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Sample records for hsp-70 gene expression

  1. [Cloning and expression analysis of HSP70 gene from Dendrobium officinale under low temperature stress].

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    Li, Dong-Bin; Gao, Han-Hui; Si, Jin-Ping; Zhu, Yu-Qiu

    2013-10-01

    To investigate HSP70 gene expression from Dendrobium officinale under low temperature stress, which will provide the molecular biological foundation for breeding the low temperature resistant strain. HSP70 gene full length cDNA was cloned by rapid amplification of cDNA ends (RACE) on the basis of HSP70 gene fragment sequences, and the structure and function of HSP70 gene were deduced. The expression of HSP70 under low temperature stress was detected by RT-PCR. The full length of HSP70 gene cDNA was 2 296 bp containing a 1 944 bp open reading frame (ORF) that encoded a protein of 647 amino acids. Its amino acids sequence had typical HSP70 characteristics and high homology with other plant's HSP70. Cold stress expression analysis showed that expression of the HSP70 gene could be induced by low temperature. The HSP70 gene of D. officinale was successfully cloned and reported for the first time which proved that the expression could be induced by low temperature. The cloning of HSP70 gene provides a stable foundation for further study of D. officinale cultivation and the breeding of the cold resistance strains.

  2. Hsp 70 expression and function during gametogenesis

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    Dix, David J.

    1997-01-01

    The dramatic transformations in nuclear content and cellular organization that occur during gametogenesis require unique regulation and execution of the mitotic and meiotic cell cycle, apoptotic cell death, DNA recombination and repair, and cellular differentiation. These processes are accompained by the constitutive and developmentally regulated expression of a number of hsp70 genes encoding 70 kDa heat shock proteins (Hsp70), including several hsp70s whose expression is unique to male germ cells. Examining the expression and function of Hsp70s in germ cells has provided significant insights into mechanisms of hsp70 gene regulation and Hsp70 protein function, as well as the developmental processes of gametogenesis. PMID:9250397

  3. PROFILES OF GENE EXPRESSION ASSOCIATED WITH TETRACYCLINE OVER EXPRESSION OF HSP70 IN MCF-7 BREAST CANCER CELLS

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    Profiles of gene expression associated with tetracycline over expression of HSP70 in MCF-7 breast cancer cells. Heat shock proteins (HSPs) protect cells from damage through their function as molecular chaperones. Some cancers reveal high levels of HSP70 expression in asso...

  4. Molecular cloning, sequencing, and expression of Eimeria tenella HSP70 partial gene.

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    Bogado, A L G; Martins, G F; Sasse, J P; Guimarães, J da S; Garcia, J L

    2017-03-15

    Members of the Eimeria genus are protozoan parasites of the subphylum Apicomplexa (Eimeriidae family), and belong to the coccidia group. Eimeria tenella is one of the most pathogenic species owing to its ability to penetrate the mucosa, and cause inflammation and damage. It is an obligate intracellular parasite that causes disease by destroying the host cells during multiplication. Heat shock protein 70 (HSP70) is a molecular chaperone that prevents cellular stress. The objective of this study was to clone, sequence, and express E. tenella HSP70 protein. After selecting the region of highest hydrophilicity in the hsp70 gene, we cloned complementary DNA (cDNA) into a pTrcHis2-TOPO vector and transformed it into TOP10 Escherichia coli cells; after induction, the bacteria expressed a 23-kDa protein with insoluble expression levels of approximately 5 mg/L. In summary, the partial hsp70 gene was successfully expressed in E. coli, producing a 23-kDa protein under insoluble conditions, and the antigen characteristics predicted by hydrophilicity analysis suggest the development of a vaccine for use in avian coccidiosis.

  5. Molecular cloning and expression of two HSP70 genes in the prawn, Macrobrachium rosenbergii

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    Liu, Jun; Yang, Wei-Jun; Zhu, Xiao-Jing; Karouna-Renier, Natalie K.; Rao, Ranga K.

    2004-01-01

    Two complementary deoxyribonucleic acid (cDNA) clones encoding 2 different 70-kDa heat shock proteins (HSPs) were isolated from the prawn Macrobrachium rosenbergii. The cDNA clones were 2448 and 2173 bp in length and contained 1950- and 1734-bp open reading frames (ORFs), respectively. The ORFs encoded 649– and 577–amino acid polypeptides, which were named Mar-HSC70 and Mar-HSP70, respectively, according to the sequence identities with other known HSC70s and HSP70s and based on their inducibility in response to heat shock stress (at 35°C). Genomic DNA sequence analysis revealed no introns in either gene. The major structural differences between the 2 proteins were a 60–amino acid segment and a 14–amino acid segment present in the N-terminal and C-terminal, respectively, of Mar-HSC70 that were not found in Mar-HSP70. Northern blotting and semiquantitative reverse transcription–polymerase chain reaction analyses indicated that the Mar-HSP70 gene was expressed under heat shock (35°C) stress in a non–tissue-specific manner. In contrast, Mar-HSC70 messenger ribonucleic acid was constitutively expressed in every tissue except muscle, and its expression in response to heat shock (at 35°C) changed only in muscle. PMID:15544169

  6. B-chromosome effects on Hsp70 gene expression does not occur at transcriptional level in the grasshopper Eyprepocnemis plorans.

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    Navarro-Domínguez, Beatriz; Cabrero, Josefa; Camacho, Juan Pedro M; López-León, María Dolores

    2016-10-01

    As intragenomic parasites, B chromosomes can elicit stress in the host genome, thus inducing a response for host adaptation to this kind of continuous parasitism. In the grasshopper Eyprepocnemis plorans, B-chromosome presence has been previously associated with a decrease in the amount of the heat-shock protein 70 (HSP70). To investigate whether this effect is already apparent at transcriptional level, we analyze the expression levels of the Hsp70 gene in gonads and somatic tissues of males and females with and without B chromosomes from two populations, where the predominant B chromosome variants (B2 and B24) exhibit different levels of parasitism, by means of quantitative real-time PCR (qPCR) on complementary DNA (cDNA). The results revealed the absence of significant differences for Hsp70 transcripts associated with B-chromosome presence in virtually all samples. This indicates that the decrease in HSP70 protein levels, formerly reported in this species, may not be a consequence of transcriptional down-regulation of Hsp70 genes, but the result of post-transcriptional regulation. These results will help to design future studies oriented to identifying factors modulating Hsp70 expression, and will also contribute to uncover the biological role of B chromosomes in eukaryotic genomes.

  7. [Downregulation of HSP70 gene expression and apoptosis in human hepatocellular carcinoma SMMC-7721 cells induced by nimesulide in vitro].

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    Yin, Guo-zhi; Tu, Kang-sheng; Han, Shao-shan; Wang, Jun; Liu, Qing-guang; Yao, Ying-min

    2012-09-01

    To investigate the effect of nimesulide on cell apoptosis and possible mechanism in human hepatocellular carcinoma SMMC-7721 cells. SMMC-7721 cells were treated with nimesulide at different concentrations. Cell viability was assessed by MTT assay. Cell apoptosis rate was determined with flow cytometry. The cleavage activity of PARP and caspase-9 and the expression of HSP70 were evaluated using RT-PCR and Western blotting. The influence of HSP70 on cell apoptosis was observed using RNA interference silencing HSP70 expression. Nimesulide significantly inhibited cell growth in SMMC-7721 cells in a time- and concentration-dependent manner, and induced cell apoptosis in a concentration-dependent manner. Moreover, nimesulide promoted the cleavage of caspase-9 and PARP and inhibited the mRNA and protein expression of HSP70. Through the specific inhibition on HSP70 gene with siRNA, cell apoptosis increased, and the apoptosis was enhanced by the cleavage activity of caspase-9 and PARP. Nimesulide could inhibit cell growth and induce apoptosis in human hepatocellular carcinoma SMMC-7721 cells via the downregulation of HSP70.

  8. Expression profiling of Plasmodium berghei HSP70 genes for generation of bright red fluorescent parasites.

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    Marion Hliscs

    Full Text Available Live cell imaging of recombinant malarial parasites encoding fluorescent probes provides critical insights into parasite-host interactions and life cycle progression. In this study, we generated a red fluorescent line of the murine malarial parasite Plasmodium berghei. To allow constitutive and abundant expression of the mCherry protein we profiled expression of all members of the P. berghei heat shock protein 70 (HSP70 family. We identified PbHSP70/1, an invariant ortholog of Plasmodium falciparum HSP70-1, as the protein with the highest expression levels during Plasmodium blood, mosquito, and liver infection. Stable allelic insertion of a mCherry expression cassette into the PbHsp70/1 locus created constitutive red fluorescent P. berghei lines, termed Pbred. We show that these parasites can be used for live imaging of infected host cells and organs, including hepatocytes, erythrocytes, and whole Anopheles mosquitoes. Quantification of the fluorescence intensity of several Pbred parasite stages revealed significantly enhanced signal intensities in comparison to GFP expressed under the control of the constitutive EF1alpha promoter. We propose that systematic transcript profiling permits generation of reporter parasites, such as the Pbred lines described herein.

  9. HSP70 Promoter-Driven Activation of Gene Expression for Immunotherapy Using Gold Nanorods and Near Infrared Light

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    Helen A. Andersson

    2014-03-01

    Full Text Available Modulation of the cytokine milieu is one approach for vaccine development. However, therapy with pro-inflammatory cytokines, such as IL-12, is limited in practice due to adverse systemic effects. Spatially-restricted gene expression circumvents this problem by enabling localized amplification. Intracellular co-delivery of gold nanorods (AuNR and a heat shock protein 70 (HSP70 promoter-driven expression vector enables gene expression in response to near infrared (NIR light. AuNRs absorb the light, convert it into heat and thereby stimulate photothermal expression of the cytokine. As proof-of-concept, human HeLa and murine B16 cancer cells were transfected with a HSP70-Enhanced Green Fluorescent Protein (EGFP plasmid and polyethylenimine (PEI-conjugated AuNRs. Exposure to either 42 °C heat-shock or NIR light induced significant expression of the reporter gene. In vivo NIR driven expression of the reporter gene was confirmed at 6 and 24 h in mice bearing B16 melanoma tumors using in vivo imaging and flow-cytometric analysis. Overall, we demonstrate a novel opportunity for site-directed, heat-inducible expression of a gene based upon the NIR-absorbing properties of AuNRs and a HSP70 promoter-driven expression vector.

  10. GENE EXPRESSION PROFILES IN ARSENIC-TREATED MCF-7 BREAST CANCER CELLS EXPRESSING DIFFERENT LEVELS OF HSP70

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    Gene expression profiles in arsenic-treated MCF-7 breast cancer cells expressing different levels of HSP70Gail Nelson, Susan Hester, Ernest Winkfield, Jill Barnes, James AllenEnvironmental Carcinogenesis Division, NHEERL, ORD, US Environmental Protection Agency, Rese...

  11. Characterization of a wheat HSP70 gene and its expression in response to stripe rust infection and abiotic stresses

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    Duan, Y.H.; Guo, J.; Ding, K.; Wang, S.J.; Zhang, H.; Dai, X.W.; Chen, Y.Y.; Govers, F.; Huang, L.L.; Kang, Z.S.

    2011-01-01

    Members of the family of 70-kD heat shock proteins (HSP70 s) play various stress-protective roles in plants. In this study, a wheat HSP70 gene was isolated from a suppression subtractive hybridization (SSH) cDNA library of wheat leaves infected by Puccinia striiformis f. sp. tritici. The gene, that

  12. Characterization of a wheat HSP70 gene and its expression in response to stripe rust infection and abiotic stresses.

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    Duan, Ying-Hui; Guo, Jun; Ding, Ke; Wang, Shu-Juan; Zhang, Hong; Dai, Xi-Wei; Chen, Yue-Ying; Govers, Francine; Huang, Li-Li; Kang, Zhen-Sheng

    2011-01-01

    Members of the family of 70-kD heat shock proteins (HSP70 s) play various stress-protective roles in plants. In this study, a wheat HSP70 gene was isolated from a suppression subtractive hybridization (SSH) cDNA library of wheat leaves infected by Puccinia striiformis f. sp. tritici. The gene, that was designated as TaHSC70, was predicted to encode a protein of 690 amino acids, with a molecular mass of 73.54 KDa and a pI of 5.01. Further analysis revealed the presence of a conserved signature that is characteristic for HSP70s and phylogenetic analysis demonstrated that TaHSC70 is a homolog of chloroplast HSP70s. TaHSC70 mRNA was present in leaves of both green and etiolated wheat seedlings and in stems and roots. The transcript level in roots was approximately threefold less than in leaves but light-dark treatment did not charge TaHSC70 expression. Following heat shock of wheat seedlings at 40°C, TaHSC70 expression increased in leaves of etiolated seedlings but remained stable at the same level in green seedlings. In addition, TaHSC70 was differentially expressed during an incompatible and compatible interaction with wheat-stripe rust, and there was a transient increase in expression upon treatment with methyl jasmonate (MeJA) treatment. Salicylic acid (SA), ethylene (ET) and abscisic acid (ABA) treatments had no influence on TaHSC70 expression. These results suggest that TaHSC70 plays a role in stress-related responses, and in defense responses elicited by infection with stripe rust fungus and does so via a JA-dependent signal transduction pathway.

  13. Mechanism of hsp70i gene bookmarking.

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    Xing, Hongyan; Wilkerson, Donald C; Mayhew, Christopher N; Lubert, Eric J; Skaggs, Hollie S; Goodson, Michael L; Hong, Yiling; Park-Sarge, Ok-Kyong; Sarge, Kevin D

    2005-01-21

    In contrast to most genomic DNA in mitotic cells, the promoter regions of some genes, such as the stress-inducible hsp70i gene that codes for a heat shock protein, remain uncompacted, a phenomenon called bookmarking. Here we show that hsp70i bookmarking is mediated by a transcription factor called HSF2, which binds this promoter in mitotic cells, recruits protein phosphatase 2A, and interacts with the CAP-G subunit of the condensin enzyme to promote efficient dephosphorylation and inactivation of condensin complexes in the vicinity, thereby preventing compaction at this site. Blocking HSF2-mediated bookmarking by HSF2 RNA interference decreases hsp70i induction and survival of stressed cells in the G1 phase, which demonstrates the biological importance of gene bookmarking.

  14. Conserved structure and expression of hsp70 paralogs in teleost fishes.

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    Metzger, David C H; Hemmer-Hansen, Jakob; Schulte, Patricia M

    2016-06-01

    The cytosolic 70KDa heat shock proteins (Hsp70s) are widely used as biomarkers of environmental stress in ecological and toxicological studies in fish. Here we analyze teleost genome sequences to show that two genes encoding inducible hsp70s (hsp70-1 and hsp70-2) are likely present in all teleost fish. Phylogenetic and synteny analyses indicate that hsp70-1 and hsp70-2 are distinct paralogs that originated prior to the diversification of the teleosts. The promoters of both genes contain a TATA box and conserved heat shock elements (HSEs), but unlike mammalian HSP70s, both genes contain an intron in the 5' UTR. The hsp70-2 gene has undergone tandem duplication in several species. In addition, many other teleost genome assemblies have multiple copies of hsp70-2 present on separate, small, genomic scaffolds. To verify that these represent poorly assembled tandem duplicates, we cloned the genomic region surrounding hsp70-2 in Fundulus heteroclitus and showed that the hsp70-2 gene copies that are on separate scaffolds in the genome assembly are arranged as tandem duplicates. Real-time quantitative PCR of F. heteroclitus genomic DNA indicates that four copies of the hsp70-2 gene are likely present in the F. heteroclitus genome. Comparison of expression patterns in F. heteroclitus and Gasterosteus aculeatus demonstrates that hsp70-2 has a higher fold increase than hsp70-1 following heat shock in gill but not in muscle tissue, revealing a conserved difference in expression patterns between isoforms and tissues. These data indicate that ecological and toxicological studies using hsp70 as a biomarker in teleosts should take this complexity into account. Copyright © 2016 Elsevier Inc. All rights reserved.

  15. EFFECT OF STRESS ON THE PERCENT BODY WEIGHT CHANGE AND MRNA EXPRESSION OF IGF-1, SURVIVINE AND HSP-70 GENE IN THE HIERARCHIAL FOLLICLES OF JAPANESE QUAIL

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    N Shit

    2014-12-01

    Full Text Available The present study was carried out to explore the effect of stress on body weight and the mRNA expression of IGF-1, Survivine and HSP-70 gene in the hierarchial follicles of Japanese quail. A total 24 birds (10 weeks were taken and stress was induced by immobilization daily for 2hrs (between 9.00 - 11.00 AM throughout the study (10 days. Four birds were sacrificed on 1, 2, 4, 6, 8 and 10 days of the treatment. Hierarchial follicles (F1, F2 & F3 were aseptically collected to quantify the expression of IGF-1, Survivine and HSP-70 gene using real-time PCR technique. The percent body weight reduction increased and reached highest (21.30% on 10th day. The fold expression of IGF-1 gene was significantly ((P=0.05 down regulated in advance to the time of experiment. However, the fold expression of survivine gene was significantly (P=0.05 up regulated and the intensity was highest (17 fold in F-3 follicle on 4th day of experiment. No significant change in the mRNA expression of HSP-70 gene was evident in this study. From this study it may be concluded that stress brings physio-molecular change through HPA activation, which not only causes tissue regression also modifies the cellular mechanism.

  16. Identification and in silico analysis of the Citrus HSP70 molecular chaperone gene family

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    Luciano G. Fietto

    2007-01-01

    Full Text Available The completion of the genome sequencing of the Arabidopsis thaliana model system provided a powerful molecular tool for comparative analysis of gene families present in the genome of economically relevant plant species. In this investigation, we used the sequences of the Arabidopsis Hsp70 gene family to identify and annotate the Citrus Hsp70 genes represented in the CitEST database. Based on sequence comparison analysis, we identified 18 clusters that were further divided into 5 subgroups encoding four mitochondrial mtHsp70s, three plastid csHsp70s, one ER luminal Hsp70 BiP, two HSP110/SSE-related proteins and eight cytosolic Hsp/Hsc70s. We also analyzed the expression profile by digital Northern of each Hsp70 transcript in different organs and in response to stress conditions. The EST database revealed a distinct population distribution of Hsp70 ESTs among isoforms and across the organs surveyed. The Hsp70-5 isoform was highly expressed in seeds, whereas BiP, mitochondrial and plastid HSp70 mRNAs displayed a similar expression profile in the organs analyzed, and were predominantly represented in flowers. Distinct Hsp70 mRNAs were also differentially expressed during Xylella infection and Citrus tristeza viral infection as well as during water deficit. This in silico study sets the groundwork for future investigations to fully characterize functionally the Citrus Hsp70 family and underscores the relevance of Hsp70s in response to abiotic and biotic stresses in Citrus.

  17. Convergent evolution of heat-inducibility during subfunctionalization of the Hsp70 gene family.

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    Krenek, Sascha; Schlegel, Martin; Berendonk, Thomas U

    2013-02-21

    Heat-shock proteins of the 70 kDa family (Hsp70s) are essential chaperones required for key cellular functions. In eukaryotes, four subfamilies can be distinguished according to their function and localisation in different cellular compartments: cytosol, endoplasmic reticulum, mitochondria and chloroplasts. Generally, multiple cytosol-type Hsp70s can be found in metazoans that show either constitutive expression and/or stress-inducibility, arguing for the evolution of different tasks and functions. Information about the hsp70 copy number and diversity in microbial eukaryotes is, however, scarce, and detailed knowledge about the differential gene expression in most protists is lacking. Therefore, we have characterised the Hsp70 gene family of Paramecium caudatum to gain insight into the evolution and differential heat stress response of the distinct family members in protists and to investigate the diversification of eukaryotic hsp70s focusing on the evolution of heat-inducibility. Eleven putative hsp70 genes could be detected in P. caudatum comprising homologs of three major Hsp70-subfamilies. Phylogenetic analyses revealed five evolutionarily distinct Hsp70-groups, each with a closer relationship to orthologous sequences of Paramecium tetraurelia than to another P. caudatum Hsp70-group. These highly diverse, paralogous groups resulted from duplications preceding Paramecium speciation, underwent divergent evolution and were subject to purifying selection. Heat-shock treatments were performed to test for differential expression patterns among the five Hsp70-groups as well as for a functional conservation within Paramecium. These treatments induced exceptionally high mRNA up-regulations in one cytosolic group with a low basal expression, indicative for the major heat inducible hsp70s. All other groups showed comparatively high basal expression levels and moderate heat-inducibility, signifying constitutively expressed genes. Comparative EST analyses for P. tetraurelia

  18. Characterization of a heat-shock-inducible hsp70 gene of the green alga Volvox carteri.

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    Cheng, Qian; Hallmann, Armin; Edwards, Lisseth; Miller, Stephen M

    2006-04-12

    The green alga Volvox carteri possesses several thousand cells, but just two cell types: large reproductive cells called gonidia, and small, biflagellate somatic cells. Gonidia are derived from large precursor cells that are created during embryogenesis by asymmetric cell divisions. The J domain protein GlsA (Gonidialess A) is required for these asymmetric divisions and is believed to function with an Hsp70 partner. As a first step toward identifying this partner, we cloned and characterized V. carteri hsp70A, which is orthologous to HSP70A of the related alga Chlamydomonas reinhardtii. Like HSP70A, V. carteri hsp70A contains multiple heat shock elements (HSEs) and is highly inducible by heat shock. Consistent with these properties, Volvox transformants that harbor a glsA antisense transgene that is driven by an hsp70A promoter fragment express Gls phenotypes that are temperature-dependent. hsp70A appears to be the only gene in the genome that encodes a cytoplasmic Hsp70, so we conclude that Hsp70A is clearly the best candidate to be the chaperone that participates with GlsA in asymmetric cell division.

  19. Identification of the HSP70-II gene in Leishmania braziliensis HSP70 locus: genomic organization and UTRs characterization

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    Puerta Concepción J

    2011-08-01

    Full Text Available Abstract Background The heat stress suffered by Leishmania sp during its digenetic life-cycle is a key trigger for its stage differentiation. In Leishmania subgenera two classes of HSP70 genes differing in their 3' UTR were described. Although the presence of HSP70-I genes was previously suggested in Leishmania (Viannia braziliensis, HSP70-II genes had been reluctant to be uncovered. Results Here, we report the existence of two types of HSP70 genes in L. braziliensis and the genomic organization of the HSP70 locus. RT-PCR experiments were used to map the untranslated regions (UTR of both types of genes. The 3' UTR-II has a low sequence identity (55-57% when compared with this region in other Leishmania species. In contrast, the 5' UTR, common to both types of genes, and the 3' UTR-I were found to be highly conserved among all Leishmania species (77-81%. Southern blot assays suggested that L. braziliensis HSP70 gene cluster may contain around 6 tandemly-repeated HSP70-I genes followed by one HSP70-II gene, located at chromosome 28. Northern blot analysis indicated that levels of both types of mRNAs are not affected by heat shock. Conclusions This study has led to establishing the composition and structure of the HSP70 locus of L. braziliensis, complementing the information available in the GeneDB genome database for this species. L. braziliensis HSP70 gene regulation does not seem to operate by mRNA stabilization as occurs in other Leishmania species.

  20. Expression analysis of HSP70 in the testis of Octopus tankahkeei under thermal stress.

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    Long, Ling-Li; Han, Ying-Li; Sheng, Zhang; Du, Chen; Wang, You-Fa; Zhu, Jun-Quan

    2015-09-01

    The gene encoding heat shock protein 70 (HSP70) was identified in Octopus tankahkeei by homologous cloning and rapid amplification of cDNA ends (RACE). The full-length cDNA (2471 bp) consists of a 5'-untranslated region (UTR) (89 bp), a 3'-UTR (426 bp), and an open reading frame (1956 bp) that encodes 651 amino acid residues with a predicted molecular mass of 71.8 kDa and an isoelectric point of 5.34. Based on the amino acid sequence analysis and multiple sequence alignment, this cDNA is a member of cytoplasmic hsp70 subfamily of the hsp70 family and was designated as ot-hsp70. Tissue expression analysis showed that HSP70 expression is highest in the testes when all examined organs were compared. Immunohistochemistry analysis, together with hematoxylin-eosin staining, revealed that the HSP70 protein was expressed in all spermatogenic cells, but not in fibroblasts. In addition, O. tankahkeei were heat challenged by exposure to 32 °C seawater for 2 h, then returned to 13 °C for various recovery time (0-24 h). Relative expression of ot-hsp70 mRNA in the testes was measured at different time points post-challenge by quantitative real-time PCR. A clear time-dependent mRNA expression of ot-hsp70 after thermal stress indicates that the HSP70 gene is inducible. Ultrastructural changes of the heat-stressed testis were observed by transmission electron microscopy. We suggest that HSP70 plays an important role in spermatogenesis and testis protection against thermal stress in O. tankahkeei. Copyright © 2015 Elsevier Inc. All rights reserved.

  1. Comparative Expression Analysis of HSP70, HSP90, IL-4, TNF, KITLG and KIT-receptor Gene between Varicocele-Induced and Non-Varicocele Testes of Dog

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    Hossein Hassanpour

    2017-09-01

    Full Text Available Background This study was designed to create an experimental varicocele model by a simple surgical procedure in dog with minimum invasion and to investigate the effect of varicocele-induced infertility on the expression of six related genes (HSP90, HSP70, IL-4, TNF, KITLG and KIT receptor. Materials and Methods In this experimental study, the proximal part of the pampini-form plexus of dog testes was partially occluded without abdominal incision which was confirmed by venographic examination. To evaluate varicocele in its acute form, dogs were castrated after 15 days and testes were dissected. Histopathologic evaluation was undertaken and the relative expression of the six genes was assessed by quantitative realtime polymerase chain reaction (PCR. Results Microscopic changes showed tubule degeneration. The Johnson score was significantly decreased in the varicocele testes when compared with non-varicocele testes. Expressions of HSP90, TNF, KITLG and the KIT-receptor gene were significantly downregulated (P=0.029, 0.047, 0.004 and 0.035 respectively in varicocele-induced testes while HSP70 was upregulated (P=0.018. IL-4 did not show differential expression (P=0.377. Conclusion We conclude that partial occlusion of the proximal part of the pampiniform plexus induces varicocele in the testis of dog. Differential expression of the mentioned genes may be responsible for the pathophysiology of varicocele and related subfertility.

  2. Hsp27 and Hsp70 Expression in Esophageal Squamous

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    Luz, Caio Cesar Floriano; Noguti, Juliana; Borges de Araújo, Leandro; Gianni, Mara Silva Dos Santos; Simão Gomes, Thiago; Ricardo, Artigiani Neto

    2017-03-01

    Twenty-eight specimens of Esophael squamous cell carcinoma (ESCC) were obtained by surgery procedures.The tissues were fixed in formalin and embedded in paraffin. In each case, all available hematoxylin and eosin stained sections were examined and a representative block was selected. The ages of these patients ranged from 40 to 93 years, with a mean age of 60 years. Results. The histological grade of tumors was 4 well-differentiated, 19 moderately differentiated and 5 poorly differentiated. Expression of Hsp27 and Hsp70 in ESCC was demonstrated in 23 (82,14%) and 26 (92,86%) cases, respectively. Adjacent normal mucosa was positive in 11 (39,29%) samples and 9 (32,15%) samples for Hsp27 and Hsp70, respectively. No relationship between the expression of Hsp27 and Hsp70 with the clinicopathological parameters, including gender, age, surgical margin, lymph node status and tumor differentiation. The median follow-up period was 60 months. Survival analysis of patients with ESCC showed no relationship with the expression of Hsp27 and Hsp70. Conclusion. Taken together, our results demonstrate that Hsp27 and Hsp70 are expressed in ESCC tissues, but they are not good prognostic factor for patients with ESCC. Creative Commons Attribution License

  3. Energy sources and levels influenced on performance parameters, thyroid hormones, and HSP70 gene expression of broiler chickens under heat stress.

