor proteins. One of these proteins, p60, appears transiently during assembly and is not a component of functionally...

  15. Exposure to non-ionizing radiation provokes changes in rat thyroid morphology and expression of HSP-90

    OpenAIRE

    Misa-Agustiño, Maria J; Jorge-Mora, Teresa; Jorge-Barreiro, Francisco J; Suarez-Quintanilla, Juan; Moreno-Piquero, Eduardo; Ares-Pena, Francisco J; López-Martín, Elena

    2015-01-01

    Non-ionizing radiation at 2.45 GHz may modify the morphology and expression of genes that codify heat shock proteins (HSP) in the thyroid gland. Diathermy is the therapeutic application of non-ionizing radiation to humans for its beneficial effects in rheumatological and musculo-skeletal pain processes. We used a diathermy model on laboratory rats subjected to maximum exposure in the left front leg, in order to study the effects of radiation on the nearby thyroid tissue. Fifty-six rats were i...

  16. NMR characterization of HtpG, the E. coli Hsp90, using sparse labeling with 13C-methyl alanine.

    Science.gov (United States)

    Pederson, Kari; Chalmers, Gordon R; Gao, Qi; Elnatan, Daniel; Ramelot, Theresa A; Ma, Li-Chung; Montelione, Gaetano T; Kennedy, Michael A; Agard, David A; Prestegard, James H

    2017-07-01

    A strategy for acquiring structural information from sparsely isotopically labeled large proteins is illustrated with an application to the E. coli heat-shock protein, HtpG (high temperature protein G), a 145 kDa dimer. It uses 13 C-alanine methyl labeling in a perdeuterated background to take advantage of the sensitivity and resolution of Methyl-TROSY spectra, as well as the backbone-centered structural information from 1 H- 13 C residual dipolar couplings (RDCs) of alanine methyl groups. In all, 40 of the 47 expected crosspeaks were resolved and 36 gave RDC data. Assignments of crosspeaks were partially achieved by transferring assignments from those made on individual domains using triple resonance methods. However, these were incomplete and in many cases the transfer was ambiguous. A genetic algorithm search for consistency between predictions based on domain structures and measurements for chemical shifts and RDCs allowed 60% of the 40 resolved crosspeaks to be assigned with confidence. Chemical shift changes of these crosspeaks on adding an ATP analog to the apo-protein are shown to be consistent with structural changes expected on comparing previous crystal structures for apo- and complex- structures. RDCs collected on the assigned alanine methyl peaks are used to generate a new solution model for the apo-protein structure.

  17. Dietary supplementation of curcumin augments heat stress tolerance through upregulation of nrf-2-mediated antioxidative enzymes and hsps in Puntius sophore.

    Science.gov (United States)

    Mahanty, Arabinda; Mohanty, Sasmita; Mohanty, Bimal P

    2017-08-01

    Heat stress is one of the major environmental concerns in global warming regime and rising temperature has resulted in mass mortalities of animals including fishes. Therefore, strategies for high temperature stress tolerance and ameliorating the effects of heat stress are being looked for. In an earlier study, we reported that Nrf-2 (nuclear factor E2-related factor 2) mediated upregulation of antioxidative enzymes and heat shock proteins (Hsps) provide survivability to fish under heat stress. In this study, we have evaluated the ameliorative potential of dietary curcumin, a potential Nrf-2 inducer in heat stressed cyprinid Puntius sophore. Fishes were fed with diet supplemented with 0.5, 1.0, and 1.5% curcumin at the rate 2% of body weight daily in three separate groups (n = 40 in each group) for 60 days. Fishes fed with basal diet (without curcumin) served as the control (n = 40). Critical thermal maxima (CTmax) was determined for all the groups (n = 10, in duplicates) after the feeding trial. Significant increase in the CTmax was observed in the group fed with 1.5% curcumin- supplemented fishes whereas it remained similar in groups fed with 0.5%, and 1% curcumin-supplemented diet, as compared to control. To understand the molecular mechanism of elevated thermotolerance in the 1.5% curcumin supplemented group, fishes were given a sub-lethal heat shock treatment (36 °C) for 6 h and expression analysis of nrf-2, keap-1, sod, catalase, gpx, and hsp27, hsp60, hsp70, hsp90, and hsp110 was carried out using RT-PCR. In the gill, expression of nrf-2, sod, catalase, gpx, and hsp60, hsp70, hsp90, and hsp110 was found to be elevated in the 1.5% curcumin-fed heat-shocked group compared to control and the basal diet-fed, heat-shocked fishes. Similarly, in the liver, upregulation in expression of nrf-2, sod, catalase, and hsp70 and hsp110 was observed in 1.5% curcumin supplemented and heat shocked group. Thus, this study showed that supplementation of curcumin

  18. Effects of heat stress on serum insulin, adipokines, AMP-activated protein kinase, and heat shock signal molecules in dairy cows*

    Science.gov (United States)

    Min, Li; Cheng, Jian-bo; Shi, Bao-lu; Yang, Hong-jian; Zheng, Nan; Wang, Jia-qi

    2015-01-01

    Heat stress affects feed intake, milk production, and endocrine status in dairy cows. The temperature-humidity index (THI) is employed as an index to evaluate the degree of heat stress in dairy cows. However, it is difficult to ascertain whether THI is the most appropriate measurement of heat stress in dairy cows. This experiment was conducted to investigate the effects of heat stress on serum insulin, adipokines (leptin and adiponectin), AMP-activated protein kinase (AMPK), and heat shock signal molecules (heat shock transcription factor (HSF) and heat shock proteins (HSP)) in dairy cows and to research biomarkers to be used for better understanding the meaning of THI as a bioclimatic index. To achieve these objectives, two experiments were performed. The first experiment: eighteen lactating Holstein dairy cows were used. The treatments were: heat stress (HS, THI average=81.7, n=9) and cooling (CL, THI average=53.4, n=9). Samples of HS were obtained on August 16, 2013, and samples of CL were collected on April 7, 2014 in natural conditions. The second experiment: HS treatment cows (n=9) from the first experiment were fed for 8 weeks from August 16, 2013 to October 12, 2013. Samples for moderate heat stress, mild heat stress, and no heat stress were obtained, respectively, according to the physical alterations of the THI. Results showed that heat stress significantly increased the serum adiponectin, AMPK, HSF, HSP27, HSP70, and HSP90 (Pheat-stressed dairy cows. When heat stress treatment lasted 8 weeks, a higher expression of HSF and HSP70 was observed under moderate heat stress. Serum HSF and HSP70 are sensitive and accurate in heat stress and they could be potential indicators of animal response to heat stress. We recommend serum HSF and HSP70 as meaningful biomarkers to supplement the THI and evaluate moderate heat stress in dairy cows in the future. PMID:26055916

  19. Cloud-Based Orchestration of a Model-Based Power and Data Analysis Toolchain

    Science.gov (United States)