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    Raghebian, Majid; Sadeghi, Ali Asghar; Aminafshar, Mehdi

    2016-12-01

    The present study was conducted to evaluate the effects of energy sources and levels on body and organs weights, thyroid hormones, and heat shock protein (HSP70) gene expression in broilers under heat stress. In a completely randomized design, 600 1-day-old Cobb chickens were assigned to five dietary treatments and four replicates. The chickens were fed diet based on corn as main energy source and energy level based on Cobb standard considered as control (C), corn-based diet with 3 % lesser energy than the control (T1), corn-based diet with 6 % lesser energy than the control (T2), corn and soybean oil-based diet according to Cobb standard (T3), and corn and soybean oil-based diet with 3 % upper energy than the control (T4). Temperature was increased to 34 °C for 8 h daily from days 12 to 41 of age to induce heat stress. The chickens in T1 and T2 had lower thyroid hormones and corticosterone levels than those in C, T3, and T4. The highest liver weight was for C and the lowest one was for T4. The highest gene expression was found in chickens fed T4 diet, and the lowest gene expression was for those in T2 group. The highest feed intake and worse feed conversion ratio was related to chickens in T2. The chickens in T3 and T4 had higher feed intake and weight gain than those in C. The results showed that the higher energy level supplied from soybean oil could enhance gene expression of HSP70 and decline the level of corticosterone and thyroid hormones and consequently improved performance.

  4. Variation in Hsp70-1A Expression Contributes to Skin Color Diversity.

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    Murase, Daiki; Hachiya, Akira; Fullenkamp, Rachel; Beck, Anita; Moriwaki, Shigeru; Hase, Tadashi; Takema, Yoshinori; Manga, Prashiela

    2016-08-01

    The wide range in human skin color results from varying levels of the pigment melanin. Genetic mechanisms underlying coloration differences have been explored, but identified genes do not account for all variation seen in the skin color spectrum. Post-transcriptional and post-translational regulation of factors that determine skin color, including melanin synthesis in epidermal melanocytes, melanosome transfer to keratinocytes, and melanosome degradation, is also critical for pigmentation. We therefore investigated proteins that are differentially expressed in melanocytes derived from either white or African American skin. Two-dimensional gel electrophoresis and mass spectrometry demonstrated that heat shock protein 70-1A (Hsp70-1A) protein levels were significantly higher in African American melanocytes compared with white melanocytes. Hsp70-1A expression significantly correlated with levels of tyrosinase, the rate-limiting melanogenic enzyme, consistent with a proposed role for Hsp70 family members in tyrosinase post-translational modification. In addition, pharmacologic inhibition and small interfering RNA-mediated downregulation of Hsp70-1A correlated with pigmentation changes in cultured melanocytes, modified human skin substitutes, and ex vivo skin. Furthermore, Hsp70-1A inhibition led to increased autophagy-mediated melanosome degradation in keratinocytes. Our data thus reveal that epidermal Hsp70-1A contributes to the diversity of skin color by regulating the amount of melanin synthesized in melanocytes and modulating autophagic melanosome degradation in keratinocytes. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  5. Preimplantation development and expression of Hsp-70 and Bax genes in bovine blastocysts derived from oocytes matured in alpha-MEM supplemented with growth factors and synthetic macromolecules.

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    Vireque, A A; Camargo, L S A; Serapião, R V; Rosa E Silva, A A M; Watanabe, Y F; Ferreira, E M; Navarro, P A A S; Martins, W P; Ferriani, R A

    2009-03-01

    In vitro culture conditions affect both the maternal and embryonic expression of genes and is likely to alter both oocyte and embryo developmental competence. The search for better and less variable culture conditions simulating those in vivo has led to the development of defined culture media, with lower impact on the molecular reprogramming of oocytes and embryos. We evaluated embryo development and relative abundance (RA) of Hsp-70 and Bax transcripts in bovine blastocysts produced from oocytes matured in a chemically defined IVM system with synthetic polymers. Immature cumulus oocyte complexes (COCs) were matured for 22-24h in alpha-MEM supplemented with IGF-1, insulin, 0.1% polyvinyl alcohol (PVA), or 0.1% polyvinylpyrrolidone (PVP), but without FSH or LH. The control group consisted of COCs matured in TCM plus FSH and 10% estrous cow serum. After fertilization, presumptive zygotes were co-cultured with cumulus cells until 224 h post-insemination. Total RNA was isolated from embryo pools, reverse transcribed into cDNA, and subjected to transcript analysis by real-time PCR. Cleavage rate was higher (P<0.05) for the control group (68.3%) than for the PVA (54.4%) and PVP-40 (58.3%) groups. Nevertheless, there was no difference among the PVA, PVP-40 and control groups in blastocyst or hatching rates. Similarly, no difference in relative abundance of Hsp-70 and Bax transcripts was detected in comparison to the control group. We inferred that bovine oocytes can be matured in serum- and gonadotrophin-free medium supplemented with PVA or PVP, enriched with IGF-I and insulin, without altering post-cleavage development and relative abundance of some genes associated with stress and apoptosis.

  6. Conserved structure and expression of hsp70 paralogs in teleost fishes

    DEFF Research Database (Denmark)

    Metzger, David C.H.; Hansen, Jakob Hemmer; Schulte, Patricia M.

    2016-01-01

    The cytosolic 70 KDa heat shock proteins (Hsp70s) are widely used as biomarkers of environmental stress in ecological and toxicological studies in fish. Here we analyze teleost genome sequences to show that two genes encoding inducible hsp70s (hsp70-1 and hsp70-2) are likely present in all teleost...... fish. Phylogenetic and synteny analyses indicate that hsp70-1 and hsp70-2 are distinct paralogs that originated prior to the diversification of the teleosts. The promoters of both genes contain a TATA box and conserved heat shock elements (HSEs), but unlike mammalian HSP70s, both genes contain...... an intron in the 5′ UTR. The hsp70-2 gene has undergone tandem duplication in several species. In addition, many other teleost genome assemblies have multiple copies of hsp70-2 present on separate, small, genomic scaffolds. To verify that these represent poorly assembled tandem duplicates, we cloned...

  7. Distinct transcripts are recognized by sense and antisense riboprobes for a member of the murine HSP70 gene family, HSP70.2, in various reproductive tissues

    Science.gov (United States)

    Murashov, A. K.; Wolgemuth, D. J.

    1996-01-01

    The expression of hsp70.2, an hsp70 gene family member, originally characterized by its high levels of expression in germ cells in the adult mouse testis, was detected in several other reproductive tissues, including epididymis, prostate, and seminal vesicles, as well as in extraembryonic tissues of mid-gestation fetuses. In addition, hybridization with RNA probes transcribed in the sense orientation surprisingly indicated the presence of slightly larger "antisense" transcripts in several tissues. The levels of antisense transcripts varied among the tissues, with the highest signal detected in the prostate and no signal being detectable in the testis. Consistent with these results, in situ hybridization analysis clearly localized the sense-orientation transcripts to pachytene spermatocytes, while no antisense-orientation transcripts were observed in adjacent sections of the same tubules. Our findings have thus shown that although hsp70.2 was expressed abundantly and in a highly stage-specific manner in the male germ line, it was also expressed in other murine tissues. Furthermore, we have made the surprising observation of antisense transcription of the hsp70.2 gene in several mouse tissues, revealing another level of complexity in the regulation and function of heat shock proteins.

  8. Genome-wide analysis of the Hsp70 family genes in pepper (Capsicum annuum L.) and functional identification of CaHsp70-2 involvement in heat stress.

    Science.gov (United States)

    Guo, Meng; Liu, Jin-Hong; Ma, Xiao; Zhai, Yu-Fei; Gong, Zhen-Hui; Lu, Ming-Hui

    2016-11-01

    Hsp70s function as molecular chaperones and are encoded by a multi-gene family whose members play a crucial role in plant response to stress conditions, and in plant growth and development. Pepper (Capsicum annuum L.) is an important vegetable crop whose genome has been sequenced. Nonetheless, no overall analysis of the Hsp70 gene family is reported in this crop plant to date. To assess the functionality of Capsicum annuum Hsp70 (CaHsp70) genes, pepper genome database was analyzed in this research. A total of 21 CaHsp70 genes were identified and their characteristics were also described. The promoter and transcript expression analysis revealed that CaHsp70s were involved in pepper growth and development, and heat stress response. Ectopic expression of a cytosolic gene, CaHsp70-2, regulated expression of stress-related genes and conferred increased thermotolerance in transgenic Arabidopsis. Taken together, our results provide the basis for further studied to dissect CaHsp70s' function in response to heat stress as well as other environmental stresses. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  9. Molecular cloning and functional analysis of the drought tolerance gene MsHSP70 from alfalfa (Medicago sativa L.).

    Science.gov (United States)

    Li, Zhenyi; Long, Ruicai; Zhang, Tiejun; Wang, Zhen; Zhang, Fan; Yang, Qingchuan; Kang, Junmei; Sun, Yan

    2017-03-01

    Heat shock proteins (HSPs) are a ubiquitously expressed class of protective proteins that play a key role in plant response to stressful conditions. This study aimed to characterize and investigate the function of an HSP gene in alfalfa (Medicago sativa). MsHSP70, which contains a 2028-bp open reading frame, was identified through homology cloning. MsHSP70 shares high sequence identity (94.47%) with HSP70 from Medicago truncatula. Expression analysis of MsHSP70 in alfalfa organs revealed a relatively higher expression level in aerial organs such as flowers, stems and leaves than in roots. MsHSP70 was induced by heat shock, abscisic acid (ABA) and hydrogen peroxide. Transgenic Arabidopsis seedlings overexpressing MsHSP70 were hyposensitive to polyethylene glycol (PEG) and ABA treatments, suggesting that exogenous expression of MsHSP70 enhanced Arabidopsis tolerance to these stresses. Examination of physiological indexes related to drought and ABA stress demonstrated that in comparison with non-transgenic plants, T3 transgenic Arabidopsis plants had an increased proline content, higher superoxide dismutase (SOD) activity, and decreased malondialdehyde (MDA) content. Furthermore, higher relative water content (RWC) was detected in transgenic plants compared with non-transgenic plants under drought stress. These findings clearly indicate that molecular manipulation of MsHSP70 in plants can have substantial effects on stress tolerance.

  10. Expression patterns and structural modelling of Hsp70 and Hsp90 in a fish-borne zoonotic nematode Anisakis pegreffii.

    Science.gov (United States)

    Chen, Hui-Yu; Cheng, Yi-Sheng; Shih, Hsiu-Hui

    2015-09-15

    Heat shock proteins (HSPs) are essential molecular chaperones that are highly conserved across organisms. They have a pivotal function in responding to thermal stress and are responsible for many cellular functions. Here, we aimed to elucidate the possible roles of Hsp70 and Hsp90 in the life cycle of the parasitic nematode Anisakis, particularly third- and fourth-stage larvae, from cold-blooded fish to warm-blooded marine mammals or accidentally to human hosts. We examined the expression profiles of Hsp70 and Hsp90 in different developmental stages of Anisakis pegreffii. The open reading frame of Hsp70 of A. pegreffii was 1950 bp, and deduced amino acid sequence showed high homology with those of other nematodes. Heatmap analysis revealed sequence identity of Hsp70 and Hsp90 in 13 important parasitic species, human and yeast. On heatmap and phylogenetic analysis, ApHsp70 and ApHsp90 shared the highest amino acid sequence identity with other nematodes and formed a monophyletic clade. The three-dimensional (3D) structure prediction of the newly characterized ApHsp70 and known ApHsp90 gene showed highly conserved motifs between A. pegreffii and other species. Quantitative real-time PCR and western blot analysis revealed higher mRNA and protein expression for ApHsp70 and ApHsp90 in fourth- than third-stage larvae, with higher mRNA and protein expression for ApHsp70 than ApHsp90. ApHsp70 and ApHsp90 may play important roles in Anisakis in response to thermal stress and might be important molecules in the development of A. pegreffii, which has implications for its control. Copyright © 2015 Elsevier B.V. All rights reserved.

  11. Molecular characterization of a ribosome-associated Hsp70-homologous gene from Rhizopus nigricans.

    Science.gov (United States)

    Cernila, Bostjan; Cresnar, Bronislava; Breskvar, Katja

    2003-10-01

    A ribosome-associated Hsp70-homologous gene (Rnssb-1) was isolated from the genomic library of the filamentous zygomycete fungus Rhizopus nigricans. The nucleotide sequence of a genomic clone encoded the N-terminal part of a protein with high similarity to the yeast SSB ribosome-associated chaperones. The missing 3' end of the gene was obtained by 3' RACE. The Northern blot analysis showed that the Rnssb-1 gene is constitutively expressed and is not induced upon heat shock at 37 degrees C. The primary structure analyses revealed that the coding region of the Rnssb-1 gene is interrupted by at least four introns. Their splicing was not inhibited by exposure of the organism to heat shock as proven by RT-PCR. A Southern blot analysis of R. nigricans genomic DNA confirmed the presence of two additional gene copies of ribosome-associated Hsp70 genes in the fungal genome.

  12. Differential GFP expression patterns induced by different heavy metals in Tg(hsp70:gfp) transgenic medaka (Oryzias latipes).

    Science.gov (United States)

    Ng, Grace Hwee Boon; Xu, Hongyan; Pi, Na; Kelly, Barry C; Gong, Zhiyuan

    2015-06-01

    Heat shock protein 70 (Hsp70) is one of the most widely used biomarker for monitoring environment perturbations in biological systems. To facilitate the analysis of hsp70 expression as a biomarker, we generated a Tg(hsp70:gfp) transgenic medaka line in which green fluorescence protein (GFP) reporter gene was driven by the medaka hsp70 promoter. Here, we characterized Tg(hsp70:gfp) medaka for inducible GFP expression by seven environment-relevant heavy metals, including mercury, arsenic, lead, cadmium, copper, chromium, and zinc. We found that four of them (mercury, arsenic, lead, and cadmium) induced GFP expression in multiple and different organs. In general, the liver, kidney, gut, and skin are among the most frequent organs to show induced GFP expression. In contrast, no detectable GFP induction was observed to copper, chromium, or zinc, indicating that the transgenic line was not responsive to all heavy metals. RT-qPCR determination of hsp70 mRNA showed similar induction and non-induction by these metals, which also correlated with the levels of metal uptake in medaka exposed to these metals. Our observations suggested that these heavy metals have different mechanisms of toxicity and/or differential bioaccumulation in various organs; different patterns of GFP expression induced by different metals may be used to determine or exclude metals in water samples tested. Furthermore, we also tested several non-metal toxicants such as bisphenol A, 2,3,7,8-tetrachlorodibenzo-p-dioxin, 4-introphenol, and lindane; none of them induced significant GFP expression in Tg(hsp70:gfp) medaka, further suggesting that the inducibility of Tg(hsp70:gfp) for GFP expression is specific to a subset of heavy metals.

  13. Cell-surface expression of Hsp70 on hematopoietic cancer cells after inhibition of HDAC activity

    DEFF Research Database (Denmark)

    Jensen, Helle; Andresen, Lars; Hansen, Karen Aagaard

    or normal ER/Golgi transport did not affect Hsp70 surface expression. Intracellular Calcium and the transcription factor Sp1, that has previously been shown to be important for the intracellular stress mediated by HDAC-inhibitors, were not involved in Hsp70 surface expression. We also found that HDAC...

  14. Adult Heat Tolerance Variation in Drosophila melanogaster is Not Related to Hsp70 Expression

    DEFF Research Database (Denmark)

    Jensen, Louise Toft; Cockerell, Fiona Elizabeth; Kristensen, Torsten Nygaard

    2010-01-01

    Expression of heat-inducible Hsp70 is considered closely linked to thermotolerance in Drosophila melanogaster and other ectotherms. However, intra-specific variation of Hsp70 expression levels and its relationship to heat resistance has only been investigated in a few studies. Although...... in adult heat tolerance. Therefore, although Hsp70 expression is a major component of the cellular heat stress response, its influence on intra-specific heat tolerance variation may be life-stage specific. J. Exp. Zool. 313A:35-44, 2010. © 2009 Wiley-Liss, Inc...... in Drosophila larvae Hsp70 expression may be a key determinant of heat tolerance, the evidence for this in adults is equivocal. We therefore examined heat-induced Hsp70 expression and several measurements of adult heat tolerance in three independent collections of D. melanogaster, measured in three laboratories...

  15. Expression of HSP70 in cerebral ischemia and neuroprotetive action of hypothermia and ketoprofen Expressão de HSP70 na isquemia cerebral e a ação neuroprotetora da hipotermia e do cetoprofeno

    Directory of Open Access Journals (Sweden)

    Daniela Pretti da Cunha Tirapelli

    2010-08-01

    Full Text Available Heat shock proteins (HSPs are molecular chaperones that bind to other proteins to shepherd them across membranes and direct them to specific locations within a cell. Several injurious stimuli can induce Hsp70 expression, including ischemia. This study aimed to investigate the pattern of expression of protein (immunohistochemistry and gene (real-time PCR Hsp70 in experimental focal cerebral ischemia in rats by occlusion of the middle cerebral artery for 1 hour and the role of neuroprotection with hypothermia (H and ketoprofen (K. The infarct volume was measured using morphometric analysis defined by triphenyl tetrazolium chloride. It was observed increases in the protein (p=0.0001 and gene (p=0.0001 Hsp70 receptor in the ischemic areas that were reduced by H (protein and gene: pProteínas de choque térmico (HSPs são chaperones moleculares que se ligam a outras proteínas para atravessar as membranas e encaminhá-las para locais específicos dentro de uma célula. Vários estímulos nocivos podem induzir a expressão de Hsp70, incluindo isquemia. Este estudo teve como objetivo investigar o padrão de expressão protéica (imunohistoquímica e gênica (PCR em tempo real de Hsp70 na isquemia cerebral focal experimental em ratos pela oclusão da artéria cerebral média durante 1 hora e o papel da neuroproteção com hipotermia (H e cetoprofeno (C. O volume de infarto foi calculado através da análise morfométrica definido por cloreto de trifenil tetrazólio. Foi observado aumento na expressão proteína (p=0,0001 e gênica (p=0,0001 de Hsp70 nas áreas isquêmicas que foram reduzidas pela H (proteína e gene: p<0,05, C (proteína: p<0,001 e H+K (proteína: p<0,01 e gene: p<0,05. O aumento de Hsp70 na área isquêmica sugere que a neuroexcitotoxicidade mediada pela Hsp70 desempenha um papel importante na morte celular e que o efeito neuroprotetor tanto da H quanto do C está diretamente envolvido com a Hsp70.

  16. Polymorphism analysis of the hsp70 stress gene in Broiler chickens (Gallus gallus of different breeds

    Directory of Open Access Journals (Sweden)

    Mazzi Carmen Maria

    2003-01-01

    Full Text Available The promoter region and the beginning of the coding region of the hsp70 stress gene were analysed in broiler chickens of a commercial breed (Hubbard-Pettersen, a breed selected for weight gain (PP1 and a non-selected breed (naked-neck Label Rouge. The naked neck gene (Naked neck, Na, which reduces feathering in birds and is thus related to heat resistance, was present in both PP1 and Label Rouge breeds. Genomic DNA was restricted with PstI and Southern blotting analysis of the samples revealed the absence of polymorphic sites for that enzyme in the promoter region and beginning of the coding region of the hsp70 gene of studied birds. PCR-SSCP analysis of these regions, however, indicated the presence of polymorphisms in the beginning of the coding region and the sequencing of the PCR products confirmed and identified two polymorphic sites in this region: a transition A ® G in position +258 and a transversion C ® G in position +276. Both mutations were considered to be silent, since they did not modify the aminoacid sequence of the protein Hsp70. The promoter region of the hsp70 gene was identical in all studied birds, indicating that the regulation pattern of this gene must be the same in all birds at the promoter region. Three different alleles (hsp70-1, hsp70-2 and hsp70-3 were identified for the hsp70 gene from the observed mutations. The allele hsp70-3 was detected in only two breeds, Hubbard-Pettersen and PP1, but at a low frequency (0,016 and 0,006, respectively.

  17. Hsp70 and Hsp90 are differentially expressed in crayfish muscle and neurons after heat stress

    Directory of Open Access Journals (Sweden)

    Liang S

    2013-12-01

    Full Text Available Shuang Liang, Xiaoqing Yu, Debra E Wood, Emmitt R Jolly Department of Biology, Case Western Reserve University, Cleveland, OH, USA Abstract: Heat shock proteins are essential cellular proteins that are highly conserved across organisms and contribute to adaptive responses of organisms during changing environmental conditions. Protein members of the families of heat shock genes can be differentially regulated in response to stressors and play critical roles in protein stability, folding, and molecular trafficking. We used a crustacean species with strong adaptability to diverse environments, the crayfish Procambarus clarkii, to study expression profiles of two well known heat shock genes, Hsp90 and Hsp70. This crayfish can withstand a broad range of temperatures, and its adaptability contributes to its value for human use as an agricultural food source and as a biological control agent against snails that transmit schistosomiasis. However, it has become a harmful invasive species in some areas. To begin to understand the thermal resilience of P. clarkii, we identified and cloned Hsp90 from crayfish by degenerate polymerase chain reaction in conjunction with rapid amplification of 3' and 5' cDNA ends, and subsequently sequenced and characterized the molecular chaperone. Sequence analysis by phylogenetic alignment and polypeptide three-dimensional structure prediction of the newly identified Hsp90 gene shows that it has conserved motifs with Hsp90 s in other species. Using quantitative polymerase chain reaction, we characterized the expression profiles of Hsp90 and Hsp70 in muscle and in central nervous system tissues. We found that Hsp70 and Hsp90 transcripts are upregulated under heat stress in both muscle and the central nervous system, but that their expression levels are more robustly increased in muscle. Keywords: crayfish, stress response, Procambarus clarkii, heat shock protein, Hsp90, schistosomiasis

  18. Molecular characterization of genes encoding cytosolic Hsp70s in the zygomycete fungus Rhizopus nigricans

    OpenAIRE

    Černila, Boštjan; Črešnar, Bronislava; Breskvar, Katja

    2003-01-01

    Previous studies have shown that some stressors, including steroid hormones 21-OH progesterone and testosterone, stimulate the accumulation of heat shock protein 70 (hsp70) messenger ribonucleic acid (mRNA) population in the zygomycete filamentous fungus Rhizopus nigricans. In this study we report the cloning of 3 R nigricans hsp70 genes (Rnhsp70-1, Rnhsp70-2, and Rnhsp70-3) encoding cytosolic Hsp70s. With a Southern blot experiment under high stringency conditions we did not detect any addit...

  19. Proteomic Profiling Reveals Upregulated Protein Expression of Hsp70 in Keloids

    Directory of Open Access Journals (Sweden)

    Ju Hee Lee

    2013-01-01

    Full Text Available Background. The biochemical characteristics of keloid-derived fibroblasts differ from those of adjacent normal fibroblasts, and these differences are thought to be the cause of abnormal fibrosis. Therefore, we investigated the characteristic proteins that are differentially expressed in keloid-derived fibroblasts using proteomics tools. Objective. We attempted to investigate the novel proteins that play important roles in the pathophysiology of keloids. Methods. Proteomics analysis was performed to identify differentially expressed proteins in keloid-derived fibroblasts. Keloid-derived fibroblasts and adjacent normal fibroblasts were analyzed with 2-DAGE. We validated these proteins with immunoblot analysis, real-time RT-PCR, and immunohistochemistry. Results. Sixteen differentially expressed protein spots were identified in keloid-derived fibroblasts. Among them, heat shock protein 70 (Hsp70 was specifically upregulated in keloid-derived fibroblasts. Also, immunohistochemistry and western blot analysis revealed increased Hsp70, TGF-β, and PCNA expressions in keloids compared to normal tissue. Conclusion. Hsp70 is overexpressed in keloid fibroblasts and tissue. The overexpression of Hsp70 may be involved in the pathogenesis of keloids, and the inhibition of Hsp70 could be a new therapeutic tool for the treatment of keloids.

  20. DNA double strand breaks and Hsp70 expression in proton irradiated living cells

    Energy Technology Data Exchange (ETDEWEB)

    Fiedler, Anja [Institute for Experimental Physics II, University of Leipzig (Germany) and Faculty of Biology, Pharmacy and Psychology, University of Leipzig (Germany)]. E-mail: afiedler@uni-leipzig.de; Reinert, Tilo [Institute for Experimental Physics II, University of Leipzig (Germany); Tanner, Judith [Clinic and Polyclinic for Radiation Oncology, University of Halle-Wittenberg (Germany); Butz, Tilman [Institute for Experimental Physics II, University of Leipzig (Germany)

    2007-07-15

    DNA double strand breaks (DSBs) in living cells can be directly provoked by ionising radiation. DSBs can be visualized by immunostaining the phosphorylated histone {gamma}H2AX. Our concern was to test the feasibility of {gamma}H2AX staining for a direct visualization of single proton hits. If single protons produce detectable foci, DNA DSBs could be used as 'biological track detectors' for protons. Ionising radiation can also damage proteins indirectly by inducing free radicals. Heat shock proteins (Hsp) help to refold or even degrade the damaged proteins. The level of the most famous heat shock protein Hsp70 is increased by ionising radiation. We investigated the expression of {gamma}H2AX and Hsp70 after cross and line patterned irradiation with counted numbers of 2.25 MeV protons on primary human skin fibroblasts. The proton induced DSBs appear more delocalised than it was expected by the ion hit accuracy. Cooling the cells before the irradiation reduces the delocalisation of DNA DSBs, which is probably caused by the reduced diffusion of DNA damaging agents. Proton irradiation seems to provoke protein damages mainly in the cytoplasm indicated by cytoplasmic Hsp70 aggregates. On the contrary, in control heat shocked cells the Hsp70 was predominantly localized in the cell nucleus. However, the irradiated area could not be recognized, all cells on the Si{sub 3}N{sub 4} window showed a homogenous Hsp70 expression pattern.

  1. HSP70 expression in the copper butterfly Lycaena tityrus across altitudes and temperatures

    DEFF Research Database (Denmark)

    Karl, I.; Sørensen, Jesper Givskov; Loeschcke, Volker

    2009-01-01

    The ability to express heat-shock proteins (HSP) under thermal stress is an essential mechanism for ectotherms to cope with unfavourable conditions. In this study, we investigate if Copper butterflies originating from different altitudes and/or being exposed to different rearing and induction...... temperatures show differences in HSP70 expression. HSP70 expression increased substantially at the higher rearing temperature in low-altitude butterflies, which might represent an adaptation to occasionally occurring heat spells. On the other hand, high-altitude butterflies showed much less plasticity...

  2. Stress protein (HSP70 family expression in intertidal benthic organisms: the example of Anthopleura elegantissima (Cnidaria: Anthozoa

    Directory of Open Access Journals (Sweden)

    Mark J. Snyder

    2004-04-01

    Full Text Available Both the physiology and distribution of intertidal organisms are strongly influenced by different kinds of physical stress. Temperature, UV radiation and desiccation in low tide conditions are usually considered the most important physical stresses. These factors impact cell metabolism, and the organism´s ability to rapidly adapt to altering environmental conditions sets its tidal distribution limits. The role of the HSP70 protein (stress protein family in thermal stress responses has been widely demonstrated. Although it has been shown that stress protein expression is a useful tool to quantify part of this physical stress, few studies have been made with different in situ intertidal organisms. To test differences in HSP70 expression under natural conditions in the intertidal environment, we chose a common cnidarian of the eastern Pacific Ocean: the anemone Anthopleura elegantissima. Polyp HSP70 expression depends on the degree of emersion and the extent of physical stress (0.1-3.6 ± 1.5 ng HSP70 mg Protein-1 were registered between fully immersed and fully emersed polyps of the same clone. The time of immersion is reflected in the recovery of polyp HSP70 levels from the high tidal exposure (reaching again as shallower clones express more HSP70 (2.5 ng HSP70 mg P-1, than deeper ones (0.1 ng HSP70 mg P-1. Also sunny and foggy environmental situations influence the stress response. Anemone clones exposed to the sunny high intertidal zone express more than three times HSP70 (2.2 ng HSP70 mg P-1 than those on a foggy high intertidal day (0.6 ng HSP70 mg P-1. For A. elegantissima, shrinking of polyps, mucus secretion, sand covering, UV absorbing molecules, and a tight patch structure work concurrently with HSP70 expression to alleviate the effects of physical stress in low tidal emersion.

  3. Cell-surface expression of Hsp70 on hematopoietic cancer cells after inhibition of HDAC activity

    DEFF Research Database (Denmark)

    Jensen, Helle; Andresen, Lars; Hansen, Karen Aagaard

    2009-01-01

    surface-negative despite effective induction of apoptosis. Interestingly, inhibition of endolysosomes or normal ER/Golgi transport did not affect Hsp70 surface expression. Intracellular calcium and the transcription factor Sp1, which has been shown previously to be important for the intracellular stress...

  4. Heritability of hsp70 expression in the beetle Tenebrio molitor: Ontogenetic and environmental effects.

    Science.gov (United States)

    Lardies, Marco A; Arias, María Belén; Poupin, María Josefina; Bacigalupe, Leonardo D

    2014-08-01

    Ectotherms constitute the vast majority of terrestrial biodiversity and are especially likely to be vulnerable to climate warming because their basic physiological functions such as locomotion, growth, and reproduction are strongly influenced by environmental temperature. An integrated view about the effects of global warming will be reached not just establishing how the increase in mean temperature impacts the natural populations but also establishing the effects of the increase in temperature variance. One of the molecular responses that are activated in a cell under a temperature stress is the heat shock protein response (HSP). Some studies that have detected consistent differences among thermal treatments and ontogenetic stages in HSP70 expression have assumed that these differences had a genetic basis and consequently expression would be heritable. We tested for changes in quantitative genetic parameters of HSP70 expression in a half-sib design where individuals of the beetle Tenebrio molitor were maintained in constant and varying thermal environments. We estimated heritability of HSP70 expression using a linear mixed modelling approach in different ontogenetic stages. Expression levels of HSP70 were consistently higher in the variable environment and heritability estimates were low to moderate. The results imply that within each ontogenetic stage additive genetic variance was higher in the variable environment and in adults compared with constant environment and larvae stage, respectively. We found that almost all the genetic correlations across ontogenetic stages and environment were positive. These suggest that directional selection for higher levels of expression in one environment will result in higher expression levels of HSP70 on the other environment for the same ontogenetic stage. Copyright © 2014 Elsevier Ltd. All rights reserved.

  5. Molecular characterization of genes encoding cytosolic Hsp70s in the zygomycete fungus Rhizopus nigricans.

    Science.gov (United States)

    Cernila, Bostjan; Cresnar, Bronislava; Breskvar, Katja

    2003-01-01

    Previous studies have shown that some stressors, including steroid hormones 21-OH progesterone and testosterone, stimulate the accumulation of heat shock protein 70 (hsp70) messenger ribonucleic acid (mRNA) population in the zygomycete filamentous fungus Rhizopus nigricans. In this study we report the cloning of 3 R nigricans hsp70 genes (Rnhsp70-1, Rnhsp70-2, and Rnhsp70-3) encoding cytosolic Hsp70s. With a Southern blot experiment under high stringency conditions we did not detect any additional highly homologous copies of the cytosolic hsp70 genes in the R nigricans genome. Sequence analyses showed that all 3 genes contain introns within the open reading frame. The dynamics of the R nigricans molecular response to progesterone, 21-OH progesterone, and testosterone, as well as to heat shock, copper ions, hydrogen peroxide, and ethanol was studied by temporal analysis of Rnhsp70-1 and Rnhsp70-2 mRNA accumulation. Northern blot experiments revealed that the Rnhsp70-2 transcript level is not affected by testosterone, whereas mRNA levels of both genes are rapidly increased with all the other stressors studied. Moreover, the decrease of transcript levels is notably delayed in ethanol stress, and a difference is observed between the profiles of Rnhsp70-1 and Rnhsp70-2 transcripts during heat stress.

  6. Molecular characterization of genes encoding cytosolic Hsp70s in the zygomycete fungus Rhizopus nigricans

    Science.gov (United States)

    Černila, Boštjan; Črešnar, Bronislava; Breskvar, Katja

    2003-01-01

    Previous studies have shown that some stressors, including steroid hormones 21-OH progesterone and testosterone, stimulate the accumulation of heat shock protein 70 (hsp70) messenger ribonucleic acid (mRNA) population in the zygomycete filamentous fungus Rhizopus nigricans. In this study we report the cloning of 3 R nigricans hsp70 genes (Rnhsp70-1, Rnhsp70-2, and Rnhsp70-3) encoding cytosolic Hsp70s. With a Southern blot experiment under high stringency conditions we did not detect any additional highly homologous copies of the cytosolic hsp70 genes in the R nigricans genome. Sequence analyses showed that all 3 genes contain introns within the open reading frame. The dynamics of the R nigricans molecular response to progesterone, 21-OH progesterone, and testosterone, as well as to heat shock, copper ions, hydrogen peroxide, and ethanol was studied by temporal analysis of Rnhsp70-1 and Rnhsp70-2 mRNA accumulation. Northern blot experiments revealed that the Rnhsp70-2 transcript level is not affected by testosterone, whereas mRNA levels of both genes are rapidly increased with all the other stressors studied. Moreover, the decrease of transcript levels is notably delayed in ethanol stress, and a difference is observed between the profiles of Rnhsp70-1 and Rnhsp70-2 transcripts during heat stress. PMID:15115284

  7. Hsp70 expression and induction as a readout for detection of immune modulatory components in food.

    Science.gov (United States)

    Wieten, Lotte; van der Zee, Ruurd; Goedemans, Renske; Sijtsma, Jeroen; Serafini, Mauro; Lubsen, Nicolette H; van Eden, Willem; Broere, Femke

    2010-01-01

    Stress proteins such as heat shock proteins (Hsps) are up-regulated in cells in response to various forms of stress, like thermal and oxidative stress and inflammation. Hsps prevent cellular damage and increase immunoregulation by the activation of anti-inflammatory T-cells. Decreased capacity for stress-induced Hsp expression is associated with immune disorders. Thus, therapeutic boosting Hsp expression might restore or enhance cellular stress resistance and immunoregulation. Especially food- or herb-derived phytonutrients may be attractive compounds to restore optimal Hsp expression in response to stress. In the present study, we explored three readout systems to monitor Hsp70 expression in a manner relevant for the immune system and evaluated novel Hsp co-inducers. First, intracellular staining and analysis by flow cytometry was used to detect stress and/or dietary compound induced Hsp70 expression in multiple rodent cell types efficiently. This system was used to screen a panel of food-derived extracts with potent anti-oxidant capacity. This strategy yielded the identity of several new enhancers of stress-induced Hsp70 expression, among them carvacrol, found in thyme and oregano. Second, CD4(+) T-cell hybridomas were generated that specifically recognized an immunodominant Hsp70 peptide. These hybridomas were used to show that carvacrol enhanced Hsp70 levels increased T-cell activation. Third, we generated a DNAJB1-luc-O23 reporter cell line to show that carvacrol increased the transcriptional activation of a heat shock promoter in the presence of arsenite. These assay systems are generally applicable to identify compounds that affect the Hsp level in cells of the immune system.

  8. Bleaching and stress in coral reef ecosystems: hsp70 expression by the giant barrel sponge Xestospongia muta.

    Science.gov (United States)

    López-Legentil, Susanna; Song, Bongkeun; McMurray, Steven E; Pawlik, Joseph R

    2008-04-01

    Sponges are a prominent component of coral reef ecosystems. Like reef-building corals, some sponges have been reported to bleach and die. The giant barrel sponge Xestospongia muta is one of the largest and most important components of Caribbean coral reef communities. Tissues of X. muta contain cyanobacterial symbionts of the Synechococcus group. Two types of bleaching have been described: cyclic bleaching, from which sponges recover, and fatal bleaching, which usually results in sponge death. We quantified hsp70 gene expression as an indicator of stress in X. muta undergoing cyclic and fatal bleaching and in response to thermal and salinity variability in both field and laboratory settings. Chlorophyll a content of sponge tissue was estimated to determine whether hsp70 expression was related to cyanobacterial abundance. We found that fatally bleached sponge tissue presented significantly higher hsp70 gene expression, but cyclically bleached tissue did not, yet both cyclic and fatally bleached tissues had lower chlorophyll a concentrations than nonbleached tissue. These results corroborate field observations suggesting that cyclic bleaching is a temporary, nonstressful state, while fatal bleaching causes significant levels of stress, leading to mortality. Our results support the hypothesis that Synechococcus symbionts are commensals that provide no clear advantage to their sponge host. In laboratory experiments, sponge pieces incubated at 30 degrees C exhibited significantly higher hsp70 expression than control pieces after 1.5 h, with sponge mortality after less than 15 h. In contrast, sponges at different salinities were not significantly stressed after the same period of time. Stress associated with increasing seawater temperatures may result in declining sponge populations in coral reef ecosystems.

  9. Reduced Hsp70 and Glutamine in Pediatric Severe Malaria Anemia

    DEFF Research Database (Denmark)

    Kempaiah, Prakasha; Dokladny, Karol; Karim, Zachary

    2016-01-01

    Severe malarial anemia [SMA, hemoglobin (Hb) cause of global morbidity and mortality among children residing in Plasmodium falciparum transmission regions. Exploration of molecular pathways through global gene expression profiling revealed that SMA was characterized...... by decreased HSPA1A, a heat shock protein (Hsp) 70 coding gene. Hsp70 is a ubiquitous chaperone that regulates Nuclear Factor-kappa B (NF-κB) signaling and production of pro-inflammatory cytokines known to be important in malaria pathogenesis (e.g., IL-1β, IL-6 and TNF-α). Since the role of host Hsp70...... in malaria pathogenesis is unexplored, we investigated Hsp70 and molecular pathways in children with SMA. Validation experiments revealed that leukocytic HSP70 transcripts were reduced in SMA relative to non-severe malaria, and that intraleukocytic hemozoin (PfHz) was associated with lower HSP70. HSP70...

  10. Hsp70 Expression Profile in Preeclampsia Model of Pregnant Rat (Rattus norvegicus) after Giving the EVOO

    Science.gov (United States)

    Irianti, E.; ilyas, S.; Rosidah; Hutahaean, S.

    2017-03-01

    Heat shock protein (Hsp) has long been known to protect cells from oxidative stress. In this case an increased expression is found on several cases of preeclampsia. One of the efforts to prevent preeclampsia is by giving antioxidants such as Extra Virgin Olive Oil (EVOO) or it’s better known as olive oil (Oleoa europaea), in the form of extra virgin known for its rich antioxidant content of tocopherols (vitamin E). The purpose of this study is to determine the expression levels of Hsp70 serum on pregnant white rat model of preeclampsia after being given EVOO. This type of research is true experiment; the subjects were female white rats and male virgin with Sprague Dawley, ± 8-11 weeks old, 180g BB s / d 200g, healthy and didn’t show any physical defects. Samples were 25 animals, divided into 5 groups, which consisted of different control and treatment given to T2 (rat model of preeclampsia), T3 (rat model of preeclampsia + EVOO 0.45g/bw/day), T4 (rat model of preeclampsia + EVOO 0.9g/bw/day) and T5 (rat model of preeclampsia + EVOO 1.8g/bw/day). The determination of each group was done by simple random sampling. Result on serum levels of Hsp70 that were tested by Elisa test in rats showed the average control was 14.64 mg / ml, group T2: 22:51 mg/ml, T3: 13.62 mg/ml, T4: 15.92 mg/ml, T5: 16:09 mg/ml. ANOVA test showed the P value was 0.001 <0.005, which meant there were significant differences on serum Hsp70 levels in the control and treatment pregnant rats group. It was known that there was a significant difference level of Hsp70 serum in group of control rats with T2 (P value <0.001) after LSD test was conducted, but not so with the group T3, T4, and T5, where the difference was not significant. There was a significant difference in the levels of Hsp70 serum on group T2 and T3 (P value 0.000), T4 (0004), T5 (0000). The gift of EVOO in the treatment group which was given EVOO with even low doses was able to control the induction of Hsp70 serum levels, which

  11. Sudden infant death syndrome: no significant expression of heat-shock proteins (HSP27, HSP70).

    Science.gov (United States)

    Doberentz, Elke; Führing, Sarah; Madea, Burkhard

    2016-03-01

    In industrialized countries, sudden infant death is the most common cause of death in young children. Although prone sleeping position is a well-known risk factor, hyperthermia might also be important. Pathognomonic findings of premortem hyperthermia do not exist. During stress, including thermal effects, heat-shock protein (HSP) expression increases. This study investigated hyperthermia as a contributing or pathogenic factor for sudden infant death syndrome (SIDS). Immunohistochemical staining for HSP27 and HSP70 in the kidney, heart, and lung from 120 SIDS cases was examined. HSP70 immunostaining was negative in kidney, heart, and lung tissues in all cases and in tissues from the control group. HSP27 staining was positive in the kidney from one case, and was positive in the lungs (respiratory epithelia in 27% of cases; vascular endothelia in 19% of cases) and was negative in the heart. In the control group HSP27 was positive in 8% of renal tubular tissues and in 29% of renal vascular endothelia. Staining for HSP27 in lung tissues was positive in respiratory epithelia in 8% of cases and for vascular endothelia in 29%, whereas tissues from the heart were positive in only 4%. The hypothesis of hyperthermia being a pathogenic factor for SIDS was not supported by immunohistochemical visualization of HSP70 or HSP27.

  12. Recombinant TgHSP70 Immunization Protects against Toxoplasma gondii Brain Cyst Formation by Enhancing Inducible Nitric Oxide Expression

    Directory of Open Access Journals (Sweden)

    Neide M. Silva

    2017-04-01

    Full Text Available Toxoplasma gondii is known to cause congenital infection in humans and animals and severe disease in immunocompromised individuals; consequently development of vaccines against the parasite is highly necessary. Under stress conditions, T. gondii expresses the highly immunogenic heat shock protein 70 (TgHSP70. Here, we assessed the protective efficacy of rTgHSP70 immunization combined with Alum in oral ME-49 T. gondii infection and the mechanisms involved on it. It was observed that immunized mice with rTgHSP70 or rTgHSP70 adsorbed in Alum presented a significantly reduced number of cysts in the brain that was associated with increased iNOS+ cell numbers in the organ, irrespective the use of the adjuvant. Indeed, ex vivo experiments showed that peritoneal macrophages pre-stimulated with rTgHSP70 presented increased NO production and enhanced parasite killing, and the protein was able to directly stimulate B cells toward antibody producing profile. In addition, rTgHSP70 immunization leads to high specific antibody titters systemically and a mixed IgG1/IgG2a response, with predominance of IgG1 production. Nonetheless, it was observed that the pretreatment of the parasite with rTgHSP70 immune sera was not able to control T. gondii internalization and replication by NIH fibroblast neither peritoneal murine macrophages, nor anti-rTgHSP70 antibodies were able to kill T. gondii by complement-mediated lysis, suggesting that these mechanisms are not crucial to resistance. Interestingly, when in combination with Alum, rTgHSP70 immunization was able to reduce inflammation in the brain of infected mice and in parallel anti-rTgHSP70 immune complexes in the serum. In conclusion, immunization with rTgHSP70 induces massive amounts of iNOS expression and reduced brain parasitism, suggesting that iNOS expression and consequently NO production in the brain is a protective mechanism induced by TgHSP70 immunization, therefore rTgHSP70 can be a good candidate for

  13. Examination the expression pattern of HSP70 heat shock protein in chicken PGCs and developing genital ridge

    Directory of Open Access Journals (Sweden)

    Mahek Anand

    2016-05-01

    Full Text Available Chicken Primordial Germ cells (PGCs are emerging pioneers in the field of applied embryology and stem cell technology. Now-a-days transgenic chickens are promising models to study human disease pathophysiology and drug designing. However, most of the molecular mechanism, which govern the stemness and pluripotency of chicken PGCs, not known in details. Recent studies have indicated the role of HSP70 in early embryonic development in many vertebrate species. Exposure of chicken to heat stress result in activation of heat shock factors which activate the transcription of HSP70. Exposure chicken eggs to acute heat stress effects HSP70 expression in PGCs and gonads. HSP70 helps in maintaining the integrity of chicken PGCs. A new emerging role of HSP70 in apoptosis has emerged. In our lab, we aim to characterize the expression of cHsp70 in chicken PGCs and gonads during embryonic development by subjecting the parents to acute levels of heat stress. Chickens whose parents subjected to heat stress showed varied expression of cHsp70 and also improved thermo tolerance. In the future we plan to study other factors and miRNAs, which is characterized as an emerging player in regulating heat shock protein response in chicken and also plays an important role in apoptosis.

  14. [The infection of dendritic cells by recombinant adenoviral vector carrying HBsAg-HSP70 chimeric gene and its biological characteristics observations].

    Science.gov (United States)

    Lei, Chun-Liang; Ou, Yang-Ling; Yang, Zhan; Tang, Xiao-Ping

    2009-02-01

    To test the infeciton efficiency of recombinant adenoviral vector carrying HBsAg-HSP70 chimeric gene and to abserve its biological characteristics. Peripheral blood mononuclear cells (PBMC) were separated from healthy blood donor and they were infected by Ad-HSP70-HBsAg on the first day of isolation. DCs were induced in medium with cytokines IL-4, GM-CSF and TNF-alpha in vitro. The biological characteristics of DC induced were analyzed by inverted fluorecent microscope, RT-PCR, flow cytometer (FACS), and mixed lymphocyte reaction (MLR). The traced gene-GFP were abserved in DCs by inverted fluorecent microscope and HSP70-HBsAg gene mRNA expression was detected by RT-PCR after the Ad-HSP70-HBsAg infection. FACS analysis shown that the expression of CD1a, CD80, CD86 and HLA-DR on surfece of two groups of DCs were similar. MLR showed that there are not a statitic difference of stimulated index (SI) between two groups. Results indicated that Ad-HSP70-HBsAg can effectively infected DCs without affecting its biological characteristics.

  15. Analysis of expression patterns of IGF-1, caspase-3 and HSP-70 in developing human tooth germs.

    Science.gov (United States)

    Kero, Darko; Kalibovic Govorko, Danijela; Medvedec Mikic, Ivana; Vukojevic, Katarina; Cigic, Livia; Saraga-Babic, Mirna

    2015-10-01

    To analyze expression patterns of IGF-1, caspase-3 and HSP-70 in human incisor and canine tooth germs during the late bud, cap and bell stages of odontogenesis. Head areas or parts of jaw containing teeth from 10 human fetuses aged between 9th and 20th developmental weeks were immunohistochemically analyzed using IGF-1, active caspase-3 and HSP-70 markers. Semi-quantitative analysis of each marker's expression pattern was also performed. During the analyzed period, IGF-1 and HSP-70 were mostly expressed in enamel organ. As development progressed, expression of IGF-1 and HSP-70 became more confined to differentiating tissues in the future cusp tip area, as well as in highly proliferating cervical loops. Few apoptotic bodies highly positive to active caspase-3 were observed in enamel organ and dental papilla from the cap stage onward. However, both enamel epithelia moderately expressed active caspase-3 throughout the investigated period. Expression patterns of IGF-1, active caspase-3 and HSP-70 imply importance of these factors for early human tooth development. IGF-1 and HSP-70 have versatile functions in control of proliferation, differentiation and anti-apoptotic protection of epithelial parts of human enamel organ. Active caspase-3 is partially involved in formation and apoptotic removal of primary enamel knot, although present findings might reflect its ability to perform other non-death functions such as differentiation of hard dental tissues secreting cells and guidance of ingrowth of proliferating cervical loops. Copyright © 2015 Elsevier Ltd. All rights reserved.

  16. Cytosolic Hsp70/Hsc70 protein expression in lymphocytes exposed to low gamma-ray dose

    Energy Technology Data Exchange (ETDEWEB)

    Manzanares A, E.; Vega C, H.R.; Letechipia de Leon, C. [Unidades Academicas de Estudios Nucleares, UAZ, A.P. 336, 98000 Zacatecas (Mexico)]. E-mail: emanz@cantera.reduaz.mx; Guzman E, L.J. [Unidad Academica de Biologia Experimental, Guadalupe, Zacatecas (Mexico); Garcia T, M. [LIBRA, Centro I and D, Campus Miguel Delibes, Valladolid 47011 (Spain)

    2004-07-01

    The purpose of this study was to evaluate the effect of low gamma ray intensity upon Hsp70 expression in human Iymphocytes. The heat shock proteins (Hsp) family, are a group of proteins present in all living organism, therefore there are highly conserved and are related to adaptation and evolution. At cellular level these proteins acts as chaperones correcting denatured proteins. When a stress agent, such heavy metals, UV, heat, etc. is affecting a cell a response to this aggression is triggered only through over expression of Hsp. Several studies has been carried out in which the cellular effect are observed, mostly of these studies uses large doses, but very few studies are related with low doses. Blood of healthy volunteers was obtained and the Iymphocytes were isolated by ficoll- histopaque gradient. Experimental lots were irradiated in a {sup 137} Cs gamma-ray. Hsp70 expression was found since 0.5 c Gy, indicating a threshold to very low doses of gamma rays. (Author)

  17. Functional analysis of Hsp70 superfamily proteins of rice (Oryza sativa).

    Science.gov (United States)

    Sarkar, Neelam K; Kundnani, Preeti; Grover, Anil

    2013-07-01

    Heat stress results in misfolding and aggregation of cellular proteins. Heat shock proteins (Hsp) enable the cells to maintain proper folding of proteins, both in unstressed as well as stressed conditions. Hsp70 genes encode for a group of highly conserved chaperone proteins across the living systems encompassing bacteria, plants, and animals. In the cellular chaperone network, Hsp70 family proteins interconnect other chaperones and play a dominant role in various cell processes. To assess the functionality of rice Hsp70 genes, rice genome database was analyzed. Rice genome contains 32 Hsp70 genes. Rice Hsp70 superfamily genes are represented by 24 Hsp70 family and 8 Hsp110 family members. Promoter and transcript expression analysis divulges that Hsp70 superfamily genes plays important role in heat stress. Ssc1 (mitochondrial Hsp70 protein in yeast) deleted yeast show compromised growth at 37 °C. Three mitochondrial rice Hsp70 sequences (i.e., mtHsp70-1, mtHsp70-2, and mtHsp70-3) complemented the Ssc1 mutation of yeast to differential extents. The information presented in this study provides detailed understanding of the Hsp70 protein family of rice, the crop species that is the major food for the world population.

  18. Expression of CAIII and Hsp70 Is Increased the Mucous Membrane of the Posterior Commissure in Laryngopharyngeal Reflux Disease.

    Science.gov (United States)

    Min, Hyun Jin; Hong, Seok Chan; Yang, Hoon Shik; Mun, Seog Kyun; Lee, Sei Young

    2016-03-01

    We tried to evaluate the difference in the expression of carbonic anhydrase (CA) III and heat shock protein (Hsp) 70 between laryngopharyngeal reflux disease (LPRD) and non-LPRD patients. The study involved 28 patients who underwent laryngeal microsurgery due to benign laryngeal disease from March to August 2008. Reflux symptom index (RSI) and reflux finding score (RFS) were measured for each person, and they were assigned either to the LPRD group (n=10) or non-LPRD group (n=18). Tissue samples were obtained from the mucosa of posterior commissure, and immunohistochemistry (IHC) staining of CAIII and Hsp70 was performed. The IHC scores were measured and compared with clinical features including RSI and RFS. Total 10 patients were assigned as LPRD group, and 18 patients were as control group. The mean IHC score of CAIII and Hsp70 was 1.70 ± 1.06 and 1.90 ± 0.88, respectively, in LPRD patients, whereas the mean IHC score of CAIII and Hsp70 was 0.78 ± 0.73 and 0.94 ± 0.87, respectively, in non-LPRD patients. The difference between two groups was statistically significant (p<0.05). CAIII and Hsp70 expressions were higher in LPRD patients that in non-LPRD patients, suggesting the possibility as one of biomomarker in LPRD diagnosis.

  19. [Effect of preventive acupuncture and moxibustion at "Guanyuan" (CV 4) on the expression of HSP 70 and HSP 70 mRNA in spleen and the contents of serum IL-2, TNF-alpha in menopausal rats].

    Science.gov (United States)

    Li, Xiao-Hong; Wang, Hong-Bin; Xu, Li-Li; Song, Xiao-Lin; Zheng, Ling; He, Yu-Wei; Zhang, Lu-Fen

    2009-04-01

    To observe the influence of preventive acupuncture (PA) and preventive moxibustion (PM) at "Guanyuan" (CV 4) on the immune function in natural climacteric rats. A total of 160 female SD rats were randomized into control, PA and PM groups, the former one group was further divided into 10 month (mon), 12 mon, 14 mon and 16 mon subgroups, and the later two groups were further divided into 12 mon, 14 mon and 16 mon subgroups, with 16 rats in each. In addition, other 16 female SD rats aged 3.5 mon were used as the young control (YC) group. "Guanyuan" (CV 4) was punctured with an acupuncture needle and the needle was retained for 20 min, or given with one ignited moxa-cone from the age of 10 mon on. The treatment was conducted twice every week, 8 weeks altogether. The expression of HSP 70 and HSP 70 mRNA of the spleen tissue was detected by using immunohistochemistry and in situ hybridization respectively, and serum IL-2 and TNF-alpha contents were assayed by using radio-immunoassay. In comparison with YC group, 1) the expression of spleen HSP 70 and HSP 70 mRNA increased significantly in 10 mon control (mon-C), 12 mon-PM and 12 mon-PA groups, and 14 mon-PA group (only HSP 70 mRNA) (P 0.05). Both preventive acupuncture and preventive moxibustion can upregulate the expression of spleen HSP 70 and HSP 70 mRNA and serum IL-2 and TNF-alpha levels, which may contribute to their effects in enhancing the immune function in menopausal rats.

  20. Association of HSP70-2 Gene 1267A/G Polymorphism With Cataract Incidence Among Guilan Population

    Directory of Open Access Journals (Sweden)

    Zivar Salehi

    2017-01-01

    Full Text Available Background: Cataract is a visible opacity of the eye lens and it is the main reason of reversible blindness in the world. Oxidative stress is known as a major factor in cataract formation HSP70-2 protein is a molecular chaperone which is essential for cell survival in stress conditions. HSP70-2 gene is located in the human leukocyte antigen class ΙΙΙ region. This gene encodes an inducible protein. One of the common polymorphism of HSP70-2 is 1267A/G which is located in coding region. The aim of this study was to analysis of 1267A/G polymorphism of hsp70-2 gene in cataract patients. Material and Methods: This case-control study included 118 cataract patients and 122 healthy people as a control groups. Genomic DNA was extracted from peripheral blood leukocytes and genotyping determination was performed by the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP method. Statistical analysis was performed by using the MedCalc software (Version 12.1. Results: The frequency of G allele was significantly higher in patients than the controls, (0.36 and 0.24, respectively. A higher frequency of the GG genotype of the HSP70-2 1267A/G polymorphism was found in the patients compared with controls (21.19% and 8.20%, respectively. The patients carrying the GG genotype were 3.2-fold at a higher risk of cataract compared with AA genotype (P=0.005. Conclusion: The finding of this study suggested that the HSP70-2 1267A/G may affect the susceptibility to cataract in the studied population. Anyway the randomized multicenter studies with greater sample size still need to confirmed our results.