    Post, Ethan; Cole, Bjorn; Dinkel, Kevin; Kim, Hongman; Lee, Erich; Nairouz, Bassem

    2016-01-01

    The proposed Europa Mission concept contains many engineering and scientific instruments that consume varying amounts of power and produce varying amounts of data throughout the mission. System-level power and data usage must be well understood and analyzed to verify design requirements. Numerous cross-disciplinary tools and analysis models are used to simulate the system-level spacecraft power and data behavior. This paper addresses the problem of orchestrating a consistent set of models, tools, and data in a unified analysis toolchain when ownership is distributed among numerous domain experts. An analysis and simulation environment was developed as a way to manage the complexity of the power and data analysis toolchain and to reduce the simulation turnaround time. A system model data repository is used as the trusted store of high-level inputs and results while other remote servers are used for archival of larger data sets and for analysis tool execution. Simulation data passes through numerous domain-specific analysis tools and end-to-end simulation execution is enabled through a web-based tool. The use of a cloud-based service facilitates coordination among distributed developers and enables scalable computation and storage needs, and ensures a consistent execution environment. Configuration management is emphasized to maintain traceability between current and historical simulation runs and their corresponding versions of models, tools and data.

  20. Adipocytes and Macrophages Interplay in the Orchestration of Tumor Microenvironment: New Implications in Cancer Progression

    Directory of Open Access Journals (Sweden)

    Luís Henrique Corrêa

    2017-09-01

    Full Text Available Inflammation has been known as one of the main keys to the establishment and progression of cancers. Chronic low-grade inflammation is also a strategic condition that underlies the causes and development of metabolic syndrome and obesity. Moreover, obesity has been largely related to poor prognosis of tumors by modulating tumor microenvironment with secretion of several inflammatory mediators by tumor-associated adipocytes (TAAs, which can modulate and recruit tumor-associated macrophages. Thus, the understanding of cellular and molecular mechanisms that underlay and link inflammation, obesity, and cancer is crucial to identify potential targets that interfere with this important route. Knowledge about the exact role of each component of the tumor microenvironment is not yet fully understood, but the new insights in literature highlight the essential role of adipocytes and macrophages interplay as key factor to determine the fate of cancer progression. In this review article, we focus on the functions of adipocytes and macrophages orchestrating cellular and molecular mechanisms that lead to inflammatory modulation in tumor microenvironment, which will be crucial to cancer establishment. We also emphasized the mechanisms by which the tumor promotes itself by recruiting and polarizing macrophages, discussing the role of adipocytes in this process. In addition, we discuss here the newest possible anticancer therapeutic treatments aiming to retard the development of the tumor based on what is known about cancer, adipocyte, and macrophage polarization.

  1. MyD88-dependent dendritic and epithelial cell crosstalk orchestrates immune responses to allergens.

    Science.gov (United States)

    Thomas, S Y; Whitehead, G S; Takaku, M; Ward, J M; Xu, X; Nakano, K; Lyons-Cohen, M R; Nakano, H; Gowdy, K M; Wade, P A; Cook, D N

    2018-05-01

    Sensitization to inhaled allergens is dependent on activation of conventional dendritic cells (cDCs) and on the adaptor molecule, MyD88. However, many cell types in the lung express Myd88, and it is unclear how signaling in these different cell types reprograms cDCs and leads to allergic inflammation of the airway. By combining ATAC-seq with RNA profiling, we found that MyD88 signaling in cDCs maintained open chromatin at select loci even at steady state, allowing genes to be rapidly induced during allergic sensitization. A distinct set of genes related to metabolism was indirectly controlled in cDCs through MyD88 signaling in airway epithelial cells (ECs). In mouse models of asthma, Myd88 expression in ECs was critical for eosinophilic inflammation, whereas Myd88 expression in cDCs was required for Th17 cell differentiation and consequent airway neutrophilia. Thus, both cell-intrinsic and cell-extrinsic MyD88 signaling controls gene expression in cDCs and orchestrates immune responses to inhaled allergens.

  2. ISOMP: An Instant Service-Orchestration Mobile M2M Platform

    Directory of Open Access Journals (Sweden)

    Cholhong Im

    2016-01-01

    Full Text Available Smartphones have greater computing power than ever before, providing convenient applications to improve our lives. In general, people find it difficult to locate suitable applications and implementing new applications often requires professional skills. In this paper, we propose a new service platform that facilitates the implementation of new applications by composing prebuilt components that provide the context information of mobile devices such as location and contacts. Our platform introduces an innovative concept named context collaboration, in which smartphones exchange context information with each other, which in turn is used to deduct useful inferences. The concept is realized by instant orchestration, which assembles some components and implements a composite component. The interactive communication interface helps a mobile device to communicate with other devices using open APIs, such as SOAP and HTTP (REST. The platform also works in heterogeneous environments, for example, between Android and iOS operating systems. Throughout the platform, mobile devices can act as smart M2M machines with context awareness, enabling intelligent tasks on behalf of users. Our platform will open up a new and innovative pathway for both enhanced mobile context awareness and M2M, which is expected to be a fundamental feature of the next generation of mobile devices.

  3. Industrial Robot Programming and UPnP Services Orchestration for the Automation of Factories

    Directory of Open Access Journals (Sweden)

    A. Valera

    2012-10-01

    Full Text Available The integration of equipment and other devices built into industrial robot cells with modern Ethernet interface technologies and low-cost mass produced devices (such as vision systems, laser scanners, force torque-sensors, PLCs and PDAs etc. enables integrators to offer more powerful and smarter solutions. Nevertheless, the programming of all these devices efficiently requires very specific knowledge about them, such as their hardware architectures and specific programming languages as well as details about the system's low level communication protocols. To address these issues, this paper describes and analyses the Plug-and-Play architecture. This is one of the most interesting service-oriented architectures (SOAs available, which exhibits characteristics that are well adapted to industrial robotics cells. To validate their programming features and applicability, a test bed was specially designed. This provides a new graphical service orchestration which was implemented using Workflow Foundation 4 of .NET. The obtained results allowed us to verify that the use of integration schemes based on SOAs reduces the system integration time and is better adapted to industrial robotic cell system integrators.