  1. Reduced heat shock response in human mononuclear cells during aging and its association with polymorphisms in HSP70 genes

    DEFF Research Database (Denmark)

    Singh, Ripudaman; Kølvraa, Steen; Bross, Peter

    2006-01-01

    was measured by genotyping the subjects for 3 single nucleotide polymorphisms, HSPA1A(A-110C), HSPA1B(A1267G), and HSPA1L(T2437C), 1 each in the 3 HSP70 genes. A significant age-related decrease in the induction of Hsp70 occurred after heat shock in both monocytes and lymphocytes. The noninducible......Age-dependent changes in heat shock response (HSR) were studied in mononuclear cells (monocytes and lymphocytes) collected from young (mean age = 22.6 +/- 1.7 years) and middle-aged (mean age = 56.3 +/- 4.7 years) subjects after 1 hour of heat shock at 42 degrees C. Genotype-specific HSR...... and inducible forms of Hsp70 decreased 1.3-fold (P induction than TT carriers in both monocytes...

  2. Polymorphism of the gene of heat shock protein hsp70 in lines of rats with normal and hypertensive status

    NARCIS (Netherlands)

    Adarichev, V A; Dymshits, G M; Krivenko, A A; Ostapchuk, Ia V; Ostaptchouk, Jana

    1998-01-01

    RFLP in the hsp70 gene encoding a major heat shock protein was analyzed in rat strains with high and normal arterial blood pressure. Dimorphism in the sets of DNA fragments was revealed after hybridization of the hsp gene leader sequence with rat DNA digested with BamHI restriction endonuclease.

  3. Extracellular HSP70 Activates ERK1/2, NF-kB and Pro-Inflammatory Gene Transcription Through Binding with RAGE in A549 Human Lung Cancer Cells.

    Science.gov (United States)

    Somensi, Nauana; Brum, Pedro Ozorio; de Miranda Ramos, Vitor; Gasparotto, Juciano; Zanotto-Filho, Alfeu; Rostirolla, Diana Carolina; da Silva Morrone, Maurilio; Moreira, José Claudio Fonseca; Pens Gelain, Daniel

    2017-01-01

    Heat shock protein 70 (HSP70) has been recently described with extracellular actions, where it is actively released in inflammatory conditions. Acting as DAMPs (damage associated molecular pattern), extracellular HSP70 (eHSP70) interacts with membrane receptors and activates inflammatory pathways. At this context, the receptor for advanced glycation endproducts (RAGE) emerges as a possible candidate for interaction with eHSP70. RAGE is a pattern-recognition receptor and its expression is increased in several diseases related to a chronic pro-inflammatory state. One of the main consequences of RAGE ligand-binding is the ERK1/2 (extracellular signal-regulated kinases)-dependent activation of NF-kB (nuclear factor kappa B), which leads to expression of TNF-α (tumor necrosis factor alpha) and other cytokines. The purpose of this work is to elucidate if eHSP70 is able to evoke RAGE-dependent signaling using A549 human lung cancer cells, which constitutively express RAGE. Immunoprecipitation and protein proximity assay were utilized to demonstrate the linkage between RAGE and eHSP70. To investigate RAGE relevance on cell response to eHSP70, siRNA was used to knockdown the receptor expression. Signaling pathways activation were evaluated by western blotting, gene reporter luciferase and real time quantitative PCR. Protein eHSP70 shown to be interacting physically with the receptor RAGE in our cell model. Treatment with eHSP70 caused ERK1/2 activation and NF-κB transactivation impaired by RAGE knockdown. Moreover, the stimulation of pro-inflammatory cytokines expression by eHSP70 was inhibited in RAGE-silenced cells. Finally, conditioned medium of eHSP70-treated A549 cells caused differential effects in monocytes cytokine expression when A549 RAGE expression is inhibited. Our results evidence eHSP70 as a novel RAGE agonist capable of influence the cross-talk between cancer and immune system cells. © 2017 The Author(s). Published by S. Karger AG, Basel.

  4. Complex patterns of geographic variation in heat tolerance and Hsp70 expression levels in the common frog Rana temporaria

    DEFF Research Database (Denmark)

    Sørensen, Jesper Givskov; Pekkonen, Minna; Lindgren, Beatrice

    2009-01-01

    1. We tested for geographical variation in heat tolerance and Hsp70 expression levels of Rana temporaria tadpoles along a 1500 km long latitudinal gradient in Sweden.   2. Temperature tolerance of the hatchling tadpoles did not differ among populations, but they tolerated stressful hot temperatur...

  5. Optimization of expression and purification of human mortalin (Hsp70): Folding/unfolding analysis

    Science.gov (United States)

    Khan, Mohd Shahnawaz; Ahmed, Anwar; Tabrez, Shams; Islam, Badar ul; Rabbani, Nayyar; Malik, Ajamaluddin; Ismael, Mohamad A.; Alsenaidy, Mohammad A.; Alsenaidy, Abdulrahman M.

    2017-12-01

    Human mortalin is a Hsp70 mitochondrial protein that plays an essential role in the biogenesis of mitochondria. The deregulation of mortalin expression and its functions could lead to several age-associated disorders and some types of cancers. In the present study, we optimized the expression and purification of recombinant human mortalin by the use of two-step chromatography. Low temperature (18 °C) and 0.5 mM (IPTG) was required for optimum mortalin expression. Chaperone activity of mortalin was assessed by the citrate synthase and insulin protection assay, which suggested their protective role in mitochondria. Folding and unfolding assessments of mortalin were carried out in the presence of guanidine hydrochloride (GdnHCl) by intrinsic fluorescence measurement, ANS (8-analino 1-nephthlene sulfonic acid) binding and CD (circular dichroism) analysis. Under denaturing conditions, mortalin showed decrease in tryptophan fluorescence intensity along with a red shift of 11 nm. Moreover, ANS binding studies illustrated decrease in hydrophobicity. CD measurement of mortalin showed a predominant helical structure. However, the secondary structure was lost at low concentration of GdnHCl (1 M). We present a simple and robust method to produce soluble mortalin and warranted that chaperones are also susceptible to unfolding and futile to maintain protein homeostasis.

  6. Microbiota Composition, HSP70 and Caspase-3 Expression as Marker for Colorectal Cancer Patients in Aceh, Indonesia.

    Science.gov (United States)

    Yusuf, Fauzi; Ilyas, Syafruddin; Damanik, Harun Al Rasyid; Fatchiyah, Fatchiyah

    2016-10-01

    to investigate the relationship between microbiota composition with HSP70 and Caspase-3 expressions in colon tissue as an initial study to develop the candidate for early detection of colorectal cancer for Indonesian patients. this is a cross-sectional study on 32 patients undergoing colonoscopy; 16 patients of colorectal cancer (CRC) while the other 16 patients are not (colitis and internal hemorrhoid). The composition of microbiota in stool samples was examined using 16S rRNA Denaturing Gradient Gel Electrophoresis (DDGE) while expression of HSP70 was examined by immunohistochemistry and Caspase-3 by using Haematoxylin-Eosin(HE) staining to determine the morphological changes in colon tissue. analysis of PCR-DDGE shows a different composition of microbiota between patients with CRC and non-CRC. All CRC patients showed disappearance of dominant band from Bifidobacterium groups. Histological observation based on Inter Class Correlation (ICC) test from all slide showed a high scores (5.2-9.2) in CRC patients and low scores (1.7-2.4) in non-CRC patients. HSP70 expression was increased significantly in CRC patients with the highest percentage of 84%, while expression of caspase-3 decreased with the highest percentage of 21%. Statistical analysis showed that the incidence of colorectal cancer was associated with the expression of HSP 70 (p<0.001), and Caspase 3 (p<0.001). bifidobacterium is an important indicator for colorectal cancer patients that show disappearance of dominant band, while expression of HSP70 increased and the Caspase-3 expression decreased significantly.

  7. Microbiota Composition, HSP70 and Caspase-3 Expression as Marker for Colorectal Cancer Patients in Aceh, Indonesia

    Directory of Open Access Journals (Sweden)

    Fauzi Yusuf

    2017-02-01

    Full Text Available Aim: to investigate the relationship between microbiota composition with HSP70 and Caspase-3 expressions in colon tissue as an initial study to develop the candidate for early detection of colorectal cancer for Indonesian patients. Methods: this is a cross-sectional study on 32 patients undergoing colonoscopy; 16 patients of colorectal cancer (CRC while the other 16 patients are not (colitis and internal hemorrhoid. The composition of microbiota in stool samples was examined using 16S rRNA Denaturing Gradient Gel Electrophoresis (DDGE while expression of HSP70 was examined by immunohistochemistry and Caspase-3 by using Haematoxylin-Eosin(HE staining to determine the morphological changes in colon tissue. Results: analysis of PCR-DDGE shows a different composition of microbiota between patients with CRC and non-CRC. All CRC patients showed disappearance of dominant band from Bifidobacterium groups. Histological observation based on Inter Class Correlation (ICC test from all slide showed a high scores (5.2-9.2 in CRC patients and low scores (1.7-2.4 in non-CRC patients. HSP70 expression was increased significantly in CRC patients with the highest percentage of 84%, while expression of caspase-3 decreased with the highest percentage of 21%. Statistical analysis showed that the incidence of colorectal cancer was associated with the expression of HSP 70 (p<0.001, and Caspase 3 (p<0.001. Conclusion: bifidobacterium is an important indicator for colorectal cancer patients that show disappearance of dominant band, while expression of HSP70 increased and the Caspase-3 expression decreased significantly.

  8. The Possible Involvement of HLA Class III Haplotype (RAGE, HSP70 and TNF Genes) in Alzheimer's Disease.

    Science.gov (United States)

    Boiocchi, Chiara; Maggioli, Elisa; Monti, Maria Cristina; Zorzetto, Michele; Sinforiani, Elena; Cereda, Cristina; Ricevuti, Giovanni; Cuccia, Mariaclara

    2015-01-01

    In the central nervous system Hsp70s seems to have a protective role in repair and removal of cellular proteins damaged by stress conditions. A protective role of Hsp70 was also shown in Alzheimer Disease. The HSP70-1 +190 G/C polymorphism is located in the gene 5'UTR region and it is implicated in alteration of the transcription binding factor; HSP70-2 +1267 A/G causes a silent mutation in the coding region and it seems to influence the mechanism of mRNA translation; HSP70-hom +2437 A/G causes a substitution Met → The (M493T) in the coding region and it seems to influence the bond with the substrate and therefore on the chaperone activity of hsp70. The aim of our study will be to investigate Alzheimer susceptibility to Hsp70 polymorphisms, taking into account our previous findings on HLA class III region, and to hypothesize a role of HLA class III haplotype configuration based on the variants of three genes: RAGE, HSP70 and TNF. We studied these polymorphisms with PCR-RFLP and PCR-TSP. We investigated 173 AD patients and 211 control subjects. Our results have shown a statistically significant decrease of the C allele frequency of the HSP70-1 +190 G/C polymorphism in AD patients vs controls (P value = 0,018), as well as the G allele of HSP70-2 +1267 G/A (p value = 0,02). We focalized our attention on haplotype reconstruction. We have observed a significant statistically decrease of GGT haplotype frequency (empirical p-value=0.0133 ); GAT haplotype was statistically significant increase in AD patients compared with control (empirical p-value=0.007). The total HLA class III haplotype are reconstructed. The causative haplotypes are the following ones: TTGATGGG ( p value =0,005; empirical p =0,0042); TTGATAGG (p value =0,45; empirical p =0,034). Patients with these haplotypes may show an earlier onset of the disease than patients with TTGGTGGG (p value=0,0138; empirical p =0,0102); TTCGTGGG (p value=0,021; empirical p =0,017); TTCGTGGA (p value =0,058; empirical p =0

  9. Analysis of association between HSP70 expression in tumor tissues and clinicopathological characteristics of HBV-related hepatocellular carcinoma

    Directory of Open Access Journals (Sweden)

    GAO Jialin

    2015-05-01

    Full Text Available ObjectiveTo investigate the relationship between heat shock protein 70 (HSP70 expression and clinicopathological characteristics of hepatitis virus B (HBV-induced hepatocellular carcinoma (HCC. MethodsStatistical analysis was conducted to explore the association between the data of HSP70 obtained from the gene expression omnibus (GEO and the clinicopathological characteristics of HCC. As for categorical data, frequency (proportion was used to describe and chi-square test or Fisher′s exact test was applied to analyze. Univariate and multivariate logistic regression analyses were used to investigate the relationship between HSP expression and the clinicopathological characteristics of HCC. The variables with a P value less than 0.10 were included for subsequent multivariate analysis. Results1 The percentage of the HCC patients whose primary tumor sizes were more than 5 cm was significantly higher in the group with high expression of HSPA4L, HSPA6, and HSPA13 than in the group with low expression (P<0.05; in comparison with the group with high expression of HSPA1A and HSPA6, the group with low expression had a significantly lower percentage of patients accompanied by liver cirrhosis (P<0.05; there was a significantly higher percentage of patients with alpha-fetoprotein (AFP levels more than 300 ng/ml in the group with high expression of HSPA5 and HSA13 compared with the group with low expression (P<0.01. 2 The survival rates of HCC patients with liver cirrhosis and/or primary tumor that was more than 5 cm significantly decreased (P<0.05. 3 Highly expressed HSPA4L might promote the growth of HCC (OR=1.019, 95% CI:1.006-10.032, P=0.003; the patients with high expression of HSPA1B might have a higher risk of developing multiple nodules (OR=1.002, 95%CI:1.000-1.004, P=0.003. ConclusionThe expression of HSPA4L and HSPA1B is, to some extent, associated with the clinicopathological characteristics of HCC, which might be a therapeutic target and

  10. The genes coding for the hsp70(dnaK) molecular chaperone machine occur in the moderate thermophilic archaeon Methanosarcina thermophila TM-1

    DEFF Research Database (Denmark)

    Hofman-Bang, H Jacob Peider; Lange, Marianne; Ahring, Birgitte Kiær

    1999-01-01

    of response by hsp70(dnaK), and a similar response by trkA. The data suggest that the moderate thermophile TM-1 has an active Hsp70(DnaK)-chaperone machine in contrast to hyperthermophilic archaea, and that trkA is a stress gene, inasmuch as it responds like classic heat-shock genes to stressors that induce...

  11. Both heat shock and water deprivation trigger Hsp70 expression in the olfactory lobe of the crab Chasmagnathus granulatus.

    Science.gov (United States)

    Frenkel, Lia; Dimant, Beatriz; Portiansky, Enrique L; Maldonado, Héctor; Delorenzi, Alejandro

    2008-10-10

    Heat-shock proteins (Hsp) are synthesized in the central nervous system in response to traumas but also after physical exercise and psychophysiological stress. Therefore, an increase in Hsp expression is a good marker of changes in metabolic activity. In the crab Chasmagnathus, a powerful memory paradigm has been established. Memory modulation is possible by water shortage. The brain areas activated by either training protocols and/or water-deprivation are still unknown. Hsp expression might be a marker to sensing the increase in metabolic activity in crab Chasmagnathus brain neuropils engaged in the physiological responses triggered by water deprivation and cognitive processing. Here, we observed an increase in brain Hsp of 70kDa (Hsp70) expression after a heat-shock treatment. Additionally, immunohistochemistry analysis revealed that, under basal conditions, some glomeruli of the olfactory lobes showed Hsp70 immunoreactivity in an on-off manner. Both a hot environment and water deprivation increased the number of glomeruli expressing Hsp70. This marker of neuropil's activity might turn out to be a powerful tool to test whether crustacean olfactory lobes not only process olfactory information but also integrate multimodal signals.

  12. Mesothelioma Cells Escape Heat Stress by Upregulating Hsp40/Hsp70 Expression via Mitogen-Activated Protein Kinases

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    Michael Roth

    2009-01-01

    Full Text Available Therapy with hyperthermal chemotherapy in pleural diffuse malignant mesothelioma had limited benefits for patients. Here we investigated the effect of heat stress on heat shock proteins (HSP, which rescue tumour cells from apoptosis. In human mesothelioma and mesothelial cells heat stress (39–42°C induced the phosphorylation of two mitogen activated kinases (MAPK Erk1/2 and p38, and increased Hsp40, and Hsp70 expression. Mesothelioma cells expressed more Hsp40 and were less sensitive to heat stress compared to mesothelial cells. Inhibition of Erk1/2 MAPK by PD98059 or by Erk1 siRNA down-regulated heat stress-induced Hsp40 and Hsp70 expression and reduced mesothelioma cell survival. Inhibition of p38MAPK by SB203580 or siRNA reduced Hsp40, but not Hsp70, expression and also increased mesothelioma cell death. Thus hyperthermia combined with suppression of p38 MAPK or Hsp40 may represent a novel approach to improve mesothelioma therapy.

  13. Schiff Base Metal Derivatives Enhance the Expression of HSP70 and Suppress BAX Proteins in Prevention of Acute Gastric Lesion

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    Shahram Golbabapour

    2013-01-01

    Full Text Available Schiff base complexes have appeared to be promising in the treatment of different diseases and disorders and have drawn a lot of attention to their biological activities. This study was conducted to evaluate the regulatory effect of Schiff base metal derivatives on the expression of heat shock proteins (HSP 70 and BAX in protection against acute haemorrhagic gastric ulcer in rats. Rats were assigned to 6 groups of 6 rats: the normal control (Tween 20 5% v/v, 5 mL/kg, the positive control (Tween 20 5% v/v, 5 mL/kg, and four Schiff base derivative groups named Schiff_1, Schiff_2, Schiff_3, and Schiff_4 (25 mg/kg. After 1 h, all of the groups received ethanol 95% (5 mL/kg but the normal control received Tween 20 (Tween 20 5% v/v, 5 mL/kg. The animals were euthanized after 60 min and the stomachs were dissected for histology (H&E, immunohistochemistry, and western blot analysis against HSP70 and BAX proteins. The results showed that the Schiff base metal derivatives enhanced the expression of HSP70 and suppressed the expression of BAX proteins during their gastroprotection against ethanol-induced gastric lesion in rats.

  14. Schiff Base Metal Derivatives Enhance the Expression of HSP70 and Suppress BAX Proteins in Prevention of Acute Gastric Lesion

    Science.gov (United States)

    Gwaram, Nura Suleiman; Al-Obaidi, Mazen M. Jamil; Soleimani, A. F.; Ali, Hapipah Mohd; Abdul Majid, Nazia

    2013-01-01

    Schiff base complexes have appeared to be promising in the treatment of different diseases and disorders and have drawn a lot of attention to their biological activities. This study was conducted to evaluate the regulatory effect of Schiff base metal derivatives on the expression of heat shock proteins (HSP) 70 and BAX in protection against acute haemorrhagic gastric ulcer in rats. Rats were assigned to 6 groups of 6 rats: the normal control (Tween 20 5% v/v, 5 mL/kg), the positive control (Tween 20 5% v/v, 5 mL/kg), and four Schiff base derivative groups named Schiff_1, Schiff_2, Schiff_3, and Schiff_4 (25 mg/kg). After 1 h, all of the groups received ethanol 95% (5 mL/kg) but the normal control received Tween 20 (Tween 20 5% v/v, 5 mL/kg). The animals were euthanized after 60 min and the stomachs were dissected for histology (H&E), immunohistochemistry, and western blot analysis against HSP70 and BAX proteins. The results showed that the Schiff base metal derivatives enhanced the expression of HSP70 and suppressed the expression of BAX proteins during their gastroprotection against ethanol-induced gastric lesion in rats. PMID:24298554

  15. Molecular karyotype and chromosomal localization of genes encoding ß-tubulin, cysteine proteinase, hsp 70 and actin in Trypanosoma rangeli

    OpenAIRE

    CB Toaldo; Steindel, M; MA Sousa; CC Tavares

    2001-01-01

    The molecular karyotype of nine Trypanosoma rangeli strains was analyzed by contour-clamped homogeneous electric field electrophoresis, followed by the chromosomal localization of ß-tubulin, cysteine proteinase, 70 kDa heat shock protein (hsp 70) and actin genes. The T. rangeli strains were isolated from either insects or mammals from El Salvador, Honduras, Venezuela, Colombia, Panama and southern Brazil. Also, T. cruzi CL-Brener clone was included for comparison. Despite the great similarity...

  16. A fraction from Petiveria alliacea induces apoptosis via a mitochondria-dependent pathway and regulates HSP70 expression

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    Susana Fiorentino

    2009-12-01

    Full Text Available To evaluate the biological activity of Petiveria alliacea extracts on tumoral cells in vitro. Materials and methods. P.alliaceafractions prepared by a bioguided purification protocol were characterized by their biological activities on two human tumoral cell lines.Morphological changes, cell viability, mitochondrial membrane depolarization, nuclear staining and activity on HSP70 were analyzed.Results. The present study demonstrates that P.alliacea fractions can induce apoptosis in a mitochondria-dependent pathway and downregulate HSP70 expression in vitro. The possible compounds present in P.alliacea responsible for the described biological activities wereidentified by dereplication methods. Conclusion. The present study provides new knowledge on the antitumoral properties of the plantspecies P. alliacea described in several ethnobotanical studies.

  17. Effect of neoadjuvant chemoradiation and postoperative radiotherapy on expression of heat shock protein 70 (HSP70 in head and neck vessels

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    Gellrich Nils-Claudius

    2011-07-01

    Full Text Available Abstract Background Preoperative radiotherapy and chemotherapy in patients with head and neck cancer result in changes to the vessels that are used to construct microsurgical anastomoses. The aim of the study was to investigate quantitative changes and HSP70 expression of irradiated neck recipient vessels and transplant vessels used for microsurgical anastomoses. Methods Of 20 patients included in this study five patients received neoadjuvant chemoradiation, another five received conventional radiotherapy and 10 patients where treated without previous radiotherapy. During surgical procedure, vessel specimens where obtained by the surgeon. Immunhistochemical staining of HSP70 was performed and quantitative measurement and evaluation of HSP70 was carried out. Results Conventional radiation and neoadjuvant chemoradiation revealed in a thickening of the intima layer of recipient vessels. A increased expression of HSP70 could be detected in the media layer of the recipient veins as well as in the transplant veins of patients treated with neoadjuvant chemoradiation. Radiation and chemoradiation decreased the HSP70 expression of the intima layer in recipient arteries. Conventional radiation led to a decrease of HSP70 expression in the media layer of recipient arteries. Conclusion Our results showed that anticancer drugs can lead to a thickening of the intima layer of transplant and recipient veins and also increase the HSP70 expression in the media layer of the recipient vessels. In contrast, conventional radiation decreased the HSP70 expression in the intima layer of arteries and the media layer of recipient arteries and veins. Comparing these results with wall thickness, it was concluded, that high levels of HSP70 may prevent the intima layer of arteries and the media layer of vein from thickening.

  18. Environmental stress-dependent effects of deletions encompassing Hsp70Ba on canalization and quantitative trait asymmetry in Drosophila melanogaster.

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    Kazuo H Takahashi

    Full Text Available Hsp70 genes may influence the expression of wing abnormalities in Drosophila melanogaster but their effects on variability in quantitative characters and developmental instability are unclear. In this study, we focused on one of the six Hsp70 genes, Hsp70Ba, and investigated its effects on within- and among-individual variability in orbital bristle number, sternopleural bristle number, wing size and wing shape under different environmental conditions. To do this, we studied a newly constructed deletion, Df(3RED5579, which encompasses Hsp70Ba and nine non-Hsp genes, in the heterozygous condition and another, Hsp70Ba(304, which deletes only Hsp70Ba, in the homozygous condition. We found no significant effect of both deletions on within-individual variation quantified by fluctuating asymmetry (FA of morphological traits. On the other hand, the Hsp70Ba(304/Hsp70Ba(304 genotype significantly increased among-individual variation quantified by coefficient of variation (CV of bristle number and wing size in female, while the Df(3RED5579 heterozygote showed no significant effect. The expression level of Hsp70Ba in the deletion heterozygote was 6 to 20 times higher than in control homozygotes, suggesting that the overexpression of Hsp70Ba did not influence developmental stability or canalization significantly. These findings suggest that the absence of expression of Hsp70Ba increases CV of some morphological traits and that HSP70Ba may buffer against environmental perturbations on some quantitative traits.

  19. Environmental Stress-Dependent Effects of Deletions Encompassing Hsp70Ba on Canalization and Quantitative Trait Asymmetry in Drosophila melanogaster

    Science.gov (United States)

    Takahashi, Kazuo H.; Daborn, Phillip J.; Hoffmann, Ary A.; Takano-Shimizu, Toshiyuki

    2011-01-01

    Hsp70 genes may influence the expression of wing abnormalities in Drosophila melanogaster but their effects on variability in quantitative characters and developmental instability are unclear. In this study, we focused on one of the six Hsp70 genes, Hsp70Ba, and investigated its effects on within-and among-individual variability in orbital bristle number, sternopleural bristle number, wing size and wing shape under different environmental conditions. To do this, we studied a newly constructed deletion, Df(3R)ED5579, which encompasses Hsp70Ba and nine non-Hsp genes, in the heterozygous condition and another, Hsp70Ba304, which deletes only Hsp70Ba, in the homozygous condition. We found no significant effect of both deletions on within-individual variation quantified by fluctuating asymmetry (FA) of morphological traits. On the other hand, the Hsp70Ba304/Hsp70Ba304 genotype significantly increased among-individual variation quantified by coefficient of variation (CV) of bristle number and wing size in female, while the Df(3R)ED5579 heterozygote showed no significant effect. The expression level of Hsp70Ba in the deletion heterozygote was 6 to 20 times higher than in control homozygotes, suggesting that the overexpression of Hsp70Ba did not influence developmental stability or canalization significantly. These findings suggest that the absence of expression of Hsp70Ba increases CV of some morphological traits and that HSP70Ba may buffer against environmental perturbations on some quantitative traits. PMID:21541022

  20. Expression of hsp70 and ferritin in embryos of the copepod Acartia tonsa (Dana) during transition between subitaneous and quiescent state

    DEFF Research Database (Denmark)

    Nilsson, Birgitte; Jepsen, Per Meyer; Rewitz, Kim Furbo

    2014-01-01

    . However, ferritin expression exhibited a strong increase from days 3 to 5 of quiescence followed by a declined. After 2 weeks of quiescence, development of the embryos recovered by the addition of oxygen and increased temperature. The expression of both genes increased 222-fold for hsp70 and 52-fold...

  1. Thermal stress in Larvae of the sand dollar (Dendraster excentricus) induces changes in hsp70 gen expression

    Science.gov (United States)

    Olivares-Bañuelos, T.; Garcia-Echauri, L.; Figueroa-Flores, S.; Carpizo-Ituarte, E.

    2010-03-01

    The relationship between temperature and individual performance is reasonably well understood, and much climate-related research has focused on potential shifts in distribution and abundance driven directly by temperature(1). It is known, that global climate change has profound implications for marine ecosystems and the economic and social systems that depend upon them. In virtually all organisms, including equinoderms, cells respond to a variety of stresses, as temperature changes, by the rapid synthesis of a highly conserved set of polypeptides termed heat shock proteins (HSPs) (2). Differential Hsp expression among species may allow functionally important genetic variation to cope with thermal stress and other environmental stressors, allowing populations to respond effectively to environmental change. The aim of the present work was to elucidate the effect of different temperatures on hsp70 expression in larvae of the sand dollar Dendraster excentricus. We are interested to understand what is how organisms that live in estuaries respond to global warming. We did thermo tolerance assays whit 3 different larval stages of the sand dollar, including 8 arms plutei, competent and metamorphic larvae. We tested 4 experimental temperatures (4, 20, 26, and 29°C) and 3 different times (1, 3 and 6 hours). After each treatment, total RNA was extracted and used to quantify hsp70 expression levels using real time PCR analysis. We detected, after 6 hours of thermal stress, the highest over-expression levels of hsp70 in both metamorphic and competent larvae. In 8 arms pluetus larvae, no significant differences in hsp70 expression were observed after the exposure to the experimental temperatures. Taking together our results, indicate that larvae of Dendraster excentricus are responding according to their development stage to cope with thermal-stress. The capability of sand dollar larvae to acclimate and respond effectively to the climatic change would be determinated by the sea

  2. Exposure to low-frequency electromagnetic fields does not alter HSP70 expression or HSF-HSE binding in HL60 cells.

    Science.gov (United States)

    Morehouse, C A; Owen, R D

    2000-05-01

    Environmental exposure to extremely low-frequency electromagnetic fields (ELF EMFs) has been identified as a possible contributor to increased cancer incidence and other diseases. In vitro studies designed to probe for biological mechanisms that might explain such relationships have included several studies of gene expression. While gene expression studies have focused on MYC, effects of ELF EMFs on the expression of beta-actin, histone H2B, beta-tubulin, SRC, FOS and JUN have also been reported. In addition, some investigators have reported both an induction of HSP70 expression and an increase in HSF-HSE binding in HL60 (human promyelocytic leukemia) cells after exposure to a 60 Hz magnetic field. In this study, HL60 cells were exposed to a weak 60 Hz magnetic field (6.3 or 8.0 microT) or to a positive control heat shock (42 or 44 degrees C). While heat shock led to reproducible induction of HSP70 expression and HSF-HSE binding, no significant effect of exposure to ELF EMFs on either of these end points was observed.

  3. Molecular karyotype and chromosomal localization of genes encoding ß-tubulin, cysteine proteinase, hsp 70 and actin in Trypanosoma rangeli

    Directory of Open Access Journals (Sweden)

    CB Toaldo

    2001-01-01

    Full Text Available The molecular karyotype of nine Trypanosoma rangeli strains was analyzed by contour-clamped homogeneous electric field electrophoresis, followed by the chromosomal localization of ß-tubulin, cysteine proteinase, 70 kDa heat shock protein (hsp 70 and actin genes. The T. rangeli strains were isolated from either insects or mammals from El Salvador, Honduras, Venezuela, Colombia, Panama and southern Brazil. Also, T. cruzi CL-Brener clone was included for comparison. Despite the great similarity observed among strains from Brazil, the molecular karyotype of all T. rangeli strains analyzed revealed extensive chromosome polymorphism. In addition, it was possible to distinguish T. rangeli from T. cruzi by the chromosomal DNA electrophoresis pattern. The localization of ß-tubulin genes revealed differences among T. rangeli strains and confirmed the similarity between the isolates from Brazil. Hybridization assays using probes directed to the cysteine proteinase, hsp 70 and actin genes discriminated T. rangeli from T. cruzi, proving that these genes are useful molecular markers for the differential diagnosis between these two species. Numerical analysis based on the molecular karyotype data revealed a high degree of polymorphism among T. rangeli strains isolated from southern Brazil and strains isolated from Central and the northern South America. The T. cruzi reference strain was not clustered with any T. rangeli strain.

  4. Molecular karyotype and chromosomal localization of genes encoding beta-tubulin, cysteine proteinase, hsp 70 and actin in Trypanosoma rangeli.

    Science.gov (United States)

    Toaldo, C B; Steindel, M; Sousa, M A; Tavares, C C

    2001-01-01

    The molecular karyotype of nine Trypanosoma rangeli strains was analyzed by contour-clamped homogeneous electric field electrophoresis, followed by the chromosomal localization of beta-tubulin, cysteine proteinase, 70 kDa heat shock protein (hsp 70) and actin genes. The T. rangeli strains were isolated from either insects or mammals from El Salvador, Honduras, Venezuela, Colombia, Panama and southern Brazil. Also, T. cruzi CL-Brener clone was included for comparison. Despite the great similarity observed among strains from Brazil, the molecular karyotype of all T. rangeli strains analyzed revealed extensive chromosome polymorphism. In addition, it was possible to distinguish T. rangeli from T. cruzi by the chromosomal DNA electrophoresis pattern. The localization of beta-tubulin genes revealed differences among T. rangeli strains and confirmed the similarity between the isolates from Brazil. Hybridization assays using probes directed to the cysteine proteinase, hsp 70 and actin genes discriminated T. rangeli from T. cruzi, proving that these genes are useful molecular markers for the differential diagnosis between these two species. Numerical analysis based on the molecular karyotype data revealed a high degree of polymorphism among T. rangeli strains isolated from southern Brazil and strains isolated from Central and the northern South America. The T. cruzi reference strain was not clustered with any T. rangeli strain.

  5. RNA polymerase II interacts with the promoter region of the noninduced hsp70 gene in Drosophila melanogaster cells.

    Science.gov (United States)

    Gilmour, D S; Lis, J T

    1986-11-01

    By using a protein-DNA cross-linking method (D. S. Gilmour and J. T. Lis, Mol. Cell. Biol. 5:2009-2018, 1985), we examined the in vivo distribution of RNA polymerase II on the hsp70 heat shock gene in Drosophila melanogaster Schneider line 2 cells. In heat shock-induced cells, a high level of RNA polymerase II was detected on the entire gene, while in noninduced cells, the RNA polymerase II was confined to the 5' end of the hsp70 gene, predominantly between nucleotides -12 and +65 relative to the start of transcription. This association of RNA polymerase II was apparent whether the cross-linking was performed by a 10-min UV irradiation of chilled cells with mercury vapor lamps or by a 40-microsecond irradiation of cells with a high-energy xenon flash lamp. We hypothesize that RNA polymerase II has access to, and a high affinity for, the promoter region of this gene before induction, and this poised RNA polymerase II may be critical in the mechanism of transcription activation.

  6. Expression of Hsp70, Igf1, and Three Oxidative Stress Biomarkers in Response to Handling and Salt Treatment at Different Water Temperatures in Yellow Perch, Perca flavescens

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    Nour Eissa

    2017-09-01

    Full Text Available Stress is a major factor that causes diseases and mortality in the aquaculture industry. The goal was to analyze the expression of stress-related biomarkers in response to different stressors in yellow perch, which is an important aquaculture candidate in North America and highly sensitive to handling in captivity. Three fish groups were established, each having four replicates, and subjected to water temperatures of 14, 20, and 26°C and acute handling stress was performed followed by a salt treatment for 144h at a salinity of 5 ppt. Serum and hepatic mRNA levels of heat shock protein (hsp70, insulin-like growth factor 1 (Igf1, glutathione peroxidase (Gpx, superoxide dismutase 1 (Sod1, and glutathione reductase (Gsr were quantified at seven times interval over 144 h using ELISA and RT-qPCR. Handling stress caused a significant down-regulation in Hsp70, Gpx, Sod1, and Gsr at a water temperature of 20°C compared to 14 and 26°C. Igf1 was significantly upregulated at 20°C and down-regulated at 14 and 26°C. Salt treatment had a transient reverse effect on the targeted biomarkers in all groups at 72 h, then caused an upregulation after 144 h, compared to the control groups. The data showed a negative strong regulatory linear relationship between igf1 with hsp70 and anti-oxidative gene expressions. These findings could provide valuable new insights into the stress responses that affect fish health and could be used to monitor the stress.

  7. Fluoro-edenite Fibers Induce Expression of Hsp70 and Inflammatory Response

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    Michael Balazy

    2007-09-01

    Full Text Available Many asbestos-like mineral fibers have been detected in the air of mountainous and volcanic areas of Italy and other parts of the world. These fibers have been suspected to be the cause of increased incidences of lung cancer and other lung diseases in these areas. However, the mechanisms of the cellular response and defense following exposure to these microscopic fibers have not been characterized. We continue to study these mechanisms to be able to propose preventive strategies in large populations. The objective of the present study was to determine comparatively biological responses of mesothelial Met-5A and monocyte-macrophage J774 cells following exposure to two types of fluoro-edenite fibers having low and high iron content (labeled 19 and 27, respectively obtained from Biancavilla (Sicily, Italy. The reference fiber was a non-iron fibrous tremolite from Val di Susa (Piemonte, Italy. The cells were treated with 5, 50, and 100 μg of fibrous matter per 1 ml for 72 hr. We identified several key mechanisms by which cells responded and counteracted the injury induced by these fibers. The fibers caused induction of the heat shock protein 70 (Hsp70, stimulated formation of reactive oxygen species (detected by using DCFH-DA as a fluorescent probe and NO• (measured as nitrite. Exposure of cells to the fibers induced lactate dehydrogenase activity and decreased viability. The fluoro-endenite type 27 was the most potent fiber tested, which indicated that iron and possibly manganese contribute significantly to this fiber toxicity. The J774 cells were more sensitive to fluoro-edenite than Met-5A cells suggesting that the primary site of the fiberinduced inflammatory response could be the macrophage rather than the pulmonary epithelium. Fluoro-edenite produces more biological alterations with respect to non-iron tremolite. Hsp70 and free radicals could be important factors in the context of mineral fiber-induced acute lung injury

  8. Effect of resistance exercise training on expression of Hsp70 and inflammatory cytokines in skeletal muscle and adipose tissue of STZ-induced diabetic rats.

    Science.gov (United States)

    Molanouri Shamsi, M; Mahdavi, M; Quinn, L S; Gharakhanlou, R; Isanegad, A

    2016-09-01

    Impairment of adipose tissue and skeletal muscles accrued following type 1 diabetes is associated with protein misfolding and loss of adipose mass and skeletal muscle atrophy. Resistance training can maintain muscle mass by changing both inflammatory cytokines and stress factors in adipose tissue and skeletal muscle. The purpose of this study was to determine the effects of a 5-week ladder climbing resistance training program on the expression of Hsp70 and inflammatory cytokines in adipose tissue and fast-twitch flexor hallucis longus (FHL) and slow-twitch soleus muscles in healthy and streptozotocin-induced diabetic rats. Induction of diabetes reduced body mass, while resistance training preserved FHL muscle weight in diabetic rats without any changes in body mass. Diabetes increased Hsp70 protein content in skeletal muscles, adipose tissue, and serum. Hsp70 protein levels were decreased in normal and diabetic rats by resistance training in the FHL, but not soleus muscle. Furthermore, resistance training decreased inflammatory cytokines in FHL skeletal muscle. On the other hand, Hsp70 and inflammatory cytokine protein levels were increased by training in adipose tissue. Also, significant positive correlations between inflammatory cytokines in adipose tissue and skeletal muscles with Hsp70 protein levels were observed. In conclusion, we found that in diabetic rats, resistance training decreased inflammatory cytokines and Hsp70 protein levels in fast skeletal muscle, increased adipose tissue inflammatory cytokines and Hsp70, and preserved FHL muscle mass. These results suggest that resistance training can maintain skeletal muscle mass in diabetes by changing inflammatory cytokines and stress factors such as Hsp70 in skeletal muscle and adipose tissue.

  9. Expression of the heat shock protein Hsp70 in chloride target cells of mayfly larvae from motorway retention pond: a biomarker of osmotic shock.

    Science.gov (United States)

    De Jong, Laetitia; Moreau, Xavier; Jean, Séverine; Scher, Olivier; Thiéry, Alain

    2006-07-31

    The aim of this work was to study, by an in toto immunohistochemical technique, the expression pattern of the heat shock protein, Hsp70, in the widely used bioindicator species Cloeon dipterum (Linnaeus 1761) (Ephemeroptera, Baetidae), living in a motorway retention pond. All sampling and measurements have been performed from March 2002 to March 2003. The water physicochemical analyses have revealed a large increase in Na(+) and Cl(-) concentrations after the de-icing road surface in winter related to motorway maintenance that correspond to an osmotic shock (from 3.1 to 105.7 mg L(-1) for Na(+) and from 3.5 to 193.9 mg L(-1) for Cl(-)). An expression of Hsp70 was observed in the chloride cells only during the osmotic shock. In contrast, the gill insertions were Hsp70 immunoreactive in specimens collected all along the year. For comparison, the expression of Hsp70 was investigated in specimens collected in a temporary pond. C. dipterum larvae living in this pond, not submitted to such osmotic shock, do not express Hsp70 neither in chloride cells nor in gill insertions. Likewise, the expression of Hsp70 was not detected in these structures during the drying period when the abiotic conditions become progressively stressful (elevation of temperature and anoxia). As chloride cells play a key role in osmoregulation, their functional integrity is crucial for the survival of the mayfly larvae in occasionally salty freshwaters. According to the well known protective role of the Hsp70 stress proteins, it is likely that the induction of Hsp70 may protect the chloride cells from osmotic shock injuries resulting from the increase in salinity. So, the Hsp70 induction in chloride cells is designed as a useful biomarker of osmotic shock. The in toto immunohistochemical detection of Hsp70 allows to characterize both the exposure situation and biological effects in target cells induced by stresses. This method could be used as a complementary qualitative approach in the biomarker

  10. Anti-stress properties and two HSP70s mRNA expressions of blunt snout bream (Megalobrama amblycephala) fed with all-plant-based diet.

    Science.gov (United States)

    Deng, Wei; Zhao, Yuhua; Wang, Weimin; Gul, Yasmeen; Cao, Junming; Huang, Yanhua; Sheng, Guangcheng; Ding, Zhujin; Du, Rui

    2014-06-01

    The influence of all-plant-based diet on fingerling blunt snout breams (Megalobrama amblycephala) was tested by examining growth performance, anti-stress properties and related gene expression. Healthy fish were randomly divided into triplicate groups per dietary treatment and fed with different formulated diets. The results showed that both weight gain, specific growth rate and protein efficiency ratio of all-plant-based diet group were significant higher than those of the control (p plant-based diet group was significantly lower than that of the control (p plant-based diets could not affect the growth performance of blunt snout breams. Compared to the control group, the lysozyme levels in serum and mucus, and glutamic-oxaloacetic transaminase activities in serum and liver decreased significantly (p plant-based diets, the superoxide dismutase activities in mucus, serum and liver as well as catalase activity in serum and liver were decreased significantly (p plant-based diet, whereas the HSP70 mRNA expression decreased significantly (p plant-based diet could decrease the anti-stress properties (non-specific immunity, stress resistance and antioxidant ability) and HSP70 mRNA expression in blunt snout breams fingerling. Although all-plant-based diets could not affect the growth performance of blunt snout breams, the application of all-plant-based diet should be discreet in the production practice.

  11. Developmental and hyperthermia-induced expression of the heat shock proteins HSP60 and HSP70 in tissues of the housefly Musca domestica: an in vitro study

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    Sunita Sharma

    2007-01-01

    Full Text Available The expression pattern of two major chaperones, the heat shock proteins (HSPs HSP60 and HSP70 was studied in vitro in tissues of the housefly Musca domestica during larval and adult stages of development to identify their immunological relatives and understand their functional significance in normal cellular activities and during thermal stress. Fluorographs of labeled polypeptides and western blots demonstrated that both HSPs are expressed constitutively and heat-induced in all the larval and adult cell types examined. The pattern of whole tissue immunocytochemical staining using anti-HSP60 and anti-HSP70 antibodies corresponded well with the observations from western blots or fluorographs. In developing oocytes, both constitutive and heat inducible expression of HSP60 were regulated in an oocyte stage-specific manner. In unstressed ovaries the expression of these proteins was less pronounced in early stage oocytes (1st - 8th than at later stages (9th and onward. The heat shock, however, induced both HSP70 and HSP60 to a significantly high level in early stage oocytes (1st-8th as compared to their respective controls. Our findings indicate the involvement of the HSP60 and HSP70 proteins in the development, growth and differentiation of both somatic and germ line tissues. Furthermore, the enhanced co-expression of HSP70 and HSP60 upon heat shock in various larval and adult cell types suggests the possible role of HSP60 in thermoprotection.

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

    Directory of Open Access Journals (Sweden)

    Yoon Jung-Hoon

    2010-06-01

    Full Text Available Abstract Background 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. Methods 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. Results 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 (H2O2, 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. Conclusions 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.

  13. Identification of several cytoplasmic HSP70 genes from the Mediterranean mussel (Mytilus galloprovincialis) and their long-term evolution in Mollusca and Metazoa.

    Science.gov (United States)

    Kourtidis, Antonis; Drosopoulou, Elena; Nikolaidis, Nikolas; Hatzi, Vasiliki I; Chintiroglou, Chariton C; Scouras, Zacharias G

    2006-04-01

    The HSP70 protein family consists one of the most conserved and important systems for cellular homeostasis under both stress and physiological conditions. The genes of this family are poorly studied in Mollusca, which is the second largest metazoan phylum. To study these genes in Mollusca, we have isolated and identified five HSP70 genes from Mytilus galloprovincialis (Mediterranean mussel) and investigated their short-term evolution within Mollusca and their long-term evolution within Metazoa. Both sequence and phylogenetic analyses suggested that the isolated genes belong to the cytoplasmic (CYT) group of the HSP70 genes. Two of these genes probably represent cognates, whereas the remaining probably represent heat-inducible genes. Phylogenetic analysis including several molluscan CYT HSP70s reveals that the cognate genes in two species have very similar sequences and form intraspecies phylogenetic clades, differently from most metazoan cognate genes studied thus far, implying either recent gene duplications or concerted evolution. The M. galloprovincialis heat-inducible genes show intraspecies phylogenetic clustering, which in combination with the higher amino acid than nucleotide identity suggests that both gene conversion and purifying selection should be responsible for their sequence homogenization. Phylogenetic analysis including several metazoan HSP70s suggests that at least two types of CYT genes were present in the common ancestor of vertebrates and invertebrates, the first giving birth to the heat-inducible genes of invertebrates, whereas the other to both the heat-inducible genes of vertebrates and the cognate genes of all metazoans. These analyses also suggest that inducible and cognate genes seem to undergo divergent evolution.

  14. LHCSR Expression under HSP70/RBCS2 Promoter as a Strategy to Increase Productivity in Microalgae.

    Science.gov (United States)

    Perozeni, Federico; Stella, Giulio Rocco; Ballottari, Matteo

    2018-01-05

    Microalgae are unicellular photosynthetic organisms considered as potential alternative sources for biomass, biofuels or high value products. However, limited biomass productivity is commonly experienced in their cultivating system despite their high potential. One of the reasons for this limitation is the high thermal dissipation of the light absorbed by the outer layers of the cultures exposed to high light caused by the activation of a photoprotective mechanism called non-photochemical quenching (NPQ). In the model organism for green algae Chlamydomonas reinhardtii, NPQ is triggered by pigment binding proteins called light-harvesting-complexes-stress-related (LHCSRs), which are over-accumulated in high light. It was recently reported that biomass productivity can be increased both in microalgae and higher plants by properly tuning NPQ induction. In this work increased light use efficiency is reported by introducing in C. reinhardtii a LHCSR3 gene under the control of Heat Shock Protein 70/RUBISCO small chain 2 promoter in a npq4 lhcsr1 background, a mutant strain knockout for all LHCSR genes. This complementation strategy leads to a low expression of LHCSR3, causing a strong reduction of NPQ induction but is still capable of protecting from photodamage at high irradiance, resulting in an improved photosynthetic efficiency and higher biomass accumulation.

  15. Expression patterns and structural modelling of Hsp70 and Hsp90 in a fish-borne zoonotic nematode Anisakis pegreffii

    National Research Council Canada - National Science Library

    Chen, Hui-Yu; Cheng, Yi-Sheng; Shih, Hsiu-Hui

    2015-01-01

    .... Here, we aimed to elucidate the possible roles of Hsp70 and Hsp90 in the life cycle of the parasitic nematode Anisakis, particularly third- and fourth-stage larvae, from cold-blooded fish to warm...

  16. Ste20-like kinase, SLK, activates the heat shock factor 1 - Hsp70 pathway.

    Science.gov (United States)

    Cybulsky, Andrey V; Guillemette, Julie; Papillon, Joan

    2016-09-01

    Expression and activation of SLK increases during renal ischemia-reperfusion injury. When highly expressed, SLK signals via c-Jun N-terminal kinase and p38 to induce apoptosis, and it exacerbates apoptosis induced by ischemia-reperfusion injury. Overexpression of SLK in glomerular epithelial cells (GECs)/podocytes in vivo induces injury and proteinuria. In response to various stresses, cells enhance expression of chaperones or heat shock proteins (e.g. Hsp70), which are involved in the folding and maturation of newly synthesized proteins, and can refold denatured or misfolded proteins. We address the interaction of SLK with the heat shock factor 1 (HSF1)-Hsp70 pathway. Increased expression of SLK in GECs (following transfection) induced HSF1 transcriptional activity. Moreover, HSF1 transcriptional activity was increased by in vitro ischemia-reperfusion injury (chemical anoxia/recovery) and heat shock, and in both instances was amplified further by SLK overexpression. HSF1 binds to promoters of target genes, such as Hsp70 and induces their transcription. By analogy to HSF1, SLK stimulated Hsp70 expression. Hsp70 was also enhanced by anoxia/recovery and was further amplified by SLK overexpression. Induction of HSF1 and Hsp70 was dependent on the kinase activity of SLK, and was mediated via polo-like kinase-1. Transfection of constitutively active HSF1 enhanced Hsp70 expression and inhibited SLK-induced apoptosis. Conversely, the proapoptotic action of SLK was augmented by HSF1 shRNA, or the Hsp70 inhibitor, pifithrin-μ. In conclusion, increased expression/activity of SLK activates the HSF1-Hsp70 pathway. Hsp70 attenuates the primary proapoptotic effect of SLK. Modulation of chaperone expression may potentially be harnessed as cytoprotective therapy in renal cell injury. Copyright © 2016 Elsevier B.V. All rights reserved.

  17. Functional analysis of the Hikeshi-like protein and its interaction with HSP70 in Arabidopsis

    Energy Technology Data Exchange (ETDEWEB)

    Koizumi, Shinya; Ohama, Naohiko; Mizoi, Junya [Laboratory of Plant Molecular Physiology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657 (Japan); Shinozaki, Kazuo [RIKEN Plant Science Center, 1-7-22 Suehiro-cho, Tsurumi, Yokohama, Kanagawa 230-0045 (Japan); Yamaguchi-Shinozaki, Kazuko, E-mail: akys@mail.ecc.u-tokyo.ac.jp [Laboratory of Plant Molecular Physiology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657 (Japan)

    2014-07-18

    Highlights: • HKL, a Hikeshi homologous gene is identified in Arabidopsis. • HKL interacts with two HSP70 isoforms and regulates the subcellular localization of HSC70-1. • The two HSP70 translocate into nucleus in response to heat stress. • Overexpression of HKL confers thermotolerance in transgenic plants. - Abstract: Heat shock proteins (HSPs) refold damaged proteins and are an essential component of the heat shock response. Previously, the 70 kDa heat shock protein (HSP70) has been reported to translocate into the nucleus in a heat-dependent manner in many organisms. In humans, the heat-induced translocation of HSP70 requires the nuclear carrier protein Hikeshi. In the Arabidopsis genome, only one gene encodes a protein with high homology to Hikeshi, and we named this homolog Hikeshi-like (HKL) protein. In this study, we show that two Arabidopsis HSP70 isoforms accumulate in the nucleus in response to heat shock and that HKL interacts with these HSP70s. Our histochemical analysis revealed that HKL is predominantly expressed in meristematic tissues, suggesting the potential importance of HKL during cell division in Arabidopsis. In addition, we show that HKL regulates HSP70 localization, and HKL overexpression conferred thermotolerance to transgenic Arabidopsis plants. Our results suggest that HKL plays a positive role in the thermotolerance of Arabidopsis plants and cooperatively interacts with HSP70.

  18. Cry1Ab treatment has no effects on viability of cultured porcine intestinal cells, but triggers Hsp70 expression.

    Directory of Open Access Journals (Sweden)

    Angelika Bondzio

    Full Text Available In vitro testing can contribute to reduce the risk that the use of genetically modified (GM crops and their proteins show unintended toxic effects. Here we introduce a porcine intestinal cell culture (IPEC-J2 as appropriate in vitro model and tested the possible toxic potential of Cry1Ab protein, commonly expressed in GM-maize. For comprehensive risk assessment we used WST-1 conversion and ATP content as metabolic markers for proliferation, lactate dehydrogenase release as indicator for cells with compromised membrane and transepithelial electrical resistance as parameter indicating membrane barrier function. The results were compared to the effects of valinomycin, a potassium ionophore, known to induce cytotoxic effects in most mammalian cell types. Whereas no toxicity was observed after Cry1Ab treatment, valinomycin induced a decrease in IPEC-J2 viability. This was confirmed by dynamic monitoring of cellular responses. Additionally, two dimensional differential in-gel electrophoresis was performed. Only three proteins were differentially expressed. The functions of these proteins were associated with responses to stress. The up-regulation of heat shock protein Hsp70 was verified by Western blotting as well as by enzyme-linked immunosorbent assay and may be related to a protective function. These findings suggest that the combination of in vitro testing and proteomic analysis may serve as a promising tool for mechanism based safety assessment.

  19. Genetics of human longevity with emphasis on the relevance of HSP70 as candidate genes

    DEFF Research Database (Denmark)

    Singh, Ripudaman; Kølvrå, Steen; Rattan, Suresh I S

    2007-01-01

    of an appropriate study design and methodology. Since aging is characterized by a progressive accumulation of molecular damage and an attenuation of the cellular defense mechanisms, the focus of studies on human longevity association with genes has now shifted to the pathways of cellular maintenance and repair...

  20. Cytosine DNA methylation at promoter of non LTR retrotransposon and heat shock protein gene (HSP70) of Entamoeba histolytica and lack of correlation with transcription status.

    Science.gov (United States)

    Agrahari, Mridula; Gaurav, Amit Kumar; Bhattacharya, Alok; Bhattacharya, Sudha

    2017-03-01

    Non LTR retrotransposons (EhLINEs and EhSINEs) occupy 11% of the Entamoeba histolytica genome. Since promoter DNA methylation at cytosines has been correlated with transcriptional silencing of transposable elements in model organisms we checked whether this was the case in EhLINE1. We located promoter activity in a 841bp fragment at 5'-end of this element by luciferase reporter assay. From RNAseq and RT-PCR analyses we selected a transcriptionally active and silent copy to study cytosine DNA methylation of the promoter region by bisulfite sequencing. None of the cytosines were methylated in either copy. Further, we looked at methylation status of a few selected cytosines in all 5'-intact EhLINE1 copies by single nucleotide incorporation opposite cytosine in bisulfite-treated DNA, where dGTP would be incorporated if the cytosine was methylated. Again we did not find evidence of cytosine methylation, indicating that expression status of this element was not correlated with promoter DNA methylation. To test for any role of cytosine methylation in transcriptional regulation of the E. histolytica Hsp70 gene in which the promoter is fully methylated under normal growth conditions, we checked methylation status and found that the promoter remained fully methylated during heat-shock as well, although transcription was greatly enhanced by heat-shock, showing that cytosine methylation is not a repressive mark for EhHsp70. Our data present direct evidence that promoter methylation, a common mode of transposon silencing, is unlikely to be involved in transcriptional regulation of EhLINE1, and reinforce the conclusion that promoter DNA methylation may not be a major contributor to transcriptional regulation in E. histolytica. Copyright © 2017 Elsevier B.V. All rights reserved.