  4. Orchestrating cytoskeleton and intracellular vesicle traffic to build functional immunological synapses.

    Science.gov (United States)

    Soares, Helena; Lasserre, Rémi; Alcover, Andrés

    2013-11-01

    Immunological synapses are specialized cell-cell contacts formed between T lymphocytes and antigen-presenting cells. They are induced upon antigen recognition and are crucial for T-cell activation and effector functions. The generation and function of immunological synapses depend on an active T-cell polarization process, which results from a finely orchestrated crosstalk between the antigen receptor signal transduction machinery, the actin and microtubule cytoskeletons, and controlled vesicle traffic. Although we understand how some of these particular events are regulated, we still lack knowledge on how these multiple cellular elements are harmonized to ensure appropriate T-cell responses. We discuss here our view on how T-cell receptor signal transduction initially commands cytoskeletal and vesicle traffic polarization, which in turn sets the immunological synapse molecular design that regulates T-cell activation. We also discuss how the human immunodeficiency virus (HIV-1) hijacks some of these processes impairing immunological synapse generation and function. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  5. AIDing Chromatin and Transcription-Coupled Orchestration of Immunoglobulin Class-Switch Recombination

    Science.gov (United States)

    Vaidyanathan, Bharat; Yen, Wei-Feng; Pucella, Joseph N.; Chaudhuri, Jayanta

    2014-01-01

    Secondary diversification of the antibody repertoire upon antigenic challenge, in the form of immunoglobulin heavy chain (IgH) class-switch recombination (CSR) endows mature, naïve B cells in peripheral lymphoid organs with a limitless ability to mount an optimal humoral immune response, thus expediting pathogen elimination. CSR replaces the default constant (CH) region exons (Cμ) of IgH with any of the downstream CH exons (Cγ, Cε, or Cα), thereby altering effector functions of the antibody molecule. This process depends on, and is orchestrated by, activation-induced deaminase (AID), a DNA cytidine deaminase that acts on single-stranded DNA exposed during transcription of switch (S) region sequences at the IgH locus. DNA lesions thus generated are processed by components of several general DNA repair pathways to drive CSR. Given that AID can instigate DNA lesions and genomic instability, stringent checks are imposed that constrain and restrict its mutagenic potential. In this review, we will discuss how AID expression and substrate specificity and activity is rigorously enforced at the transcriptional, post-transcriptional, post-translational, and epigenetic levels, and how the DNA-damage response is choreographed with precision to permit targeted activity while limiting bystander catastrophe. PMID:24734031

  6. Memory CD8+ T Cells: Orchestrators and Key Players of Innate Immunity?

    Directory of Open Access Journals (Sweden)

    Grégoire Lauvau

    2016-09-01

    Full Text Available Over the past decades, the dichotomy between innate and adaptive immune responses has largely dominated our understanding of immunology. Upon primary encounter with microbial pathogens, differentiation of adaptive immune cells into functional effectors usually takes several days or even longer, making them contribute to host protection only late during primary infection. However, once generated, antigen-experienced T lymphocytes can persist in the organism and constitute a pool of memory cells that mediate fast and effective protection to a recall infection with the same microbial pathogen. Herein, we challenge this classical paradigm by highlighting the "innate nature" of memory CD8+ T cells. First, within the thymus or in the periphery, naïve CD8+ T cells may acquire phenotypic and functional characteristics of memory CD8+ T cells independently of challenge with foreign antigens. Second, both the "unconventional" and the "conventional" memory cells can rapidly express protective effector functions in response to sets of inflammatory cytokines and chemokines signals, independent of cognate antigen triggering. Third, memory CD8+ T cells can act by orchestrating the recruitment, activation, and licensing of innate cells, leading to broad antimicrobial states. Thus, collectively, memory CD8+ T cells may represent important actors of innate immune defenses.

  7. Nuclear organization in the spinal cord depends on motor neuron lamination orchestrated by catenin and afadin function

    OpenAIRE

    Dewitz, C.; Pimpinella, S.; Hackel, P.; Akalin, A.; Jessell, T.M.; Zampieri, N.

    2018-01-01

    Motor neurons in the spinal cord are found grouped in nuclear structures termed pools, whose position is precisely orchestrated during development. Despite the emerging role of pool organization in the assembly of spinal circuits, little is known about the morphogenetic programs underlying the patterning of motor neuron subtypes. We applied three-dimensional analysis of motor neuron position to reveal the roles and contributions of cell adhesive function by inactivating N-cadherin, catenin, a...

  8. Placental heat shock proteins: no immunohistochemical evidence for a differential stress response in preterm labour.

    Science.gov (United States)

    Divers, M J; Bulmer, J N; Miller, D; Lilford, R J

    1995-01-01

    The aetiology of idiopathic preterm labour remains obscure. The hypothesis that a stress response induced by low-grade bacterial infection in utero-placental tissues was investigated. Distribution of cognate and inducible isoforms of heat shock proteins (HSP) 70 kD, HSP 60 kD and HSP 90 kD were investigated in an immunohistochemical study of placental and decidual tissues before and after labour at varying gestations. Subjects were pregnant women undergoing singleton delivery after idiopathic preterm labour at less than 34 weeks' gestation (n = 23); spontaneous term labour at 37-42 weeks' gestation (n =24); preterm caesarean sections at less than 34 weeks' gestation for preeclampsia or intrauterine growth retardation (n=14); elective caesarean section at 37-42 weeks' gestation for cephalopelvic disproportion (n = 6). HSP expression was constant throughout the third trimester of pregnancy and did not change following the onset of labour, regardless of gestational age. A stress response in decidual tissues as determined by immunohistochemical analysis is apparently not associated with preterm labour.

  9. Effects of heat stress on serum insulin, adipokines, AMP-activated protein kinase, and heat shock signal molecules in dairy cows.

    Science.gov (United States)

    Min, Li; Cheng, Jian-bo; Shi, Bao-lu; Yang, Hong-jian; Zheng, Nan; Wang, Jia-qi

    2015-06-01

    Heat stress affects feed intake, milk production, and endocrine status in dairy cows. The temperature-humidity index (THI) is employed as an index to evaluate the degree of heat stress in dairy cows. However, it is difficult to ascertain whether THI is the most appropriate measurement of heat stress in dairy cows. This experiment was conducted to investigate the effects of heat stress on serum insulin, adipokines (leptin and adiponectin), AMP-activated protein kinase (AMPK), and heat shock signal molecules (heat shock transcription factor (HSF) and heat shock proteins (HSP)) in dairy cows and to research biomarkers to be used for better understanding the meaning of THI as a bioclimatic index. To achieve these objectives, two experiments were performed. The first experiment: eighteen lactating Holstein dairy cows were used. The treatments were: heat stress (HS, THI average=81.7, n=9) and cooling (CL, THI average=53.4, n=9). Samples of HS were obtained on August 16, 2013, and samples of CL were collected on April 7, 2014 in natural conditions. The second experiment: HS treatment cows (n=9) from the first experiment were fed for 8 weeks from August 16, 2013 to October 12, 2013. Samples for moderate heat stress, mild heat stress, and no heat stress were obtained, respectively, according to the physical alterations of the THI. Results showed that heat stress significantly increased the serum adiponectin, AMPK, HSF, HSP27, HSP70, and HSP90 (Pdairy cows. When heat stress treatment lasted 8 weeks, a higher expression of HSF and HSP70 was observed under moderate heat stress. Serum HSF and HSP70 are sensitive and accurate in heat stress and they could be potential indicators of animal response to heat stress. We recommend serum HSF and HSP70 as meaningful biomarkers to supplement the THI and evaluate moderate heat stress in dairy cows in the future.