  1. Sequence analysis of the 3’-untranslated region of HSP70 (type I genes in the genus Leishmania: its usefulness as a molecular marker for species identification

    Directory of Open Access Journals (Sweden)

    Requena Jose M

    2012-04-01

    Full Text Available Abstract Background The Leishmaniases are a group of clinically diverse diseases caused by parasites of the genus Leishmania. To distinguish between species is crucial for correct diagnosis and prognosis as well as for treatment decisions. Recently, sequencing of the HSP70 coding region has been applied in phylogenetic studies and for identifying of Leishmania species with excellent results. Methods In the present study, we analyzed the 3’-untranslated region (UTR of Leishmania HSP70-type I gene from 24 strains representing eleven Leishmania species in the belief that this non-coding region would have a better discriminatory capacity for species typing than coding regions. Results It was observed that there was a remarkable degree of sequence conservation in this region, even between species of the subgenus Leishmania and Viannia. In addition, the presence of many microsatellites was a common feature of the 3´-UTR of HSP70-I genes in the Leishmania genus. Finally, we constructed dendrograms based on global sequence alignments of the analyzed Leishmania species and strains, the results indicated that this particular region of HSP70 genes might be useful for species (or species complex typing, improving for particular species the discrimination capacity of phylogenetic trees based on HSP70 coding sequences. Given the large size variation of the analyzed region between the Leishmania and Viannia subgenera, direct visualization of the PCR amplification product would allow discrimination between subgenera, and a HaeIII-PCR-RFLP analysis might be used for differentiating some species within each subgenera. Conclusions Sequence and phylogenetic analyses indicated that this region, which is readily amplified using a single pair of primers from both Old and New World Leishmania species, might be useful as a molecular marker for species discrimination.

  2. Diversity of cytosolic HSP70 Heat Shock Protein from decapods and their phylogenetic placement within Arthropoda.

    Science.gov (United States)

    Baringou, Stephane; Rouault, Jacques-Deric; Koken, Marcel; Hardivillier, Yann; Hurtado, Luis; Leignel, Vincent

    2016-10-10

    The 70kDa heat shock proteins (HSP70) are considered the most conserved members of the HSP family. These proteins are primordial to the cell, because of their implications in many cellular pathways (e. g., development, immunity) and also because they minimize the effects of multiple stresses (e. g., temperature, pollutants, salinity, radiations). In the cytosol, two ubiquitous HSP70s with either a constitutive (HSC70) or an inducible (HSP70) expression pattern are found in all metazoan species, encoded by 5 or 6 genes (Drosophila melanogaster or yeast and human respectively). The cytosolic HSP70 protein family is considered a major actor in environmental adaptation, and widely used in ecology as an important biomarker of environmental stress. Nevertheless, the diversity of cytosolic HSP70 remains unclear amongst the Athropoda phylum, especially within decapods. Using 122 new and 311 available sequences, we carried out analyses of the overall cytosolic HSP70 diversity in arthropods (with a focus on decapods) and inferred molecular phylogenies. Overall structural and phylogenetic analyses showed a surprisingly high diversity in cytosolic HSP70 and revealed the existence of several unrecognised groups. All crustacean HSP70 sequences present signature motifs and molecular weights characteristic of non-organellar HSP70, with multiple specific substitutions in the protein sequence. The cytosolic HSP70 family in arthropods appears to be constituted of at least three distinct groups (annotated as A, B and C), which comprise several subdivisions, including both constitutive and inducible forms. Group A is constituted by several classes of Arthropods, while group B and C seem to be specific to Malacostraca and Hexapoda/Chelicerata, respectively. The HSP70 organization appeared much more complex than previously suggested, and far beyond a simple differentiation according to their expression pattern (HSC70 versus HSP70). This study proposes a new classification of cytosolic

  3. Phosphatidylcholine-specific phospholipase C/heat shock protein 70 (Hsp70)/transcription factor B-cell translocation gene 2 signaling in rat bone marrow stromal cell differentiation to cholinergic neuron-like cells.

    Science.gov (United States)

    Shao, Jing; Sun, Chunhui; Su, Le; Zhao, Jing; Zhang, Shangli; Miao, Junying

    2012-12-01

    Although bone marrow stromal cells (BMSCs) can differentiate into neuron-like cells, the mechanisms underlying neuronal differentiation are not well understood. We recently found that inhibition of phosphatidylcholine-specific phospholipase C (PC-PLC) by its inhibitor D609 promoted BMSCs' differentiation into cholinergic neuron-like cells. Using the effective small molecule D609 and gene microarray technology, we investigated the change of gene expression profile to identify key mediators involved in the neuronal differentiation. We selected heat shock protein 70 (Hsp70) and transcription factor B-cell translocation gene 2 (Btg2) that were maximally up-regulated for further study. We found that functional suppression of Hsp70 blocked D609-induced increase of Btg2 expression and cholinergic neuronal differentiation of BMSCs. These results demonstrated that Hsp70 was the pivotal factor in PC-PLC-medicated neuronal differentiation of BMSCs, and Btg2 might be its downstream target. Our findings provide new clues for controlling BMSCs' differentiation into cholinergic neuron-like cells and provide a putative strategy for neurodegenerative diseases therapies. Crown Copyright © 2012. Published by Elsevier Ltd. All rights reserved.

  4. Impact of metals on histopathology and expression of HSP 70 in different tissues of Milk fish (Chanos chanos) of Kaattuppalli Island, South East Coast, India.

    Science.gov (United States)

    Rajeshkumar, Sivakumar; Munuswamy, Natesan

    2011-04-01

    Histological and Immunohistochemical studies were carried out to document the possible impact of heavy metal contamination in different tissues of Chanos chanos. Heavy metals such as Cu, Pb, Zn, Cd, Mn and Fe were predominant in water, sediment and biota of Kaattuppalli Island and varied significantly between two different sites. Histological changes such as swelling of muscle fiber and break down of muscle bundles were noted in the muscle. Similarly gill filament cell proliferation, increase in intercellular spaces and primary and secondary lamellar epithelium were evident in gills. The hepatocytes showed damage of central vein and rupture of irregular hepatic plate with more number of vacuoles in the fish collected from polluted site compared to that from the less polluted site. The impact of pollution was also assessed in different tissues by immunohistochemistry using primary antibody (mouse monoclonal HSP70 antibody 1:2000) and secondary antibody (HRP conjugated antibody) for expression of stress protein. Immunostaining analysis showed expression of HSP70 with high intensity in the tissues of fish collected from polluted site compared to less polluted sites. Further, HSP70 positive cells were analyzed from six locations per fish tissue section. One-way analysis of variance (ANOVA) followed by least significant difference (LSD) was used to check if the expression was significant. Results indicate that the values are statistically significant at the two different sampling sites (P<0.05). Copyright © 2011 Elsevier Ltd. All rights reserved.

  5. Examination the expression pattern of HSP70 heat shock protein in chicken PGCs and developing genital ridge

    OpenAIRE

    Mahek Anand; Roland Tóth; Alayu Kidane; Alexandra Nagy; Bence Lázár; Eszter Patakiné Várkonyi; Krisztina Liptói; Elen Gocza

    2016-01-01

    Chicken Primordial Germ cells (PGCs) are emerging pioneers in the field of applied embryology and stem cell technology. Now-a-days transgenic chickens are promising models to study human disease pathophysiology and drug designing. However, most of the molecular mechanism, which govern the stemness and pluripotency of chicken PGCs, not known in details. Recent studies have indicated the role of HSP70 in early embryonic development in many vertebrate species. Exposure of chicken to heat...

  6. A Rhodiola rosea root extract protects skeletal muscle cells against chemically induced oxidative stress by modulating heat shock protein 70 (HSP70) expression.

    Science.gov (United States)

    Hernández-Santana, Aaron; Pérez-López, Verónica; Zubeldia, Jose María; Jiménez-del-Rio, Miguel

    2014-04-01

    Rhodiola rosea is a perennial plant in the Crassulaceae family, recently postulated to exert its adaptogenic functions partially by modulating the expression of molecular factors such as heat shock proteins (HSP). The aim of this study was to analyze the efficacy of a Rhodiola rosea extract (Rhodiolife) in protecting murine skeletal muscle cells (C2 C12 myotubes) from chemically induced oxidative stress and to establish whether modulation of HSP70 expression is observed. C2 C12 cells treated with Rhodiolife did not experience any loss of viability (p > 0.05) at concentrations of 1-100 µg/mL for up to 24 h. In control cultures, viability decreased 25% following exposure to 2 mM H2 O2 (1 h). However, no significant decrease in viability in cells pre-treated with extract at concentrations as low as 1 µg/mL was observed. HSP70 mRNA levels were up-regulated two-fold in cell cultures treated with Rhodiolife (10 µg/mL), and expression was further enhanced by exposure to H2 O2 (six-fold, p < 0.05). HSP70 protein levels were maintained in pre-treated cell cultures compared to controls but was significantly lower (-50%) in cells lacking treatment exposed to H2 O2 . The present results indicate that Rhodiolife protects C2 C12 myotubes against peroxide-induced oxidative stress through the modulation of the molecular chaperone HSP70. Copyright © 2013 John Wiley & Sons, Ltd.

  7. Hsp70 architecture: the formation of novel polymeric structures of Hsp70.1 and Hsc70 after proteotoxic stress.

    Science.gov (United States)

    Steel, Rohan; Cross, Ryan S; Ellis, Sarah L; Anderson, Robin L

    2012-01-01

    Heat induces Hsp70.1 (HSPA1) and Hsc70 (HSPA8) to form complex detergent insoluble cytoplasmic and nuclear structures that are distinct from the cytoskeleton and internal cell membranes. These novel structures have not been observed by earlier immunofluorescence studies as they are obscured by the abundance of soluble Hsp70.1/Hsc70 present in cells. While resistant to detergents, these Hsp70 structures display complex intracellular dynamics and are efficiently disaggregated by ATP, indicating that this pool of Hsp70.1/Hsc70 retains native function and regulation. Hsp70.1 promotes the repair of proteotoxic damage and cell survival after stress. In heated fibroblasts expressing Hsp70.1, Hsp70.1 and Hsc70 complexes are efficiently disaggregated before the cells undergo-heat induced apoptosis. In the absence of Hsp70.1, fibroblasts have increased rates of heat-induced apoptosis and maintain stable insoluble Hsc70 structures. The differences in the intracellular distribution of Hsp70.1 and Hsc70, combined with the ability of Hsp70.1, but not Hsc70, to promote the disaggregation of insoluble Hsp70.1/Hsc70 complexes, indicate that these two closely related proteins perform distinctly different cellular functions in heated cells.

  8. Genetic variation and recombination of RdRp and HSP 70h genes of Citrus tristeza virus isolates from orange trees showing symptoms of citrus sudden death disease.

    Science.gov (United States)

    Gomes, Clarissa P C; Nagata, Tatsuya; de Jesus, Waldir C; Neto, Carlos R Borges; Pappas, Georgios J; Martin, Darren P

    2008-01-16

    Citrus sudden death (CSD), a disease that rapidly kills orange trees, is an emerging threat to the Brazilian citrus industry. Although the causal agent of CSD has not been definitively determined, based on the disease's distribution and symptomatology it is suspected that the agent may be a new strain of Citrus tristeza virus (CTV). CTV genetic variation was therefore assessed in two Brazilian orange trees displaying CSD symptoms and a third with more conventional CTV symptoms. A total of 286 RNA-dependent-RNA polymerase (RdRp) and 284 heat shock protein 70 homolog (HSP70h) gene fragments were determined for CTV variants infecting the three trees. It was discovered that, despite differences in symptomatology, the trees were all apparently coinfected with similar populations of divergent CTV variants. While mixed CTV infections are common, the genetic distance between the most divergent population members observed (24.1% for RdRp and 11.0% for HSP70h) was far greater than that in previously described mixed infections. Recombinants of five distinct RdRp lineages and three distinct HSP70h lineages were easily detectable but respectively accounted for only 5.9 and 11.9% of the RdRp and HSP70h gene fragments analysed and there was no evidence of an association between particular recombinant mosaics and CSD. Also, comparisons of CTV population structures indicated that the two most similar CTV populations were those of one of the trees with CSD and the tree without CSD. We suggest that if CTV is the causal agent of CSD, it is most likely a subtle feature of population structures within mixed infections and not merely the presence (or absence) of a single CTV variant within these populations that triggers the disease.

  9. Genetic variation and recombination of RdRp and HSP 70h genes of Citrus tristeza virus isolates from orange trees showing symptoms of citrus sudden death disease

    Directory of Open Access Journals (Sweden)

    Pappas Georgios J

    2008-01-01

    Full Text Available Abstract Background Citrus sudden death (CSD, a disease that rapidly kills orange trees, is an emerging threat to the Brazilian citrus industry. Although the causal agent of CSD has not been definitively determined, based on the disease's distribution and symptomatology it is suspected that the agent may be a new strain of Citrus tristeza virus (CTV. CTV genetic variation was therefore assessed in two Brazilian orange trees displaying CSD symptoms and a third with more conventional CTV symptoms. Results A total of 286 RNA-dependent-RNA polymerase (RdRp and 284 heat shock protein 70 homolog (HSP70h gene fragments were determined for CTV variants infecting the three trees. It was discovered that, despite differences in symptomatology, the trees were all apparently coinfected with similar populations of divergent CTV variants. While mixed CTV infections are common, the genetic distance between the most divergent population members observed (24.1% for RdRp and 11.0% for HSP70h was far greater than that in previously described mixed infections. Recombinants of five distinct RdRp lineages and three distinct HSP70h lineages were easily detectable but respectively accounted for only 5.9 and 11.9% of the RdRp and HSP70h gene fragments analysed and there was no evidence of an association between particular recombinant mosaics and CSD. Also, comparisons of CTV population structures indicated that the two most similar CTV populations were those of one of the trees with CSD and the tree without CSD. Conclusion We suggest that if CTV is the causal agent of CSD, it is most likely a subtle feature of population structures within mixed infections and not merely the presence (or absence of a single CTV variant within these populations that triggers the disease.

  10. A fraction from Petiveria alliacea induces apoptosis via a mitochondria-dependent pathway and regulates HSP70 expression

    OpenAIRE

    Cifuentes, Maria Claudia; Grupo de Inmunobiología y Biología Celular, Facultad de Ciencias, Pontificia Universidad Javeriana, Bogotá; Castañeda, Diana Mercedes; Grupo de Inmunobiología y Biología Celular, Facultad de Ciencias, Pontificia Universidad Javeriana, Bogotá; Urueña, Claudia Patricia; Grupo de Inmunobiología y Biología Celular, Facultad de Ciencias, Pontificia Universidad Javeriana, Bogotá; Fiorentino, Susana; Grupo de Inmunobiología y Biología Celular, Facultad de Ciencias, Pontificia Universidad Javeriana, Bogotá

    2009-01-01

    Una fracción de Petiveria alliacea induce apoptosis dependiente de la mitocondria y regula la expresión de la proteína HSP70. Objetivo: Evaluar la actividad biológica in vitro de extractos de Petiveria alliacea utilizando líneas celulares tumorales. Materiales y Métodos: Las fracciones de P.alliacea fueron preparadas mediante un protocolo de purificación biodirigido. La actividad biológica fue caracterizada utilizando dos líneas celulares tumorales de origen humano, donde se analizaron los ca...

  11. Heat adaptation from regular hot water immersion decreases proinflammatory responses, HSP70 expression, and physical heat stress.

    Science.gov (United States)

    Yang, Fwu-Lin; Lee, Chia-Chi; Subeq, Yi-Maun; Lee, Chung-Jen; Ke, Chun-Yen; Lee, Ru-Ping

    2017-10-01

    Hot-water immersion (HWI) is a type of thermal therapy for treating various diseases. In our study, the physiological responses to occasional and regular HWI have been explored. The rats were divided into a control group, occasional group (1D), and regular group (7D). The 1D and 7D groups received 42°C during 15mins HWI for 1 and 7 days, respectively. The blood samples were collected for proinflammatory cytokines examinations, the heart, liver and kidney were excised for subsequent IHC analysis to measure the level of heat shock protein 70 (HSP70). The results revealed that the body temperature increased significantly during HWI on Day 3 and significantly declined on Days 6 and 7. For the 7D group, body weight, heart rate, hematocrit, platelet, osmolarity, and lactate level were lower than those in the 1D group. Furthermore, the levels of granulocyte counts, tumor necrosis factor-α, and interleukin-6 were lower in the 7D group than in the 1D group. The induction of HSP70 in the 1D group was higher than in the other groups. Physiological responses to occasional HWI are disadvantageous because of heat stress. However, adaptation to heat from regular HWI resulted in decreased proinflammatory responses and physical heat stress. Copyright © 2017 Elsevier Ltd. All rights reserved.

  12. Organization and evolution of hsp70 clusters strikingly differ in two species of Stratiomyidae (Diptera inhabiting thermally contrasting environments

    Directory of Open Access Journals (Sweden)

    Bettencourt Brian R

    2011-03-01

    Full Text Available Abstract Background Previously, we described the heat shock response in dipteran species belonging to the family Stratiomyidae that develop in thermally and chemically contrasting habitats including highly aggressive ones. Although all species studied exhibit high constitutive levels of Hsp70 accompanied by exceptionally high thermotolerance, we also detected characteristic interspecies differences in heat shock protein (Hsp expression and survival after severe heat shock. Here, we analyzed genomic libraries from two Stratiomyidae species from thermally and chemically contrasting habitats and determined the structure and organization of their hsp70 clusters. Results Although the genomes of both species contain similar numbers of hsp70 genes, the spatial distribution of hsp70 copies differs characteristically. In a population of the eurytopic species Stratiomys singularior, which exists in thermally variable and chemically aggressive (hypersaline conditions, the hsp70 copies form a tight cluster with approximately equal intergenic distances. In contrast, in a population of the stenotopic Oxycera pardalina that dwells in a stable cold spring, we did not find hsp70 copies in tandem orientation. In this species, the distance between individual hsp70 copies in the genome is very large, if they are linked at all. In O. pardalina we detected the hsp68 gene located next to a hsp70 copy in tandem orientation. Although the hsp70 coding sequences of S. singularior are highly homogenized via conversion, the structure and general arrangement of the hsp70 clusters are highly polymorphic, including gross aberrations, various deletions in intergenic regions, and insertion of incomplete Mariner transposons in close vicinity to the 3'-UTRs. Conclusions The hsp70 gene families in S. singularior and O. pardalina evolved quite differently from one another. We demonstrated clear evidence of homogenizing gene conversion in the S. singularior hsp70 genes, which form

  13. Induction of hsp70, hsp90, and catalase activity in planarian Dugesia japonica exposed to cadmium.

    Science.gov (United States)

    Zhang, Xiufang; Mo, Yehua; Zhou, Luming; Wang, Yinan; Wang, Zhongchen; Zhao, Bosheng

    2016-08-01

    The hsp70 and hsp90 expression patterns and catalase (CAT) activity in the freshwater planaria Dugesia japonica exposed to cadmium (Cd) under laboratory conditions were investigated. Planaria were exposed to a range of Cd concentrations (0-150 μg Cd/L) for 24 h. The expression levels of hsp70 and hsp90 were determined by relative quantitative real-time polymerase chain reaction. Within the overall dose range in the experiment, the expression level of hsp70 and the activity of CAT in D. japonica were altered significantly. Hsp70 was induced in D. japonica upon Cd exposure concentrations as low as 9.375 μg Cd/L. No significant effect on the expression level of hsp90 was observed. Our findings demonstrated that stress gene hsp70, but not hsp90, was responsive to Cd contamination in D. japonica CAT activity was significantly induced at concentrations of 18.75, 37.5, and 75 μg Cd/L after 24-h exposure. We recommend that the use of hsp70 as a biomarker should be complemented by evidence of changes in other parameters, such as CAT activity, in D. japonica. © The Author(s) 2014.

  14. Phylogenetic analysis of HSP70 and cyt b gene sequences for Chinese Leishmania isolates and ultrastructural characteristics of Chinese Leishmania sp.

    Science.gov (United States)

    Yuan, Dongmei; Qin, Hanxiao; Zhang, Jianguo; Liao, Lin; Chen, Qiwei; Chen, Dali; Chen, Jianping

    2017-02-01

    Leishmaniasis is a worldwide epidemic disease caused by the genus Leishmania, which is still endemic in the west and northwest areas of China. Some viewpoints of the traditional taxonomy of Chinese Leishmania have been challenged by recent phylogenetic researches based on different molecular markers. However, the taxonomic positions and phylogenetic relationships of Chinese Leishmania isolates remain controversial, which need for more data and further analysis. In this study, the heat shock protein 70 (HSP70) gene and cytochrome b (cyt b) gene were used for phylogenetic analysis of Chinese Leishmania isolates from patients, dogs, gerbils, and sand flies in different geographic origins. Besides, for the interesting Leishmania sp. in China, the ultrastructure of three Chinese Leishmania sp. strains (MHOM/CN/90/SC10H2, SD, GL) were observed by transmission electron microscopy. Bayesian trees from HSP70 and cyt b congruently indicated that the 14 Chinese Leishmania isolates belong to three Leishmania species including L. donovani complex, L. gerbilli, and L. (Sauroleishmania) sp. Their identity further confirmed that the undescribed Leishmania species causing visceral Leishmaniasis (VL) in China is closely related to L. tarentolae. The phylogenetic results from HSP70 also suggested the classification of subspecies within L. donovani complex: KXG-918, KXG-927, KXG-Liu, KXG-Xu, 9044, SC6, and KXG-65 belong to L. donovani; Cy, WenChuan, and 801 were proposed to be L. infantum. Through transmission electron microscopy, unexpectedly, the Golgi apparatus were not observed in SC10H2, SD, and GL, which was similar to previous reports of reptilian Leishmania. The statistical analysis of microtubule counts separated SC10H2, SD, and GL as one group from any other reference strain (L. donovani MHOM/IN/80/DD8; L. tropica MHOM/SU/74/K27; L. gerbilli MRHO/CN/60/GERBILLI). The ultrastructural characteristics of Leishmania sp. partly lend support to the phylogenetic inference that

  15. thermal stress and Hsp70 as selective agents

    Indian Academy of Sciences (India)

    Madhu Sudhan

    Introduction. Stress is as ancient as life itself, and shapes the evolution of genes, genomes, and organisms – a fact attested to by the kingdom-spanning conservation of heat shock proteins. (Hsps) and Hsp70 in particular. Hsp70 is the principal eukaryotic stress-inducible heat shock protein and promotes the ability of ...

  16. HSP70 induced by Hantavirus infection interacts with viral nucleocapsid protein and its overexpression suppresses virus infection in Vero E6 cells.

    Science.gov (United States)

    Yu, Lu; Ye, Ling; Zhao, Rong; Liu, Yan Fang; Yang, Shou Jing

    2009-07-15

    Hantavirus (HTV) infection is known to induce innate cellular response, a more specified cellular response in the host cells. However, whether it stimulates synthesis of stress proteins, particularly associations of viral proteins, is entirely unknown. The primary focus of this research is using Vero E6 cells infected with Hantaan 76-118 (HTNV) as an in vitro infection model to examine the individual contribution of HTV infection to heat shock response. This study shows that HTNV infection rapidly induced HSP70 expression in Vero E6 cells, which underwent a nucleo-cytoplasmic shuttle that lasted for more than 3 d. The increased HSP70 was preceded by induction of HSP70 mRNA. The physical association of HSP70 with viral nucleocapsid protein (NP) in infected cells was demonstrated by co-localization and immunoprecipation. Vero E6 cells that constitutively overexpress HSP70 after stable transfection with HSP70 gene, when infected with HTNV, showed selectively reduced NP synthesis. These findings suggest HSP70 is actively involved in the control of the expression level of viral structural proteins and possibly involved in virus assembly by binding of NP to HSP70. Overexpression of HSP70 does not favor viral propagation.

  17. Combined effects of thermal stress and Cd on lysosomal biomarkers and transcription of genes encoding lysosomal enzymes and HSP70 in mussels, Mytilus galloprovincialis

    Energy Technology Data Exchange (ETDEWEB)

    Izagirre, Urtzi; Errasti, Aitzpea; Bilbao, Eider; Múgica, María; Marigómez, Ionan, E-mail: ionan.marigomez@ehu.es

    2014-04-01

    Highlights: • Thermal stress and Cd caused lysosomal enlargement and membrane destabilisation. • hex, gusb and ctsl but not hsp70 were up-regulated at elevated temperature but down-regulated by Cd. • Thermal stress influenced lysosomal responses to Cd exposure. • The presence of Cd jeopardised responsiveness against thermal stress. - Abstract: In estuaries and coastal areas, intertidal organisms may be subject to thermal stress resulting from global warming, together with pollution. In the present study, the combined effects of thermal stress and exposure to Cd were investigated in the endo-lysosomal system of digestive cells in mussels, Mytilus galloprovincialis. Mussels were maintained for 24 h at 18 °C and 26 °C seawater temperature in absence and presence of 50 μg Cd/L seawater. Cadmium accumulation in digestive gland tissue, lysosomal structural changes and membrane stability were determined. Semi-quantitative PCR was applied to reveal the changes elicited by the different experimental conditions in hexosaminidase (hex), β-glucuronidase (gusb), cathepsin L (ctsl) and heat shock protein 70 (hsp70) gene transcription levels. Thermal stress provoked lysosomal enlargement whilst Cd-exposure led to fusion of lysosomes. Both thermal stress and Cd-exposure caused lysosomal membrane destabilisation. hex, gusb and ctsl genes but not hsp70 gene were transcriptionally up-regulated as a result of thermal stress. In contrast, all the studied genes were transcriptionally down-regulated in response to Cd-exposure. Cd bioaccumulation was comparable at 18 °C and 26 °C seawater temperatures but interactions between thermal stress and Cd-exposure were remarkable both in lysosomal biomarkers and in gene transcription. hex, gusb and ctsl genes, reacted to elevated temperature in absence of Cd but not in Cd-exposed mussels. Therefore, thermal stress resulting from global warming might influence the use and interpretation of lysosomal biomarkers in marine pollution

  18. Hsp70 expression and metabolite composition in response to short-term thermal changes in Folsomia candida (Collembola)

    DEFF Research Database (Denmark)

    Waagner, Dorthe; Heckmann, Lars-Henrik; Malmendal, Anders

    2010-01-01

    protein Hsp70 was assessed at the mRNA level using real time quantitative polymerase chain reaction (QPCR). Hsp70 was rapidly induced and significantly increased by the temperature increase. The relative concentrations of low molecular weight metabolites were analysed in F. candida using nuclear magnetic...

  19. Mutant HSP70 Reverses Autoimmune Depigmentation in Vitiligo

    Science.gov (United States)

    Mosenson, Jeffrey A.; Zloza, Andrew; Nieland, John D.; Garrett-Mayer, Elizabeth; Eby, Jonathan M.; Huelsmann, Erica J.; Kumar, Previn; Denman, Cecele J.; Lacek, Andrew T.; Kohlhapp, Frederick J.; Alamiri, Ahmad; Hughes, Tasha; Bines, Steven D.; Kaufman, Howard L.; Overbeck, Andreas; Mehrotra, Shikhar; Hernandez, Claudia; Nishimura, Michael I.; Guevara-Patino, Jose A.; Le Poole, I. Caroline

    2013-01-01

    Vitiligo is an autoimmune disease characterized by destruction of melanocytes, leaving 0.5% of the population with progressive depigmentation. Current treatments offer limited efficacy. We report that modified inducible heat shock protein 70 (HSP70i) prevents T cell–mediated depigmentation. HSP70i is the molecular link between stress and the resultant immune response. We previously showed that HSP70i induces an inflammatory dendritic cell (DC) phenotype and is necessary for depigmentation in vitiligo mouse models. Here, we observed a similar DC inflammatory phenotype in vitiligo patients. In a mouse model of depigmentation, DNA vaccination with a melanocyte antigen and the carboxyl terminus of HSP70i was sufficient to drive autoimmunity. Mutational analysis of the HSP70i substrate-binding domain established the peptide QPGVLIQVYEG as invaluable for DC activation, and mutant HSP70i could not induce depigmentation. Moreover, mutant HSP70iQ435A bound human DCs and reduced their activation, as well as induced a shift from inflammatory to tolerogenic DCs in mice. HSP70iQ435A-encoding DNA applied months before spontaneous depigmentation prevented vitiligo in mice expressing a transgenic, melanocyte-reactive T cell receptor. Furthermore, use of HSP70iQ435A therapeutically in a different, rapidly depigmenting model after loss of differentiated melanocytes resulted in 76% recovery of pigmentation. Treatment also prevented relevant T cells from populating mouse skin. In addition, ex vivo treatment of human skin averted the disease-related shift from quiescent to effector T cell phenotype. Thus, HSP70iQ435A DNA delivery may offer potent treatment opportunities for vitiligo. PMID:23447019

  20. Hsp70 expression and free radical release after exposure to non-thermal radio-frequency electromagnetic fields and ultrafine particles in human Mono Mac 6 cells.

    Science.gov (United States)

    Simkó, M; Hartwig, C; Lantow, M; Lupke, M; Mattsson, M-O; Rahman, Q; Rollwitz, J

    2006-02-08

    The contemporary urban environment has become increasingly complex in its composition, leading to discussions regarding possible novel health effects. Two factors that recently have received considerable attention are ultrafine particles (UFP; spectrum. Several studies have shown biological effects of both these exposures in various cell systems. Here we investigate if exposure to UFP (12-14 nm, 100 microg/ml) and RF-electromagnetic fields (EMF; 2 W/kg specific absorption rate (SAR); continuous wave (CW) or modulated (217Hz or GSM-nonDTX)), alone or in combination influences levels of the superoxide radical anion or the stress protein heat-shock protein (Hsp70) in the human monocyte cell line Mono Mac 6. Heat treatment (42-43 degrees C, 1h) was used as positive control for both stress reaction and for heat development in the RF exposure setup. Our results clearly show that Mono Mac 6 cells are capable to internalise UFP, and that this phagocytic activity is connected to an increased release of free radicals. This increase (40-45% above negative control) is stronger than the effect of heat treatment. On the other hand, none of the employed RF exposures showed any effects on free radical levels. Co-exposure of RF and UFP did not potentiate the UFP effect either. Our investigations showed a significantly increased Hsp70 expression level by heat treatment in a time-dependent manner, whereas UFP, RF, or UFP+RF were without any effect. Therefore, we conclude that in the investigated Mono Mac 6 cells, RF exposure alone or in combination with UFP cannot influence stress-related responses.

  1. Regulation of Hsp27 and Hsp70 expression in human and mouse skin construct models by caveolae following exposure to the model sulfur mustard vesicant, 2-chloroethyl ethyl sulfide.

    Science.gov (United States)

    Black, Adrienne T; Hayden, Patrick J; Casillas, Robert P; Heck, Diane E; Gerecke, Donald R; Sinko, Patrick J; Laskin, Debra L; Laskin, Jeffrey D

    2011-06-01

    Dermal exposure to the vesicant sulfur mustard causes marked inflammation and tissue damage. Basal keratinocytes appear to be a major target of sulfur mustard. In the present studies, mechanisms mediating skin toxicity were examined using a mouse skin construct model and a full-thickness human skin equivalent (EpiDerm-FT™). In both systems, administration of the model sulfur mustard vesicant, 2-chloroethyl ethyl sulfide (CEES, 100-1000μM) at the air surface induced mRNA and protein expression of heat shock proteins 27 and 70 (Hsp27 and Hsp70). CEES treatment also resulted in increased expression of caveolin-1, the major structural component of caveolae. Immunohistochemistry revealed that Hsp27, Hsp70 and caveolin-1 were localized in basal and suprabasal layers of the epidermis. Caveolin-1 was also detected in fibroblasts in the dermal component of the full thickness human skin equivalent. Western blot analysis of caveolar membrane fractions isolated by sucrose density centrifugation demonstrated that Hsp27 and Hsp70 were localized in caveolae. Treatment of mouse keratinocytes with filipin III or methyl-β-cyclodextrin, which disrupt caveolar structure, markedly suppressed CEES-induced Hsp27 and Hsp70 mRNA and protein expression. CEES treatment is known to activate JNK and p38 MAP kinases; in mouse keratinocytes, inhibition of these enzymes suppressed CEES-induced expression of Hsp27 and Hsp70. These data suggest that MAP kinases regulate Hsp 27 and Hsp70; moreover, caveolae-mediated regulation of heat shock protein expression may be important in the pathophysiology of vesicant-induced skin toxicity. Copyright © 2011 Elsevier Inc. All rights reserved.

  2. Suppressed expression of homotypic multinucleation, extracellular domains of CD172α (SIRP-α) and CD47 (IAP) receptors in TAMs upregulated by Hsp70-peptide complex in Dalton's lymphoma.

    Science.gov (United States)

    Gautam, P K; Acharya, A

    2014-07-01

    CD172α and CD47 are members of glycoprotein expressed on macrophages and various immune cells, promote immune recognition and T cell stimulation that priming phagocytosis of pathogens and apoptotic bodies and malignant cell. Tumour-releasing immunosuppressive factor promotes tumour growth and transforms the tumour resident M1 phenotype of macrophage to M2 phenotype (TAMs) that promotes tumour progression by downregulating the expression of different surface receptor including CD172α and CD47. Recent studies have reported that CD172α and CD47 are involved in the pathogenesis and promote malignancies such as lymphoma, leukaemia, melanoma, lung cancer and multiple myeloma, and their expression varies during infection and malignancies. Autologous Hsp70 is well recognized for its role in activating macrophages leading to enhance production of inflammatory cytokines. It has been observed that Hsp70 derived from normal tissues do not elicit tumour immunity, while Hsp70 preparation from tumour cell was able to elicit tumour immunity. However, the role of exogenous autologous hsp70 on the formation of giant cells is completely unknown. Therefore, in the present study, we sought to investigate the effect of Hsp70-peptide complex on the expression of CD172α and CD47 receptors in normal peritoneal macrophages (NMO) and TAMs. Finding shows that the expression of CD172α and CD47 enhances in TAMs and it reverts back the suppressed function of TAMs into M1 state of immunoregulatory phenotype that promotes tumour regression by enhanced multinucleation and phagocytosis of malignant cells and significantly enhances the homotypic fusion of macrophages and polykaryon formation in vitro by enhancing the expression of SIRPα and IAP. © 2014 John Wiley & Sons Ltd.

  3. Influence of Ginkgo Biloba extract (EGb 761) on expression of IL-1 Beta, IL-6, TNF-alfa, HSP-70, HSF-1 and COX-2 after noise exposure in the rat cochlea.

    Science.gov (United States)

    Dogan, Remzi; Sjostrand, Alev Pektas; Yenıgun, Alper; Karatas, Ersin; Kocyigit, Abdurrahim; Ozturan, Orhan

    2017-10-14

    The objective of this study was to investigate the influence of Ginkgo Biloba in early treatment of noise induced hearing loss on expression of IL-6, IL-1 Beta, TNF-alfa, HSP-70, HSF-1 and COX-2 in the rat cochlea. Thirty two female rats were randomly divided into four groups (Acoustic Trauma, Ginkgo Biloba, Acoustic Trauma+Ginkgo Biloba, Non Treatment). Auditory brainstem response (ABR) was applied in all the groups. At the end of the study, IL-1Beta, IL-6, TNF-alpha, HSP-70, HSF-1 and COX-2 were studied in cochlear tissue with ELISA and Western blot analysis. There were significant increases in ABR values measured at days 1 and 7 compared to baseline values in Group 3. IL-1 Beta, IL-6 and TNF-alpha values were significantly higher in Group 1 than in the other groups. Whereas HSP-70 and HSF-1 values were found to be significantly lower in Group 1 compared to those in Group 2 and Group 3. COX-2 of Group 1 was significantly higher than the other groups. Ginkgo Biloba is helpful in the treatment of noise induced hearing loss and exerts its effect by inhibiting expression of IL-1 Beta, IL-6, TNF-alpha and COX-2 and increasing HSP-70 and HSF-1 values in rat cochlea. Copyright © 2017 Elsevier B.V. All rights reserved.

  4. Tumor imaging and targeting potential of an Hsp70-derived 14-mer peptide.

    Directory of Open Access Journals (Sweden)

    Mathias Gehrmann

    Full Text Available We have previously used a unique mouse monoclonal antibody cmHsp70.1 to demonstrate the selective presence of a membrane-bound form of Hsp70 (memHsp70 on a variety of leukemia cells and on single cell suspensions derived from solid tumors of different entities, but not on non-transformed cells or cells from corresponding 'healthy' tissue. This antibody can be used to image tumors in vivo and target them for antibody-dependent cellular cytotoxicity. Tumor-specific expression of memHsp70 therefore has the potential to be exploited for theranostic purposes. Given the advantages of peptides as imaging and targeting agents, this study assessed whether a 14-mer tumor penetrating peptide (TPP; TKDNNLLGRFELSG, the sequence of which is derived from the oligomerization domain of Hsp70 which is expressed on the cell surface of tumor cells, can also be used for targeting membrane Hsp70 positive (memHsp70+ tumor cells, in vitro.The specificity of carboxy-fluorescein (CF- labeled TPP (TPP to Hsp70 was proven in an Hsp70 knockout mammary tumor cell system. TPP specifically binds to different memHsp70+ mouse and human tumor cell lines and is rapidly taken up via endosomes. Two to four-fold higher levels of CF-labeled TPP were detected in MCF7 (82% memHsp70+ and MDA-MB-231 (75% memHsp70+ cells compared to T47D cells (29% memHsp70+ that exhibit a lower Hsp70 membrane positivity. After 90 min incubation, TPP co-localized with mitochondrial membranes in memHsp70+ tumors. Although there was no evidence that any given vesicle population was specifically localized, fluorophore-labeled cmHsp70.1 antibody and TPP preferentially accumulated in the proximity of the adherent surface of cultured cells. These findings suggest a potential association between membrane Hsp70 expression and cytoskeletal elements that are involved in adherence, the establishment of intercellular synapses and/or membrane reorganization.This study demonstrates the specific binding and rapid

  5. Investigating the Chaperone Properties of a Novel Heat Shock Protein, Hsp70.c, from Trypanosoma brucei

    Directory of Open Access Journals (Sweden)

    Adélle Burger

    2014-01-01

    Full Text Available The neglected tropical disease, African Trypanosomiasis, is fatal and has a crippling impact on economic development. Heat shock protein 70 (Hsp70 is an important molecular chaperone that is expressed in response to stress and Hsp40 acts as its co-chaperone. These proteins play a wide range of roles in the cell and they are required to assist the parasite as it moves from a cold blooded insect vector to a warm blooded mammalian host. A novel cytosolic Hsp70, from Trypanosoma brucei, TbHsp70.c, contains an acidic substrate binding domain and lacks the C-terminal EEVD motif. The ability of a cytosolic Hsp40 from Trypanosoma brucei J protein 2, Tbj2, to function as a co-chaperone of TbHsp70.c was investigated. The main objective was to functionally characterize TbHsp70.c to further expand our knowledge of parasite biology. TbHsp70.c and Tbj2 were heterologously expressed and purified and both proteins displayed the ability to suppress aggregation of thermolabile MDH and chemically denatured rhodanese. ATPase assays revealed a 2.8-fold stimulation of the ATPase activity of TbHsp70.c by Tbj2. TbHsp70.c and Tbj2 both demonstrated chaperone activity and Tbj2 functions as a co-chaperone of TbHsp70.c. In vivo heat stress experiments indicated upregulation of the expression levels of TbHsp70.c.

  6. PREFERENTIAL SECRETION OF INDUCIBLE HSP70 BY VITILIGO MELANOCYTES UNDER STRESS

    Science.gov (United States)

    Mosenson, Jeffrey A.; Flood, Kelsey; Klarquist, Jared; Eby, Jonathan M.; Koshoffer, Amy; Boissy, Raymond E.; Overbeck, Andreas; C.Tung, Rebecca; Poole, I. Caroline Le

    2014-01-01

    SUMMARY Inducible HSP70 (HSP70i) chaperones peptides from stressed cells, protecting them from apoptosis. Upon extracellular release, HSP70i serves an adjuvant function, enhancing immune responses to bound peptides. We questioned whether HSP70i differentially protects control and vitiligo melanocytes from stress and subsequent immune responses. We compared expression of HSP70i in skin samples, evaluated the viability of primary vitiligo and control melanocytes exposed to bleaching phenols, and measured secreted HSP70i. We determined whether HSP70i traffics to melanosomes to contact immunogenic proteins by cell fractionation, western blotting, electron microscopy and confocal microscopy. Viability of vitiligo and control melanocytes was equally affected under stress. However, vitiligo melanocytes secreted increased amounts of HSP70i in response to MBEH, corroborating with aberrant HSP70i expression in patient skin. Intracellular HSP70i colocalized with melanosomes, and more so in response to MBEH in vitiligo melanocytes. Thus whereas either agent is cytotoxic to melanocytes, MBEH preferentially induces immune responses to melanocytes. PMID:24354861

  7. Cytotoxicity and Expression of c-fos, HSP70, and GADD45/153 Proteins in Human Liver Carcinoma (HepG2 Cells Exposed to Dinitrotoluenes

    Directory of Open Access Journals (Sweden)

    Paul B. Tchounwou

    2005-08-01

    ŽÂ¼g/mL or within the dose range of 0-200 μg/mL for 2,6-DNT. The 45-kDa growth arrest and damage protein was significantly expressed at the dose range of 200 – 250μg/mL for 2,6-DNT and at the dose range of 200 - 400μg/mL for 2,4-DNT. Expression of 153-kDa growth arrest and DNA damage protein was significant at the 100, 200, and 250μg/mL doses for 2,6-DNT and at the 200 μg/mL dose for 2,4-DNT. Overall, these results indicate the potential of DNTs to induce cytotoxic, proteotoxic (HSP70, and genotoxic (GADD45/153 effects, as well as oxidative stress and pro-inflammatory reactions (c-fos.

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

    Science.gov (United States)

    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 NH4Cl) 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 (H2O2) 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.

  9. Heat shock protein-70 (Hsp-70 suppresses paraquat-induced neurodegeneration by inhibiting JNK and caspase-3 activation in Drosophila model of Parkinson's disease.

    Directory of Open Access Journals (Sweden)

    Arvind Kumar Shukla

    Full Text Available Parkinson's disease (PD is one of the most common neurodegenerative disorders with limited clinical interventions. A number of epidemiological as well as case-control studies have revealed an association between pesticide exposure, especially of paraquat (PQ and occurrence of PD. Hsp70, a molecular chaperone by function, has been shown as one of the modulators of neurological disorders. However, paucity of information regarding the protective role of Hsp70 on PQ-induced PD like symptoms led us to hypothesize that modulation of hsp70 expression in the dopaminergic neurons would improve the health of these cells. We took advantage of Drosophila, which is a well-established model for neurological research and also possesses genetic tools for easy manipulation of gene expression with limited ethical concern. Over-expression of hsp70 was found to reduce PQ-induced oxidative stress along with JNK and caspase-3 mediated dopaminergic neuronal cell death in exposed organism. Further, anti-apoptotic effect of hsp70 was shown to confer better homeostasis in the dopaminergic neurons of PQ-exposed organism as evidenced by their improved locomotor performance and survival. The study has merit in the context of human concern since we observed protection of dopaminergic neurons in PQ-exposed organism by over-expressing a human homologue of hsp70, HSPA1L, in these cells. The effect was parallel to that observed with Drosophila hsp70. These findings reflect the potential therapeutic applicability of hsp70 against PQ-induced PD like symptoms in an organism.

  10. Recent Gene Duplication and Subfunctionalization Produced a Mitochondrial GrpE, the Nucleotide Exchange Factor of the Hsp70 Complex, Specialized in Thermotolerance to Chronic Heat Stress in Arabidopsis1[W][OA

    Science.gov (United States)

    Hu, Catherine; Lin, Siou-ying; Chi, Wen-tzu; Charng, Yee-yung

    2012-01-01

    The duplication and divergence of heat stress (HS) response genes might help plants adapt to varied HS conditions, but little is known on the topic. Here, we examined the evolution and function of Arabidopsis (Arabidopsis thaliana) mitochondrial GrpE (Mge) proteins. GrpE acts as a nucleotide-exchange factor in the Hsp70/DnaK chaperone machinery. Genomic data show that AtMge1 and AtMge2 arose from a recent whole-genome duplication event. Phylogenetic analysis indicated that duplication and preservation of Mges occurred independently in many plant species, which suggests a common tendency in the evolution of the genes. Intron retention contributed to the divergence of the protein structure of Mge paralogs in higher plants. In both Arabidopsis and tomato (Solanum lycopersicum), Mge1 is induced by ultraviolet B light and Mge2 is induced by heat, which suggests regulatory divergence of the genes. Consistently, AtMge2 but not AtMge1 is under the control of HsfA1, the master regulator of the HS response. Heterologous expression of AtMge2 but not AtMge1 in the temperature-sensitive Escherichia coli grpE mutant restored its growth at 43°C. Arabidopsis T-DNA knockout lines under different HS regimes revealed that Mge2 is specifically required for tolerating prolonged exposure to moderately high temperature, as compared with the need of the heat shock protein 101 and the HS-associated 32-kD protein for short-term extreme heat. Therefore, with duplication and subfunctionalization, one copy of the Arabidopsis Mge genes became specialized in a distinct type of HS. We provide direct evidence supporting the connection between gene duplication and adaptation to environmental stress. PMID:22128139

  11. Effects of emodin and vitamin C on growth performance, biochemical parameters and two HSP70s mRNA expression of Wuchang bream (Megalobrama amblycephala Yih) under high temperature stress.

    Science.gov (United States)

    Ming, Jianhua; Xie, Jun; Xu, Pao; Ge, Xianping; Liu, Wenbin; Ye, Jinyun

    2012-05-01

    In order to study the effects of dietary emodin, high-dose vitamin C (Vc) and their combination on growth of Wuchang bream (Megalobrama amblycephala Y.) and its resistance to high temperature stress, 1200 healthy Wuchang bream with initial body weight of 133.44 ± 2.11 g were randomly divided into four groups: a control group fed with basal diet (containing 50.3 mg/kg Vc) and three treated groups fed with basal diets supplemented with 60 mg/kg emodin, 700 mg/kg Vc, and the combination of 60 mg/kg emodin + 700 mg/kg Vc, respectively. After feeding for 60 days, the growth performance of Wuchang bream was measured. Then 25 fish per tank were exposed to heat stress of 34 °C. The biochemical parameters of blood and liver, and expression levels of liver two HSP70s mRNA before and after heat stress were determined and the cumulative mortality of each group under heat stress was counted. The results showed that before stress, compared with the control, the weight gain (WG) and specific growth rate (SGR), serum total protein (TP), lysozyme (LSZ), and alkaline phosphatase (ALP) levels, liver superoxide dismutase (SOD) activity and expression level of HSP70 mRNA significantly increased in emodin and Vc groups while feed conversion rate (FCR), serum cortisol (COR), triglyceride (TG) and liver malondialdehyde (MDA) contents decreased (P stress, compared with the control, the serum TP, LSZ, ALP levels, liver SOD, CAT activities, and expression levels of HSC70 and HSP70 mRNAs increased in emodin and Vc groups in varying degrees and serum COR, glucose, glutamic-pyruvic transaminase (GPT), glutamic-oxaloacetic transaminase (GOT), TG and liver MDA levels decreased to some extent. Although these parameters had similar changing trend as above ones in combination group, it did not show any synergism either. Statistics showed that under heat stress, the cumulative mortalities of emodin and Vc groups, except at 6 h in emodin group, were significantly lower than that of the control (P 0

  12. Hsp70 plays an important role in high-fat diet induced gestational hyperglycemia in mice.

    Science.gov (United States)

    Xing, Baoheng; Wang, Lili; Li, Qin; Cao, Yalei; Dong, Xiujuan; Liang, Jun; Wu, Xiaohua

    2015-12-01

    Gestational diabetes mellitus (GDM) has emerged as an epidemic disease during the last decade, affecting about 2 to 5% pregnant women. Even among women who have gestational hyperglycemia may also be positively related to adverse outcomes as GDM. Since heat shock protein (Hsp) 70 has been reported to be associated with diabetes and insulin resistance and its expression was reported to be negatively regulated by the membrane-permeable Hsp70 inhibitor MAL3-101 while positively regulated by the Hsp70 activator BGP-15, we investigated whether Hsp70 played a role in a gestational hyperglycemia mouse model. Mice were divided into non-pregnant and pregnant groups, and each comprised three subgroups: control, high-fat diet (HFD) + MAL3-101, and HFD + BGP-15. We examined the serum levels of triglycerides, total cholesterol, glucose, and insulin, as well as conducted thermal detection of brown adipose tissue (BAT). The role of Hsp70 in BAT apoptosis was also investigated by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay and caspase-3 staining. Higher serum level of Hsp70 was associated with increased bodyweight gain after pregnancy in mice fed HFD. Circulating Hsp70 was elevated in control pregnant mice compared to control non-pregnant mice. BGP-induced serum Hsp70 expression reduced triglycerides, total cholesterol, glucose, and insulin levels in the serum. Additionally, thermal detection of BAT, TUNEL, and caspase-3 staining revealed relationship correlation between Hsp70 and BAT functions. Hsp70 level is associated with hyperglycemia during pregnancy. Our results support the role of Hsp70 in facilitating BAT activities and protecting BAT cells from apoptosis via caspase-3 pathway.

  13. Overexpression of the cochaperone CHIP enhances Hsp70-dependent folding activity in mammalian cells

    NARCIS (Netherlands)

    Kampinga, HH; Kanon, B; Salomons, FA; Kabakov, AE; Patterson, C

    CHIP is a cochaperone of Hsp70 that inhibits Hsp70-dependent refolding in vitro. However, the effect of altered expression of CHIP on the fate of unfolded proteins in mammalian cells has not been determined. Surprisingly, we found that overexpression of CHIP in fibroblasts increased the refolding of

  14. Ionizing radiation improves glioma-specific targeting of superparamagnetic iron oxide nanoparticles conjugated with cmHsp70.1 monoclonal antibodies (SPION-cmHsp70.1)

    Science.gov (United States)

    Shevtsov, Maxim A.; Nikolaev, Boris P.; Ryzhov, Vyacheslav A.; Yakovleva, Ludmila Y.; Marchenko, Yaroslav Y.; Parr, Marina A.; Rolich, Valerij I.; Mikhrina, Anastasiya L.; Dobrodumov, Anatolii V.; Pitkin, Emil; Multhoff, Gabriele

    2015-12-01

    The stress-inducible 72 kDa heat shock protein Hsp70 is known to be expressed on the membrane of highly aggressive tumor cells including high-grade gliomas, but not on the corresponding normal cells. Membrane Hsp70 (mHsp70) is rapidly internalized into tumor cells and thus targeting of mHsp70 might provide a promising strategy for theranostics. Superparamagnetic iron oxide nanoparticles (SPIONs) are contrast negative agents that are used for the detection of tumors with MRI. Herein, we conjugated the Hsp70-specific antibody (cmHsp70.1) which is known to recognize mHsp70 to superparamagnetic iron nanoparticles to assess tumor-specific targeting before and after ionizing irradiation. In vitro experiments demonstrated the selectivity of SPION-cmHsp70.1 conjugates to free and mHsp70 in different tumor cell types (C6 glioblastoma, K562 leukemia, HeLa cervix carcinoma) in a dose-dependent manner. High-resolution MRI (11 T) on T2-weighted images showed the retention of the conjugates in the C6 glioma model. Accumulation of SPION-cmHsp70.1 nanoparticles in the glioma resulted in a nearly 2-fold drop of values in comparison to non-conjugated SPIONs. Biodistribution analysis using NLR-M2 measurements showed a 7-fold increase in the tumor-to-background (normal brain) uptake ratio of SPION-cmHsp70.1 conjugates in glioma-bearing rats in comparison to SPIONs. This accumulation within Hsp70-positive glioma was further enhanced after a single dose (10 Gy) of ionizing radiation. Elevated accumulation of the magnetic conjugates in the tumor due to radiosensitization proves the combination of radiotherapy and application of Hsp70-targeted agents in brain tumors.The stress-inducible 72 kDa heat shock protein Hsp70 is known to be expressed on the membrane of highly aggressive tumor cells including high-grade gliomas, but not on the corresponding normal cells. Membrane Hsp70 (mHsp70) is rapidly internalized into tumor cells and thus targeting of mHsp70 might provide a promising strategy

  15. HSP70-Inducible hNIS-IRES-eGFP Reporter Imaging: Response to Heat Shock

    Directory of Open Access Journals (Sweden)

    Jiantu Che

    2007-11-01

    Full Text Available A retroviral vector pQHSP70/hNIS-IRES-eGFP (pQHNIG70 was constructed containing the hNIS-IRES-eGFP dual-reporter genes under the control of an inducible human heat shock protein (HSP70 promoter and RG2-pQHSP70/hNIS-IRES-eGFP (RG2-pQHNIG70 transduced cells were generated. Heat-induced expression of both reporter genes in RG2-pQHNIG70 cells was validated by enhanced green fluorescent protein (eGFP fluorescence-activated cell sorter, in vitro radiotracer assays, and immunoblot and immunocytochemistry. A 2.2- to 6.1-fold (131I−, a 6.1- to 14.4-fold (99mTcO4−, and a 5.1- to 39-fold (fluorescence increase above baseline was observed in response to graded hyperthermia (39–43°C. Increases in eGFP fluorescence and radiotracer uptake were first noted at 6 hours, reached a maximum at 24 hours, and fell toward baseline at 72 hours. A stable ratio of radiotracer uptake to eGFP fluorescence and to heat shock protein (HSP70 protein was demonstrated over a wide range of expression levels, induced by different levels of heating. We also demonstrate that the local application of heat on RG2-pQHNIG70 xenografts can effectively induce hNIS and eGFP gene expression in vivo and that this expression can be efficiently visualized by fluorescence, scintigraphic, and micro–positron emission tomography imaging. Endogenous HSP70 protein and reporter expression was confirmed by postmortem tissue evaluations (immunoblot and immunohistochemistry. The pQHNIG70 reporter system can be used to study stress and drug responses in transduced cells and tissues.