  10. Proteolytic cleavage orchestrates cofactor insertion and protein assembly in [NiFe]-hydrogenase biosynthesis.

    Science.gov (United States)

    Senger, Moritz; Stripp, Sven T; Soboh, Basem

    2017-07-14

    Metalloenzymes catalyze complex and essential processes, such as photosynthesis, respiration, and nitrogen fixation. For example, bacteria and archaea use [NiFe]-hydrogenases to catalyze the uptake and release of molecular hydrogen (H 2 ). [NiFe]-hydrogenases are redox enzymes composed of a large subunit that harbors a NiFe(CN) 2 CO metallo-center and a small subunit with three iron-sulfur clusters. The large subunit is synthesized with a C-terminal extension, cleaved off by a specific endopeptidase during maturation. The exact role of the C-terminal extension has remained elusive; however, cleavage takes place exclusively after assembly of the [NiFe]-cofactor and before large and small subunits form the catalytically active heterodimer. To unravel the functional role of the C-terminal extension, we used an enzymatic in vitro maturation assay that allows synthesizing functional [NiFe]-hydrogenase-2 of Escherichia coli from purified components. The maturation process included formation and insertion of the NiFe(CN) 2 CO cofactor into the large subunit, endoproteolytic cleavage of the C-terminal extension, and dimerization with the small subunit. Biochemical and spectroscopic analysis indicated that the C-terminal extension of the large subunit is essential for recognition by the maturation machinery. Only upon completion of cofactor insertion was removal of the C-terminal extension observed. Our results indicate that endoproteolytic cleavage is a central checkpoint in the maturation process. Here, cleavage temporally orchestrates cofactor insertion and protein assembly and ensures that only cofactor-containing protein can continue along the assembly line toward functional [NiFe]-hydrogenase. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  11. 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.

  12. Acute heat stress induces differential gene expressions in the testes of a broiler-type strain of Taiwan country chickens.

    Science.gov (United States)

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

    2015-01-01

    The expression of testicular genes following acute heat stress has been reported in layer-type roosters, but few similar studies have been conducted on broilers. This study investigated the effect of acute heat stress on the gene expression in the testes of a broiler-type strain of Taiwan country chickens. Roosters were subjected to acute heat stress (38°C) for 4 h, and then exposed to 25°C, with testes collected 0, 2, and 6 h after the cessation of heat stress, using non-heat-stressed roosters as controls (n = 3 roosters per group). The body temperature and respiratory rate increased significantly (pstress. The numbers of apoptotic cells increased 2 h after the acute heat stress (79 ± 7 vs. 322 ± 192, control vs. heat stress; pstressed chickens from those of the controls, including genes involved in the response to stimulus, protein metabolism, signal transduction, cell adhesion, transcription, and apoptosis. The mRNA expressions of upregulated genes, including HSP25, HSP90AA1, HSPA2, and LPAR2, and of downregulated genes, including CDH5, CTNNA3, EHF, CIRBP, SLA, and NTF3, were confirmed through quantitative real-time polymerase chain reaction (qRT-PCR). Moreover, numerous transcripts in the testes exhibited distinct expressions between the heat-stressed broiler-type and layer-type chickens. We concluded that the transcriptional responses of testes to acute heat stress may differ between the broiler-type and layer-type roosters. Whether the differential expression patterns associate with the heat-tolerance in the strains require a further exploration.

  13. Temporal patterns of cardiac performance and genes encoding heat shock proteins and metabolic sensors of an intertidal limpet Cellana toreuma during sublethal heat stress.

    Science.gov (United States)

    Zhang, Shu; Han, Guo-dong; Dong, Yun-wei

    2014-04-01

    Intertidal invertebrates develop effective physiological adaptations to cope with the rapidly changing thermal environment in the intertidal zone. In the present study, the temporal patterns of heart rate, protein carbonyl groups, and genes encoding heat shock proteins (hsp70 and hsp90) and metabolic sensors (ampkα, ampkβ and sirt1) were measured to study the effect of sublethal heat stress on the cardiac function, oxidative stress, heat shock response and cellular metabolism of an intertidal limpet Cellana toreuma. All the physiological parameters are sensitive to temperature and duration of heat stress. Spearman correlation analysis revealed that the correlations between heart rate and levels of heat shock proteins mRNA and metabolic sensors mRNA were statistically significant. These results further suggest that cardiac function plays crucial roles in cellular energy metabolism and heat shock responses. The significant increase of protein carbonyl groups at 34°C after 4h exposure indicated that the failure of cardiac function and the increase of anaerobic metabolism partly leads to the increase of protein carbonyl groups. Generally, the physiological responses to heat stress are sensitive to temperature and are energy-consumptive, as indicated by the upregulation of metabolic sensors mRNA. However, the upregulation of heat shock proteins and metabolic sensors at the post-transcriptional level and related functions need to be confirmed in further experiments. Copyright © 2014 Elsevier Ltd. All rights reserved.

  14. Reactive oxygen species promote heat shock protein 90-mediated HBV capsid assembly

    International Nuclear Information System (INIS)

    Kim, Yoon Sik; Seo, Hyun Wook; Jung, Guhung

    2015-01-01

    Hepatitis B virus (HBV) infection induces reactive oxygen species (ROS) production and has been associated with the development of hepatocellular carcinoma (HCC). ROS are also an important factor in HCC because the accumulated ROS leads to abnormal cell proliferation and chromosome mutation. In oxidative stress, heat shock protein 90 (Hsp90) and glutathione (GSH) function as part of the defense mechanism. Hsp90 prevents cellular component from oxidative stress, and GSH acts as antioxidants scavenging ROS in the cell. However, it is not known whether molecules regulated by oxidative stress are involved in HBV capsid assembly. Based on the previous study that Hsp90 facilitates HBV capsid assembly, which is an important step for the packing of viral particles, here, we show that ROS enrich Hsp90-driven HBV capsid formation. In cell-free system, HBV capsid assembly was facilitated by ROS with Hsp90, whereas it was decreased without Hsp90. In addition, GSH inhibited the function of Hsp90 to decrease HBV capsid assembly. Consistent with the result of cell-free system, ROS and buthionine sulfoximine (BS), an inhibitor of GSH synthesis, increased HBV capsid formation in HepG2.2.15 cells. Thus, our study uncovers the interplay between ROS and Hsp90 during HBV capsid assembly. - Highlights: • We examined H 2 O 2 and GSH modulate HBV capsid assembly. • H 2 O 2 facilitates HBV capsid assembly in the presence of Hsp90. • GSH inhibits function of Hsp90 in facilitating HBV capsid assembly. • H 2 O 2 and GSH induce conformation change of Hsp90

  15. Reactive oxygen species promote heat shock protein 90-mediated HBV capsid assembly

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Yoon Sik, E-mail: yumshak@naver.com; Seo, Hyun Wook, E-mail: suruk@naver.com; Jung, Guhung, E-mail: drjung@snu.ac.kr

    2015-02-13

    Hepatitis B virus (HBV) infection induces reactive oxygen species (ROS) production and has been associated with the development of hepatocellular carcinoma (HCC). ROS are also an important factor in HCC because the accumulated ROS leads to abnormal cell proliferation and chromosome mutation. In oxidative stress, heat shock protein 90 (Hsp90) and glutathione (GSH) function as part of the defense mechanism. Hsp90 prevents cellular component from oxidative stress, and GSH acts as antioxidants scavenging ROS in the cell. However, it is not known whether molecules regulated by oxidative stress are involved in HBV capsid assembly. Based on the previous study that Hsp90 facilitates HBV capsid assembly, which is an important step for the packing of viral particles, here, we show that ROS enrich Hsp90-driven HBV capsid formation. In cell-free system, HBV capsid assembly was facilitated by ROS with Hsp90, whereas it was decreased without Hsp90. In addition, GSH inhibited the function of Hsp90 to decrease HBV capsid assembly. Consistent with the result of cell-free system, ROS and buthionine sulfoximine (BS), an inhibitor of GSH synthesis, increased HBV capsid formation in HepG2.2.15 cells. Thus, our study uncovers the interplay between ROS and Hsp90 during HBV capsid assembly. - Highlights: • We examined H{sub 2}O{sub 2} and GSH modulate HBV capsid assembly. • H{sub 2}O{sub 2} facilitates HBV capsid assembly in the presence of Hsp90. • GSH inhibits function of Hsp90 in facilitating HBV capsid assembly. • H{sub 2}O{sub 2} and GSH induce conformation change of Hsp90.

  16. Photosynthetic and transcriptional responses of the marine diatom Thalassiosira pseudonana to the combined effect of temperature stress and copper exposure

    DEFF Research Database (Denmark)

    Leung, Priscilla T. Y.; Yi, Andy Xianliang; Ip, Jack C. H.

    2017-01-01

    ) was positively modulated by increasing temperatures. Hsp 90, hsp90-2 and sit1 (related to silica shell formation) were highly expressed at 30°C under 1000μg/L Cu, while the genes encoding light harvesting proteins (3HfcpA and 3HfcpB) and silaffin precursor sil3 were significantly up-regulated at 15°C under 200μg...

  17. Heat shock proteins in relation to heat stress tolerance of creeping bentgrass at different N levels.

    Science.gov (United States)

    Wang, Kehua; Zhang, Xunzhong; Goatley, Mike; Ervin, Erik

    2014-01-01

    Heat stress is a primary factor causing summer bentgrass decline. Changes in gene expression at the transcriptional and/or translational level are thought to be a fundamental mechanism in plant response to environmental stresses. Heat stress redirects protein synthesis in higher plants and results in stress protein synthesis, particularly heat shock proteins (HSPs). The goal of this work was to analyze the expression pattern of major HSPs in creeping bentgrass (Agrostis stolonifera L.) during different heat stress periods and to study the influence of nitrogen (N) on the HSP expression patterns. A growth chamber study on 'Penn-A4' creeping bentgrass subjected to 38/28°C day/night for 50 days, was conducted with four nitrate rates (no N-0, low N-2.5, medium N-7.5, and high N-12.5 kg N ha-1) applied biweekly. Visual turfgrass quality (TQ), normalized difference vegetation index (NDVI), photochemical efficiency of photosystem II (Fv/Fm), shoot electrolyte leakage (ShEL), and root viability (RV) were monitored, along with the expression pattern of HSPs. There was no difference in measured parameters between treatments until week seven, except TQ at week five. At week seven, grass at medium N had better TQ, NDVI, and Fv/Fm accompanied by lower ShEL and higher RV, suggesting a major role in improved heat tolerance. All the investigated HSPs (HSP101, HSP90, HSP70, and sHSPs) were up-regulated by heat stress. Their expression patterns indicated cooperation between different HSPs and their roles in bentgrass thermotolerance. In addition, their production seems to be resource dependent. This study could further improve our understanding about how different N levels affect bentgrass thermotolerance.

  18. TaHsfA6f is a transcriptional activator that regulates a suite of heat stress protection genes in wheat (Triticum aestivum L.) including previously unknown Hsf targets.

    Science.gov (United States)

    Xue, Gang-Ping; Drenth, Janneke; McIntyre, C Lynne

    2015-02-01

    Heat stress is a significant environmental factor adversely affecting crop yield. Crop adaptation to high-temperature environments requires transcriptional reprogramming of a suite of genes involved in heat stress protection. This study investigated the role of TaHsfA6f, a member of the A6 subclass of heat shock transcription factors, in the regulation of heat stress protection genes in Triticum aestivum (bread wheat), a poorly understood phenomenon in this crop species. Expression analysis showed that TaHsfA6f was expressed constitutively in green organs but was markedly up-regulated during heat stress. Overexpression of TaHsfA6f in transgenic wheat using a drought-inducible promoter resulted in up-regulation of heat shock proteins (HSPs) and a number of other heat stress protection genes that included some previously unknown Hsf target genes such as Golgi anti-apoptotic protein (GAAP) and the large isoform of Rubisco activase. Transgenic wheat plants overexpressing TaHsfA6f showed improved thermotolerance. Transactivation assays showed that TaHsfA6f activated the expression of reporter genes driven by the promoters of several HSP genes (TaHSP16.8, TaHSP17, TaHSP17.3, and TaHSP90.1-A1) as well as TaGAAP and TaRof1 (a co-chaperone) under non-stress conditions. DNA binding analysis revealed the presence of high-affinity TaHsfA6f-binding heat shock element-like motifs in the promoters of these six genes. Promoter truncation and mutagenesis analyses identified TaHsfA6f-binding elements that were responsible for transactivation of TaHSP90.1-A1 and TaGAAP by TaHsfA6f. These data suggest that TaHsfA6f is a transcriptional activator that directly regulates TaHSP, TaGAAP, and TaRof1 genes in wheat and its gene regulatory network has a positive impact on thermotolerance. © The Author 2014. Published by Oxford University Press on behalf of the Society for Experimental Biology.

  19. Automatic provisioning, deployment and orchestration for load-balancing THREDDS instances

    Science.gov (United States)

    Cofino, A. S.; Fernández-Tejería, S.; Kershaw, P.; Cimadevilla, E.; Petri, R.; Pryor, M.; Stephens, A.; Herrera, S.

    2017-12-01

    THREDDS is a widely used web server to provide to different scientific communities with data access and discovery. Due to THREDDS's lack of horizontal scalability and automatic configuration management and deployment, this service usually deals with service downtimes and time consuming configuration tasks, mainly when an intensive use is done as is usual within the scientific community (e.g. climate). Instead of the typical installation and configuration of a single or multiple independent THREDDS servers, manually configured, this work presents an automatic provisioning, deployment and orchestration cluster of THREDDS servers. This solution it's based on Ansible playbooks, used to control automatically the deployment and configuration setup on a infrastructure and to manage the datasets available in THREDDS instances. The playbooks are based on modules (or roles) of different backends and frontends load-balancing setups and solutions. The frontend load-balancing system enables horizontal scalability by delegating requests to backend workers, consisting in a variable number of instances for the THREDDS server. This implementation allows to configure different infrastructure and deployment scenario setups, as more workers are easily added to the cluster by simply declaring them as Ansible variables and executing the playbooks, and also provides fault-tolerance and better reliability since if any of the workers fail another instance of the cluster can take over it. In order to test the solution proposed, two real scenarios are analyzed in this contribution: The JASMIN Group Workspaces at CEDA and the User Data Gateway (UDG) at the Data Climate Service from the University of Cantabria. On the one hand, the proposed configuration has provided CEDA with a higher level and more scalable Group Workspaces (GWS) service than the previous one based on Unix permissions, improving also the data discovery and data access experience. On the other hand, the UDG has improved its

  20. Molecular characterization and expression analysis of a heat shock protein 90 gene from disk abalone (Haliotis discus).

    Science.gov (United States)

    Wang, Ning; Whang, Ilson; Lee, Jae-Seong; Lee, Jehee

    2011-06-01

    Heat shock protein 90s (hsp90s) are chaperones that contribute to the proper folding of cellular proteins and help animals cope with the cellular protein damages in stress conditions. In this study, an hsp90 gene was isolated from disc abalone (Haliotis discus). The complete nucleotide sequence of the hsp90 gene contains an open reading frame of 2,184 base pairs, encoding an 84 kDa protein. Disk abalone hsp90 shares high sequence similarity with other hsp90 family proteins. Although the phylogenetic analysis did not classify it into the hsp90α group, the inductivity of this gene was confirmed by heat shock and lipopolysaccharide (LPS) challenge test. Disk abalone hsp90 gene displayed a rapid and reversible induction response to both an exposure of typical heat shock and the LPS challenge. Once given the sublethal heat shock treatment, the transcription of disk abalone hsp90 gene was significantly up-regulated. With a recovery of 12 h, the transcription of disk abalone hsp90 gene gradually attenuated to the control level. These observations reflected the feedback regulation of abalone heat shock responses faithfully. In response to LPS challenge, the transcription of disk abalone hsp90 gene was significantly increased within 2 h and it approached maximum induction at 4 h later and recovered finally the reference level in 24 h. Take all together, the cloning and expression analysis of disk abalone hsp90 gene provided useful molecular information of abalone responses in stress conditions and potential ways to monitor the chronic stressors in abalone culture environments and diagnose the animal health status.

  1. Silica Nanoparticles Induce Oxidative Stress and Autophagy but Not Apoptosis in the MRC-5 Cell Line

    Directory of Open Access Journals (Sweden)

    Sorina Nicoleta Petrache Voicu

    2015-12-01

    Full Text Available This study evaluated the in vitro effects of 62.5 µg/mL silica nanoparticles (SiO2 NPs on MRC-5 human lung fibroblast cells for 24, 48 and 72 h. The nanoparticles’ morphology, composition, and structure were investigated using high resolution transmission electron microscopy, selected area electron diffraction and X-ray diffraction. Our study showed a decreased cell viability and the induction of cellular oxidative stress as evidenced by an increased level of reactive oxygen species (ROS, carbonyl groups, and advanced oxidation protein products after 24, 48, and 72 h, as well as a decreased concentration of glutathione (GSH and protein sulfhydryl groups. The protein expression of Hsp27, Hsp60, and Hsp90 decreased at all time intervals, while the level of protein Hsp70 remained unchanged during the exposure. Similarly, the expression of p53, MDM2 and Bcl-2 was significantly decreased for all time intervals, while the expression of Bax, a marker for apoptosis, was insignificantly downregulated. These results correlated with the increase of pro-caspase 3 expression. The role of autophagy in cellular response to SiO2 NPs was demonstrated by a fluorescence-labeled method and by an increased level of LC3-II/LC3-I ratio. Taken together, our data suggested that SiO2 NPs induced ROS-mediated autophagy in MRC-5 cells as a possible mechanism of cell survival.

  2. Investigating hsp Gene Expression in Liver of Channa striatus under Heat Stress for Understanding the Upper Thermal Acclimation

    Directory of Open Access Journals (Sweden)

    Gopal Krishna Purohit

    2014-01-01

    Full Text Available Changes in hsp gene expression profiles in murrel Channa striatus experimentally exposed to temperature stress (36°C for 4, 15, and 30 days were investigated; fish collected from aquaculture ponds and maintained in laboratory at the pond temperature (25 ± 1°C served as control. Channa collected from a hot spring runoff (36°C was included in the study to examine the hsp profiles beyond 30 days of exposure. Gene expression analyses of a battery of hsps in liver tissues were carried out by quantitative RT-PCR and protein expressions were analyzed by immunoblotting. hsps could be grouped into three clusters based on similarity in response to heat stress: hsp70, hsp78, and hsp60, whose transcript level continued to increase with duration of exposure; hsp90 and hsp110 that increased to a much higher level and then decreased; hsp27 and hsp47 that did not significantly vary as compared to control. The results suggest that Hsp70, Hsp78, and Hsp60 are involved in thermal acclimation and long term survival at high temperature. Fish living in the hot spring runoff appears to continuously express hsps that can be approximated by long term induction of hsps in farmed fish if temperature of their environment is raised to 36°C.

  3. Increased gray matter volume of left pars opercularis in male orchestral musicians correlate positively with years of musical performance.

    Science.gov (United States)

    Abdul-Kareem, Ihssan A; Stancak, Andrej; Parkes, Laura M; Sluming, Vanessa

    2011-01-01

    To compare manual volumetry of gray matter (GM) / white matter (WM) of Broca's area subparts: pars opercularis (POP) and pars triangularis (PTR) in both hemispheres between musicians and nonmusician, as it has been shown that these regions are crucial for musical abilities. A previous voxel-based morphometric (VBM) study conducted in our laboratory reported increased GM density in Broca's area of left hemisphere in male orchestral musicians. Functional segregation of POP/PTR justified separate volumetric analysis of these parts. We used the same cohort for the VBM study. Manual morphometry (stereology) was used to compare volumes between 26/26 right-handed orchestral musicians/nonmusicians. As expected, musicians showed significantly increased GM volume in the Broca's area, specifically in the left POP. No significant results were detected in right POP, left/right PTR GM volumes, and WM volumes for all regions. Results were positively correlated with years of musical performance (r = 0.7, P = 0.0001). This result corroborates the VBM study and is in line with the hypothesis of critical involvement of POP in hearing-action integration being an integral component of frontoparietotemporal mirror neuron network. We hypothesize that increased size of musicians' left POP represent use-dependent structural adaptation in response to intensive audiomotor skill acquisition. Copyright © 2010 Wiley-Liss, Inc.

  4. Nuclear Organization in the Spinal Cord Depends on Motor Neuron Lamination Orchestrated by Catenin and Afadin Function

    Directory of Open Access Journals (Sweden)

    Carola Dewitz

    2018-02-01

    Full Text Available Motor neurons in the spinal cord are found grouped in nuclear structures termed pools, whose position is precisely orchestrated during development. Despite the emerging role of pool organization in the assembly of spinal circuits, little is known about the morphogenetic programs underlying the patterning of motor neuron subtypes. We applied three-dimensional analysis of motor neuron position to reveal the roles and contributions of cell adhesive function by inactivating N-cadherin, catenin, and afadin signaling. Our findings reveal that nuclear organization of motor neurons is dependent on inside-out positioning, orchestrated by N-cadherin, catenin, and afadin activities, controlling cell body layering on the medio-lateral axis. In addition to this lamination-like program, motor neurons undergo a secondary, independent phase of organization. This process results in segregation of motor neurons along the dorso-ventral axis of the spinal cord, does not require N-cadherin or afadin activity, and can proceed even when medio-lateral positioning is perturbed.

  5. Nuclear Organization in the Spinal Cord Depends on Motor Neuron Lamination Orchestrated by Catenin and Afadin Function.

    Science.gov (United States)

    Dewitz, Carola; Pimpinella, Sofia; Hackel, Patrick; Akalin, Altuna; Jessell, Thomas M; Zampieri, Niccolò

    2018-02-13

    Motor neurons in the spinal cord are found grouped in nuclear structures termed pools, whose position is precisely orchestrated during development. Despite the emerging role of pool organization in the assembly of spinal circuits, little is known about the morphogenetic programs underlying the patterning of motor neuron subtypes. We applied three-dimensional analysis of motor neuron position to reveal the roles and contributions of cell adhesive function by inactivating N-cadherin, catenin, and afadin signaling. Our findings reveal that nuclear organization of motor neurons is dependent on inside-out positioning, orchestrated by N-cadherin, catenin, and afadin activities, controlling cell body layering on the medio-lateral axis. In addition to this lamination-like program, motor neurons undergo a secondary, independent phase of organization. This process results in segregation of motor neurons along the dorso-ventral axis of the spinal cord, does not require N-cadherin or afadin activity, and can proceed even when medio-lateral positioning is perturbed. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

  6. Dietary ascorbic acid modulates the expression profile of stress protein genes in hepatopancreas of adult Pacific abalone Haliotis discus hannai Ino.

    Science.gov (United States)

    Wu, Chenglong; Wang, Jia; Xu, Wei; Zhang, Wenbing; Mai, Kangsen

    2014-12-01

    This study was conducted to investigate the effects of dietary ascorbic acid (AA) on transcriptional expression patterns of antioxidant proteins, heat shock proteins (HSP) and nuclear factor kappa B (NF-κB) in the hepatopancreas of Pacific abalone Haliotis discus hannai Ino (initial average length: 84.36 ± 0.24 mm) using real-time quantitative PCR assays. L-ascorbyl-2-molyphosphate (LAMP) was added to the basal diet to formulate four experimental diets containing 0.0, 70.3, 829.8 and 4967.5 mg AA equivalent kg(-1) diets, respectively. Each diet was fed to triplicate groups of adult abalone in acrylic tanks (200 L) in a flow-through seawater system. Each tank was stocked with 15 abalone. Animals were fed once daily (17:00) to apparent satiation for 24 weeks. The results showed that the dietary AA (70.3 mg kg(-1)) could significantly up-regulate the expression levels of Cu/Zn superoxide dismutase (SOD), catalase (CAT), glutathione-S-transferase (GST), feritin (FT) and heat shock protein 26 (HSP26) in the hepatopancreas of abalone in this treatment compared to the controls. However, the expression levels of Mn-SOD, glutathione peroxidase (GPX), thioredoxin peroxidase (TPx), selenium-binding protein (SEBP), HSP70 and HSP90 were significantly down-regulated. Compared with those in the group with 70.3 mg kg(-1) dietary AA, the expression levels of CAT, GST and HSP26 were decreased in abalone fed with very high dietary AA (4967.5 mg kg(-1)). In addition, significant up-regulations of expression levels of Mn-SOD, GPX, TPx, SEBP, FT, HSP70, HSP90 and NF-κB were observed in abalone fed with apparently excessive dietary AA (829.8 and 4967.5 mg kg(-1)) as compared to those fed 70.3 mg kg(-1) dietary AA. These findings showed that dietary AA influenced the expression levels of antioxidant proteins, heat shock proteins and NF-κB in the hepatopancreas of abalone at transcriptional level. Levels of dietary AA that appeared adequate (70.3 mg kg(-1)) reduced the oxidative stress

  7. Aberrant expression and secretion of heat shock protein 90 in patients with bullous pemphigoid.

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    Stefan Tukaj

    Full Text Available The cell stress chaperone heat shock protein 90 (Hsp90 has been implicated in inflammatory responses and its inhibition has proven successful in different mouse models of autoimmune diseases, including epidermolysis bullosa acquisita. Here, we investigated expression levels and secretory responses of Hsp90 in patients with bullous pemphigoid (BP, the most common subepidermal autoimmune blistering skin disease. In comparison to healthy controls, the following observations were made: (i Hsp90 was highly expressed in the skin of BP patients, whereas its serum levels were decreased and inversely associated with IgG autoantibody levels against the NC16A immunodominant region of the BP180 autoantigen, (ii in contrast, neither aberrant levels of circulating Hsp90 nor any correlation of this protein with serum autoantibodies was found in a control cohort of autoimmune bullous disease patients with pemphigus vulgaris, (iii Hsp90 was highly expressed in and restrictedly released from peripheral blood mononuclear cells of BP patients, and (iv Hsp90 was potently induced in and restrictedly secreted from human keratinocyte (HaCaT cells by BP serum and isolated anti-BP180 NC16A IgG autoantibodies, respectively. Our results reveal an upregulated Hsp90 expression at the site of inflammation and an autoantibody-mediated dysregulation of the intracellular and extracellular distribution of this chaperone in BP patients. These findings suggest that Hsp90 may play a pathophysiological role and represent a novel potential treatment target in BP.

  8. Heat shock protein 90 in neurodegenerative diseases

    Directory of Open Access Journals (Sweden)

    Rodina Anna

    2010-06-01

    Full Text Available Abstract Hsp90 is a molecular chaperone with important roles in regulating pathogenic transformation. In addition to its well-characterized functions in malignancy, recent evidence from several laboratories suggests a role for Hsp90 in maintaining the functional stability of neuronal proteins of aberrant capacity, whether mutated or over-activated, allowing and sustaining the accumulation of toxic aggregates. In addition, Hsp90 regulates the activity of the transcription factor heat shock factor-1 (HSF-1, the master regulator of the heat shock response, mechanism that cells use for protection when exposed to conditions of stress. These biological functions therefore propose Hsp90 inhibition as a dual therapeutic modality in neurodegenerative diseases. First, by suppressing aberrant neuronal activity, Hsp90 inhibitors may ameliorate protein aggregation and its associated toxicity. Second, by activation of HSF-1 and the subsequent induction of heat shock proteins, such as Hsp70, Hsp90 inhibitors may redirect neuronal aggregate formation, and protect against protein toxicity. This mini-review will summarize our current knowledge on Hsp90 in neurodegeneration and will focus on the potential beneficial application of Hsp90 inhibitors in neurodegenerative diseases.

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

    Directory of Open Access Journals (Sweden)

    Haiping Tang

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

  10. Positive associations between upregulated levels of stress-induced phosphoprotein 1 and matrix metalloproteinase-9 in endometriosis/adenomyosis.

    Science.gov (United States)

    Wang, Hsin-Shih; Tsai, Chia-Lung; Chang, Pi-Yueh; Chao, Angel; Wu, Ren-Chin; Chen, Shun-Hua; Wang, Chin-Jung; Yen, Chih-Feng; Lee, Yun-Shien; Wang, Tzu-Hao

    2018-01-01

    Stress-induced phosphoprotein-1 (STIP1), an adaptor protein that coordinates the functions of HSP70 and HSP90 in protein folding, has been implicated in the development of human gynecologic malignancies. This case-control study investigates STIP1 serum levels and tissue expression in relation to endometriosis/adenomyosis in Taiwanese population. Female patients with surgically confirmed endometriosis/adenomyosis were compared with women free of endometriosis/adenomyosis. Serum STIP1 levels were measured using an enzyme-linked immunosorbent assay and surgical tissues were analyzed by immunohistochemistry. Both epithelial and stromal cells in surgical tissues of endometriosis and adenomyosis expressed STIP1 and MMP-9. Notably, MMP-9 expression was significantly decreased when STIP1 expression was knocked-down. In vitro experiments revealed that STIP1 was capable of binding to the MMP-9 promoter and enhanced its transcriptional expression. The preoperative serum STIP1 levels of patients with endometriosis/adenomyosis were significantly higher than those of the controls. In brief, our data suggest an association between STIP1 levels and endometriosis/adenomyosis.

  11. Characterization of six small HSP genes from Chironomus riparius (Diptera, Chironomidae): Differential expression under conditions of normal growth and heat-induced stress.

    Science.gov (United States)

    Martín-Folgar, Raquel; de la Fuente, Mercedes; Morcillo, Gloria; Martínez-Guitarte, José-Luis

    2015-10-01

    Small heat shock proteins (sHSPs) comprise the most numerous, structurally diverse, and functionally uncharacterized family of heat shock proteins. Several Hsp genes (Hsp 90, 70, 40, and 27) from the insect Chironomus riparius are widely used in aquatic toxicology as biomarkers for environmental toxins. Here, we conducted a comparative study and characterized secondary structure of the six newly identified sHsp genes Hsp17, Hsp21, Hsp22, Hsp23, Hsp24, and Hsp34. A characteristic α-crystallin domain is predicted in all the new proteins. Phylogenetic analysis suggests a strong relation to other sHSPs from insects and interesting evidence regarding evolutionary origin and duplication events. Comparative analysis of transcription profiles for Hsp27, Hsp70, and the six newly identified genes revealed that Hsp17, Hsp21, and Hsp22 are constitutively expressed under normal conditions, while under two different heat shock conditions these genes are either not activated or are even repressed (Hsp22). In contrast, Hsp23, Hsp24, and Hsp34 are significantly activated along with Hsp27 and Hsp70 during heat stress. These results strongly suggest functional differentiation within the small HSP subfamily and provide new data to help understand the coping mechanisms induced by stressful environmental stimuli. Copyright © 2015 Elsevier Inc. All rights reserved.

  12. Characterize and Gene Expression of Heat Shock Protein 90 in Marine Crab Charybdis japonica following Bisphenol A and 4-Nonylphenol Exposures.

    Science.gov (United States)

    Park, Kiyun; Kwak, Ihn-Sil

    2014-01-01

    Heat shock protein 90 (HSP90) is a highly conserved molecular chaperone important in the maturation of a broad spectrum of protein. In this study, an HSP90 gene was isolated from Asian paddle crab, Charybdis japonica, as a bio-indicator to monitor the marine ecosystem. This work reports the responses of C. japonica HSP90 mRNA expression to cellular stress by endocrine disrupting chemicals (EDCs), such as bisphenol A (BPA) and 4-nonylphenol (NP) using real-time. reverse transcription polymerase chain reaction. The deduced amino acid sequence of HSP90 from C. japonica shared a high degree of homology with their homologues in other species. In a phylogenetic analysis, C. japonica HSP90 is evolutionally related with an ortholog of the other crustacean species. The expression of HSP90 gene was almost distributed in all the examined tissues of the C. japonica crab but expression levels varied among the different body parts of the crabs. We examined HSP90 mRNA expression pattern in C. japonica crabs exposed to EDCs for various exposure times. The expression of HSP90 transcripts was significantly increased in C. japonica crabs exposed to BPA and NP at different concentrations for 12, 24, 48 and 96 hours. The mRNA expression of HSP90 gene was significantly induced in a concentration- and time-dependent manner after BPA or NP exposures for 96 hours. Taken together, expression analysis of Asian paddle crab HSP90 gene provided useful molecular information about crab responses in stress conditions and potential ways to monitor the EDCs stressors in marine environments.

  13. First field demonstration of cloud datacenter workflow automation employing dynamic optical transport network resources under OpenStack and OpenFlow orchestration.

    Science.gov (United States)

    Szyrkowiec, Thomas; Autenrieth, Achim; Gunning, Paul; Wright, Paul; Lord, Andrew; Elbers, Jörg-Peter; Lumb, Alan

    2014-02-10

    For the first time, we demonstrate the orchestration of elastic datacenter and inter-datacenter transport network resources using a combination of OpenStack and OpenFlow. Programmatic control allows a datacenter operator to dynamically request optical lightpaths from a transport network operator to accommodate rapid changes of inter-datacenter workflows.

  14. Characterization