  16. Over-expression of gene encoding heat shock protein 70 from Mycobacterium tuberculosis and its evaluation as vaccine adjuvant

    Directory of Open Access Journals (Sweden)

    J Dhakal

    2013-01-01

    Full Text Available Background: Heat shock proteins (Hsps are evolutionary ancient and highly conserved molecular chaperons found in prokaryotes as well as eukaryotes. Hsp70 is a predominant member of Hsp family. Microbial Hsp70s (mHsp70s have acquired special significance in immunity since they have been shown to be potent activators of the innate immune system and generate specific immune responses against tumours and infectious agents. Objectives: The present study was aimed to clone express and purify recombinant Hsp70 from the Mycobacterium tuberculosis and characterise it immunologically. The study also aimed at determining the potential of recombinant M. tuberculosis heat shock protein (rMTB-Hsp70 as adjuvant or antigen carrier. Materials and Methods: Cloning of M. tuberculosis heat shock protein (MTB-Hsp70 amplicon was carried out using the pGEMT-Easy vector although for expression, pProExHTb prokaryotic expression vector was used. Purification of recombinant Hsp70 was carried out by nickel-nitrilotriacetic acid (Ni-NTA affinity chromatography. For immunological characterization and determining the adjuvant effect of MTB-Hsp70, BALB/c mice were used. The data obtained was statistically analysed. Results: Hsp70 gene was cloned, sequenced and the sequence data were submitted to National Center for Biotechnology Information (NCBI. Recombinant MTB-Hsp70 was successfully over-expressed using the prokaryotic expression system and purified to homogeneity. The protein was found to be immunodominant. Significant adjuvant effect was produced by the rMTB-Hsp70 when inoculated with recombinant outer membrane protein 31; however, effect was less than the conventionally used the Freund′s adjuvant. Conclusion: Protocol standardised can be followed for bulk production of rHsp70 in a cost-effective manner. Significant adjuvant effect was produced by rMTB-Hsp70; however, the effect was than Freund′s adjuvant. Further, studies need to be carried out to explore its

  17. Metabolic fates and effects of nitrite in brown trout under normoxic and hypoxic conditions: blood and tissue nitrite metabolism and interactions with branchial NOS, Na+/K+-ATPase and hsp70 expression

    DEFF Research Database (Denmark)

    Jensen, Frank Bo; Gerber, Lucie; Hansen, Marie Niemann

    2015-01-01

    Nitrite secures essential nitric oxide (NO) bioavailability in hypoxia at low endogenous concentrations, whereas it becomes toxic at high concentrations. We exposed brown trout to normoxic and hypoxic water in the absence and presence of added ambient nitrite to decipher the cellular metabolism...... and effects of nitrite at basal and elevated concentrations under different oxygen regimes. We also tested hypotheses concerning the influence of nitrite on branchial nitric oxide synthase (NOS), Na+/K+-ATPase (nka) and heat shock protein (hsp70) mRNA expression. Basal plasma and erythrocyte nitrite levels...... were higher in hypoxia than normoxia, suggesting increased NOS activity. Nitrite exposure strongly elevated nitrite concentrations in plasma, erythrocytes, heart tissue and white muscle, which was associated with an extensive metabolism of nitrite to nitrate and to iron-nitrosylated and S...

  18. Plasma exosomes are enriched in Hsp70 and modulated by stress and cortisol in rainbow trout.

    Science.gov (United States)

    Faught, Erin; Henrickson, Lynsi; Vijayan, Mathilakath M

    2017-02-01

    Exosomes are endosomally derived vesicles that are secreted from cells and contain a suite of molecules, including proteins and nucleic acids. Recent studies suggest the possibility that exosomes in circulation may be affecting recipient target cell function, but the modes of action are unclear. Here, we tested the hypothesis that exosomes are in circulation in fish plasma and that these vesicles are enriched with heat shock protein 70 (Hsp70). Exosomes were isolated from rainbow trout (Oncorhynchus mykiss) plasma using differential centrifugation, and their presence was confirmed by transmission electron microscopy and the exosomal marker acetylcholinesterase. Plasma exosomes were enriched with Hsp70, and this stress protein was transiently elevated in trout plasma in response to a heat shock in vivo Using trout hepatocytes in primary culture, we tested whether stress levels of cortisol, the principle corticosteroid in teleosts, regulates exosomal Hsp70 content. As expected, a 1-h heat shock (+15°C above ambient) increased Hsp70 expression in hepatocytes, and this led to higher Hsp70 enrichment in exosomes over a 24-h period. However, cortisol treatment significantly reduced the expression of Hsp70 in exosomes released from either unstressed or heat-shocked hepatocytes. This cortisol-mediated suppression was not specific to Hsp70 as beta-actin expression was also reduced in exosomes released from hepatocytes treated with the steroid. Our results suggest that circulating Hsp70 is released from target tissues via exosomes, and their release is modulated by stress and cortisol. Overall, we propose a novel role for extracellular vesicular transport of Hsp70 in the organismal stress response. © 2017 Society for Endocrinology.

  19. Hsp70 positively regulates porcine circovirus type 2 replication in vitro.

    Science.gov (United States)

    Liu, Jie; Bai, Juan; Zhang, Lili; Jiang, Zhihua; Wang, Xianwei; Li, Yufeng; Jiang, Ping

    2013-12-01

    The Hsp70 chaperone plays a central role in multiple processes within cells. Porcine circovirus type 2 (PCV2) is the essential causal agent of post-weaning multisystemic wasting syndrome (PMWS), which has spread worldwide. But the mechanism of PCV2 replication remains poorly understood. In this study, we firstly found the positive effect of heat stress on the replication of PCV2 in the continuous porcine monocytic cell line 3D4/31. Downregulation of Hsp70 by the specific chaperone inhibitor Quercetin or RNA interference and upregulation of Hsp70 by expression from a recombinant adenovirus showed that Hsp70 enhanced PCV2 genome replication and virion production. A specific interaction between Hsp70 and PCV2 Cap was confirmed by colocalization by confocal microscopy and co-immunoprecipitation. Furthermore, the NF-κB pathway was activated and caspase-3 activity was reduced when Hsp70 was overexpressed in PCV2-infected 3D4/31 cells. These data suggested that Hsp70 positively regulated PCV2 replication, which being helpful for understanding the molecular mechanism of PCV2 infection. © 2013 Elsevier Inc. All rights reserved.

  20. Hsp70A and GlsA interact as partner chaperones to regulate asymmetric division in Volvox.

    Science.gov (United States)

    Cheng, Qian; Pappas, Valeria; Hallmann, Armin; Miller, Stephen M

    2005-10-15

    GlsA, a J-protein chaperone, is required for the asymmetric divisions that set aside germ and somatic cell precursors during embryogenesis in Volvox carteri, and previous evidence indicated that this function requires an intact Hsp70-binding site. To determine if Hsp70A, the only known cytoplasmic Hsp70 in V. carteri, is the chaperone partner of GlsA, we investigated the localization of the two proteins during critical stages of embryogenesis and tested their capacity to interact. We found that a substantial fraction of Hsp70A co-localizes with GlsA, both in interphase and mitotic blastomeres. In addition, Hsp70A coimmunoprecipitated with GlsA, and co-expression of GlsA and Hsp70A variants partially rescued the Gls phenotype of a glsA mutant, whereas neither variant by itself rescued the mutant phenotype. Immunofluorescence analysis demonstrated that GlsA is about equally abundant in all blastomeres at all cleavage stages examined but that Hsp70A is more abundant in anterior (asymmetrically dividing) blastomeres than in posterior (symmetrically dividing) blastomeres during the period of asymmetric division. We conclude that Hsp70A and GlsA function as chaperone partners that regulate asymmetric division and that the relative abundance of Hsp70A in asymmetrically dividing embryos may determine which blastomeres divide asymmetrically and which do not.

  1. In Vitro Evaluation of Influenza M2 and Leishmania major HSP70 (221-604) Chimer Protein

    Science.gov (United States)

    Fotouhi, Fatemeh; Farahmand, Behrokh; Heidarchi, Behnaz; Esghaei, Maryam; Rafati, Sima; Tavassoti Kheiri, Masoumeh

    2014-01-01

    Background: Permanent antigenic variation of influenza viruses causes a major concern to develop an effective human influenza vaccine. Conserved antigens are new vaccine candidates because it is not necessary to match the prepared vaccine with circulating strains. Ion channel M2 protein is conserved among all influenza A viruses, allowing the virus to enter host cells. Objectives: To prepare an effective vaccine against influenza A viruses, a chimerical DNA plasmid encoding Influenza virus M2 protein and Leishmania major HSP70 was constructed. Materials and Methods: Influenza A/New Caledonia/20/99 (H1N1) was inoculated into MDCK cell line and total RNA was extracted. The full length M2 gene was amplified by RT-PCR using designed specific primers, cloned into pGEM-T Easy cloning vector and completely sequenced. The M2 gene was then subcloned into the pcDNA upstream of HSP70 gene. Recombinant plasmids were transfected into COS-7 cells to evaluate protein expression. Results: The recombinant plasmids were confirmed by PCR, restriction enzyme analysis and sequencing. Three dimensional structure of chimer protein was assessed using specific software. Transient protein expression in eukaryotic cells was confirmed by specific mRNA detection, indirect Immunofluorescence test and western blotting. Conclusions: M2-HSP70 chimer protein was successfully expressed in eukaryotic cells. Computational studies of chimer peptide sequence revealed that fusing HSP to the C-terminal of M2 protein does not mask the predominant epitope of M2. HSP70 is a molecular chaperon and immunostimulatory component. Genetically fusing antigens to HSPs leads to the enrichment of DNA vaccine potency. The immunogenicity of this construct with different formulation would be evaluated in further investigations. PMID:25485058

  2. Combined pharmacological induction of Hsp70 suppresses prion protein neurotoxicity in Drosophila.

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    Yan Zhang

    Full Text Available Prion diseases are rare and aggressive neurodegenerative disorders caused by the accumulation of misfolded, toxic conformations of the prion protein (PrP. Therapeutic strategies directed at reducing the levels of PrP offer the best chance of delaying or halting disease progression. The challenge, though, is to define pharmacologic targets that result in reduced PrP levels. We previously reported that expression of wild type hamster PrP in flies induces progressive locomotor dysfunction and accumulation of pathogenic PrP conformations, while co-expression of human Hsp70 delayed these changes. To validate the therapeutic potential of Hsp70, we treated flies with drugs known to induce Hsp70 expression, including the Hsp90 inhibitor 17-DMAG and the glucocorticoid dexamethasone. Although the individual treatment with these compounds produced no significant benefits, their combination significantly increased the level of inducible Hsp70, decreased the level of total PrP, reduced the accumulation of pathogenic PrP conformers, and improved locomotor activity. Thus, the combined action of two pharmacological activators of Hsp70 with distinct targets results in sustained high levels of inducible Hsp70 with improved behavioral output. These findings can have important therapeutic applications for the devastating prion diseases and other related proteinopathies.

  3. Characterisation of the Plasmodium falciparum Hsp70-Hsp90 organising protein (PfHop).

    Science.gov (United States)

    Gitau, Grace W; Mandal, Pradipta; Blatch, Gregory L; Przyborski, Jude; Shonhai, Addmore

    2012-03-01

    Malaria is caused by Plasmodium species, whose transmission to vertebrate hosts is facilitated by mosquito vectors. The transition from the cold blooded mosquito vector to the host represents physiological stress to the parasite, and additionally malaria blood stage infection is characterised by intense fever periods. In recent years, it has become clear that heat shock proteins play an essential role during the parasite's life cycle. Plasmodium falciparum expresses two prominent heat shock proteins: heat shock protein 70 (PfHsp70) and heat shock protein 90 (PfHsp90). Both of these proteins have been implicated in the development and pathogenesis of malaria. In eukaryotes, Hsp70 and Hsp90 proteins are functionally linked by an essential adaptor protein known as the Hsp70-Hsp90 organising protein (Hop). In this study, recombinant P. falciparum Hop (PfHop) was heterologously produced in E. coli and purified by nickel affinity chromatography. Using specific anti-PfHop antisera, the expression and localisation of PfHop in P. falciparum was investigated. PfHop was shown to co-localise with PfHsp70 and PfHsp90 in parasites at the trophozoite stage. Gel filtration and co-immunoprecipitation experiments suggested that PfHop was present in a complex together with PfHsp70 and PfHsp90. The association of PfHop with both PfHsp70 and PfHsp90 suggests that this protein may mediate the functional interaction between the two chaperones.

  4. Extracts Obtained from Pterocarpus angolensis DC and Ziziphus mucronata Exhibit Antiplasmodial Activity and Inhibit Heat Shock Protein 70 (Hsp70 Function

    Directory of Open Access Journals (Sweden)

    Tawanda Zininga

    2017-07-01

    Full Text Available Malaria parasites are increasingly becoming resistant to currently used antimalarial therapies, therefore there is an urgent need to expand the arsenal of alternative antimalarial drugs. In addition, it is also important to identify novel antimalarial drug targets. In the current study, extracts of two plants, Pterocarpus angolensis and Ziziphus mucronata were obtained and their antimalarial functions were investigated. Furthermore, we explored the capability of the extracts to inhibit Plasmodium falciparum heat shock protein 70 (Hsp70 function. Heat shock protein 70 (Hsp70 are molecular chaperones whose function is to facilitate protein folding. Plasmodium falciparum the main agent of malaria, expresses two cytosol-localized Hsp70s: PfHsp70-1 and PfHsp70-z. The PfHsp70-z has been reported to be essential for parasite survival, while inhibition of PfHsp70-1 function leads to parasite death. Hence both PfHsp70-1 and PfHsp70-z are potential antimalarial drug targets. Extracts of P. angolensis and Z. mucronata inhibited the basal ATPase and chaperone functions of the two parasite Hsp70s. Furthermore, fractions of P. angolensis and Z. mucronata inhibited P. falciparum 3D7 parasite growth in vitro. The extracts obtained in the current study exhibited antiplasmodial activity as they killed P. falciparum parasites maintained in vitro. In addition, the findings further suggest that some of the compounds in P. angolensis and Z. mucronata may target parasite Hsp70 function.

  5. Loss of the Inducible Hsp70 Delays the Inflammatory Response to Skeletal Muscle Injury and Severely Impairs Muscle Regeneration

    Science.gov (United States)

    Howard, Travis M.; Ahn, Bumsoo; Ferreira, Leonardo F.

    2013-01-01

    Skeletal muscle regeneration following injury is a highly coordinated process that involves transient muscle inflammation, removal of necrotic cellular debris and subsequent replacement of damaged myofibers through secondary myogenesis. However, the molecular mechanisms which coordinate these events are only beginning to be defined. In the current study we demonstrate that Heat shock protein 70 (Hsp70) is increased following muscle injury, and is necessary for the normal sequence of events following severe injury induced by cardiotoxin, and physiological injury induced by modified muscle use. Indeed, Hsp70 ablated mice showed a significantly delayed inflammatory response to muscle injury induced by cardiotoxin, with nearly undetected levels of both neutrophil and macrophage markers 24 hours post-injury. At later time points, Hsp70 ablated mice showed sustained muscle inflammation and necrosis, calcium deposition and impaired fiber regeneration that persisted several weeks post-injury. Through rescue experiments reintroducing Hsp70 intracellular expression plasmids into muscles of Hsp70 ablated mice either prior to injury or post-injury, we confirm that Hsp70 optimally promotes muscle regeneration when expressed during both the inflammatory phase that predominates in the first four days following severe injury and the regenerative phase that predominates thereafter. Additional rescue experiments reintroducing Hsp70 protein into the extracellular microenvironment of injured muscles at the onset of injury provides further evidence that Hsp70 released from damaged muscle may drive the early inflammatory response to injury. Importantly, following induction of physiological injury through muscle reloading following a period of muscle disuse, reduced inflammation in 3-day reloaded muscles of Hsp70 ablated mice was associated with preservation of myofibers, and increased muscle force production at later time points compared to WT. Collectively our findings indicate that

  6. Expression of Heat Stress Protein 70 mRNA in Patients with Chronic Ob-structive Pulmonary Disease and Its Significance

    Institute of Scientific and Technical Information of China (English)

    ZHAO Jianping; XIE Jungang; XU Yongjian; ZHANG Zhenxiang; ZHANG Ning

    2005-01-01

    The effects of cigarette smoke extract (CSE) on the expression of heat stress protein 70(Hsp70) in human bronchi smooth muscle cells were investigated in vitro, and the changes in Hsp70 mRNA in the patients with chronic obstructive pulmonary disease and their significance were explored. Human bronchi smooth muscle cells were cultured with CSE at the different concentrations. The expression of Hsp70 mRNA and Hsp70 was detected by reverse translation-polymerase chain reaction (RT-PCR) and Western blotting respectively. Levels of Hsp70 mRNA and Hsp70 in lymphocytes from 20 patients with COPD and 20 healthy smoking control subjects were measured by RT-PCR and Western blotting. The results showed the expression of both Hsp70 mRNA and Hsp70 was decreased conformably in human bronchi smooth muscle cells treated with CSE at certain concentration in vitro. The A values of the Hsp70 mRNA expression were 0. 24±0.11 and 0.42±0.13 respectively in COPD patients and healthy smoking controls with the difference being significant (P<0.01). There was also significant difference in the A values of the Hsp70 expression between COPD patients and healthy smoking controls (20.9±9.9 vs 44.8±15.3, P<0.01). The levels of Hsp70 mRNA had strongly positive correlation with Hsp70 protein (r = 0. 85, P<0.01). It was suggested that the expression of Hsp70 mRNA was in concordance with the expression of Hsp70, which could provide a basis on the study of Hsp70 gene regulation and Hsp70 gene in the development of COPD.

  7. Differential role for ERK2 in anoxia-induced activation of transcription and translation of Hsp70 in NIH 3T3 cells

    DEFF Research Database (Denmark)

    Ossum, Carlo; Lauritsen, Anders N.; Karottki, Dorina Gabriela

    2011-01-01

    and transcription was involved. Inhibition of the MAP kinase p38, which was transiently activated during chemical anoxia, had no effect on the increase in Hsp70 expression whereas an inhibitor of reactive oxygen species and inhibition of the phosphatase PP1 and PP2a inhibited the increase in Hsp70 expression......Hsp70 has the ability to enhance the recovery of stressed cells by its ability to catalyze the reassembly of damaged proteins. Such a chaperoning function is essential for the Hsp70-mediated protection against anoxic stress that causes protein denaturation. We have studied induction of both....... Inhibition of ERK2 by the MEK inhibitor PD98059 resulted in strong inhibition of Hsp70 protein expression and simultaneous stimulation of hsp70 transcription....

  8. Peptides and aptamers targeting HSP70: a novel approach for anticancer chemotherapy.

    Science.gov (United States)

    Rérole, Anne-Laure; Gobbo, Jessica; De Thonel, Aurelie; Schmitt, Elise; Pais de Barros, Jean Paul; Hammann, Arlette; Lanneau, David; Fourmaux, Eric; Demidov, Oleg N; Deminov, Oleg; Micheau, Olivier; Lagrost, Laurent; Colas, Pierre; Kroemer, Guido; Garrido, Carmen

    2011-01-15

    The inhibition of heat shock protein 70 (HSP70) is an emerging strategy in cancer therapy. Unfortunately, no specific inhibitors are clinically available. By yeast two-hybrid screening, we have identified multiple peptide aptamers that bind HSP70. When expressed in human tumor cells, two among these peptide aptamers-A8 and A17-which bind to the peptide-binding and the ATP-binding domains of HSP70, respectively, specifically inhibited the chaperone activity, thereby increasing the cells' sensitivity to apoptosis induced by anticancer drugs. The 13-amino acid peptide from the variable region of A17 (called P17) retained the ability to specifically inhibit HSP70 and induced the regression of subcutaneous tumors in vivo after local or systemic injection. This antitumor effect was associated with an important recruitment of macrophages and T lymphocytes into the tumor bed. Altogether, these data indicate that peptide aptamers or peptides that target HSP70 may be considered as novel lead compounds for cancer therapy. © 2011 AACR.

  9. Heat stress induces expression of HSP genes in genetically divergent chickens.

    Science.gov (United States)

    Cedraz, Haniel; Gromboni, Juliana Gracielle Gonzaga; Garcia, Antonio Amandio Pinto; Farias Filho, Ronaldo Vasconcelos; Souza, Teillor Machado; Oliveira, Eduardo Ribeiro de; Oliveira, Elizangela Bonfim de; Nascimento, Carlos Souza do; Meneghetti, Camila; Wenceslau, Amauri Arias

    2017-01-01

    Chickens are animals that are sensitive to thermal stress, which may decrease their production level in terms that it affects feed intake and thus, decreasing body weight gain. The Heat Shock Factors (HSF) and Heat Shock Proteins (HSP) genes are involved in the key cellular defense mechanisms during exposure in hot environments. Aimed with this study to analyze the expression of HSF1, HSF3, HSP70 and HSP90 genes in two local breeds (Peloco and Caneluda) and a commercial broiler line (Cobb 500®) to verify differences in resistance of these chicken to Heat stress treatment. Chicken were submitted to heat stress under an average temperature of 39°C ± 1. Under stress environment, the HSP70 and HSP90 genes were more expressed in backyard chickens than in broiler. There was a difference in HSP70 and HSP90 expression between Caneluda and Cobb and between Peloco and Cobb under stress and comfort environment respectively. HSP70 expression is higher in local breeds during heat stress than in a commercial broiler line. No significant differences were observed in the expression of HSF1 and HSF3 genes between breeds or environments. HSP70 and HSP90 genes are highly expressed, HSF1 and HSF3 genes did not have high expression in all genetic groups. HSP70 and HSP90 are highly expressed in Peloco and Caneluda within heat stress, these breeds proved to be very resistant to high temperature.

  10. Influence of In Vitro IL-2 or IL-15 Alone or in Combination with Hsp 70 Derived 14-Mer Peptide (TKD on the Expression of NK Cell Activatory and Inhibitory Receptors on Peripheral Blood T Cells, B Cells and NKT Cells.

    Directory of Open Access Journals (Sweden)

    Ilona Hromadnikova

    Full Text Available Previous studies from Multhoff and colleagues reported that plasma membrane Hsp70 acts as a tumour-specific recognition structure for activated NK cells, and that the incubation of NK cells with Hsp70 and/or a 14-mer peptide derived from the N-terminal sequence of Hsp70 (TKDNNLLGRFELSG, TKD, aa 450-463 plus a low dose of IL-2 triggers NK cell proliferation and migration, and their capacity to kill cancer cells expressing membrane Hsp70. Herein, we have used flow cytometry to determine the influence of in vitro stimulation of peripheral blood mononuclear cells from healthy individuals with IL-2 or IL-15, either alone or in combination with TKD peptide on the cell surface expression of CD94, NK cell activatory receptors (CD16, NK2D, NKG2C, NKp30, NKp44, NKp46, NKp80, KIR2DL4, DNAM-1 and LAMP1 and NK cell inhibitory receptors (NKG2A, KIR2DL2/L3, LIR1/ILT-2 and NKR-P1A by CD3+CD56+ (NKT, CD3+CD4+, CD3+CD8+ and CD19+ populations. NKG2D, DNAM-1, LAMP1 and NKR-P1A expression was upregulated after the stimulation with IL-2 or IL-15 alone or in combination with TKD in NKT, CD8+ T cells and B cells. CD94 was upregulated in NKT and CD8+ T cells. Concurrently, an increase in a number of CD8+ T cells expressing LIR1/ILT-2 and CD4+ T cells positive for NKR-P1A was observed. The proportion of CD8+ T cells that expressed NKG2D was higher after IL-2/TKD treatment, when compared with IL-2 treatment alone. In comparison with IL-15 alone, IL-15/TKD treatment increased the proportion of NKT cells that were positive for CD94, LAMP1 and NKRP-1A. The more potent effect of IL-15/TKD on cell surface expression of NKG2D, LIR1/ILT-2 and NKRP-1A was observed in B cells compared with IL-15 alone. However, this increase was not of statistical significance. IL-2/TKD induced significant upregulation of LAMP1 in CD8+ T cells compared with IL-2 alone. Besides NK cells, other immunocompetent cells present within the fraction of peripheral blood mononuclear cells were influenced by

  11. Hsp70 stabilizes lysosomes and reverts Niemann-Pick disease-associated lysosomal pathology

    DEFF Research Database (Denmark)

    Kirkegaard, Thomas; Roth, Anke G; Petersen, Nikolaj H T

    2010-01-01

    inhibition of ASM, effectively revert the Hsp70-mediated stabilization of lysosomes. Notably, the reduced ASM activity in cells from patients with Niemann-Pick disease (NPD) A and B-severe lysosomal storage disorders caused by mutations in the sphingomyelin phosphodiesterase 1 gene (SMPD1) encoding for ASM...

  12. Restoring Anticancer Immune Response by Targeting Tumor-Derived Exosomes With a HSP70 Peptide Aptamer.

    Science.gov (United States)

    Gobbo, Jessica; Marcion, Guillaume; Cordonnier, Marine; Dias, Alexandre M M; Pernet, Nicolas; Hammann, Arlette; Richaud, Sarah; Mjahed, Hajare; Isambert, Nicolas; Clausse, Victor; Rébé, Cédric; Bertaut, Aurélie; Goussot, Vincent; Lirussi, Frédéric; Ghiringhelli, François; de Thonel, Aurélie; Fumoleau, Pierre; Seigneuric, Renaud; Garrido, Carmen

    2016-03-01

    Exosomes, via heat shock protein 70 (HSP70) expressed in their membrane, are able to interact with the toll-like receptor 2 (TLR2) on myeloid-derived suppressive cells (MDSCs), thereby activating them. We analyzed exosomes from mouse (C57Bl/6) and breast, lung, and ovarian cancer patient samples and cultured cancer cells with different approaches, including nanoparticle tracking analysis, biolayer interferometry, FACS, and electron microscopy. Data were analyzed with the Student's t and Mann-Whitney tests. All statistical tests were two-sided. We showed that the A8 peptide aptamer binds to the extracellular domain of membrane HSP70 and used the aptamer to capture HSP70 exosomes from cancer patient samples. The number of HSP70 exosomes was higher in cancer patients than in healthy donors (mean, ng/mL ± SD = 3.5 ± 1.7 vs 0.17 ± 0.11, respectively, P = .004). Accordingly, all cancer cell lines examined abundantly released HSP70 exosomes, whereas "normal" cells did not. HSP70 had higher affinity for A8 than for TLR2; thus, A8 blocked HSP70/TLR2 association and the ability of tumor-derived exosomes to activate MDSCs. Treatment of tumor-bearing C57Bl/6 mice with A8 induced a decrease in the number of MDSCs in the spleen and inhibited tumor progression (n = 6 mice per group). Chemotherapeutic agents such as cisplatin or 5FU increase the amount of HSP70 exosomes, favoring the activation of MDSCs and hampering the development of an antitumor immune response. In contrast, this MDSC activation was not observed if cisplatin or 5FU was combined with A8. As a result, the antitumor effect of the drugs was strongly potentiated. A8 might be useful for quantifying tumor-derived exosomes and for cancer therapy through MDSC inhibition. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  13. Inducible HSP70 antagonizes IL-1β cytocidal effects through inhibiting NF-kB activation via destabilizing TAK1 in HeLa cells.

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    Xiang Cao

    Full Text Available BACKGROUND: Despite several reports describing the HSP70-mediated cytoprotection against IL-1, the precise mechanism for this phenomenon remains to be determined. METHODS/PRINCIPAL FINDINGS: Here we used HeLa cells, a human epithelial carcinoma cell line, to evaluate the role of inducible HSP70 in response of IL-1β stimulation. We found that inducible HSP70 antagonized the cytotoxicity of IL-1β and improved the survival of HeLa cells. Further investigation demonstrated that increased expression level of inducible HSP70 reduced the complex of TAK1 and HSP90, and promoted the degradation of TAK1 protein via proteasome pathway. By overexpression and RNAi knockdown, we showed that inducible HSP70 modulated the NF-kB but not MAPKs signalings through influencing the stability of TAK1 protein in HeLa cells. Moreover, overexpression of HSP70 attenuated the production of iNOS upon IL-1β stimulation, validating that inducible HSP70 serves as a cytopretective factor to antagonize the cytocidal effects of IL-1β in HeLa cells. CONCLUSIONS/SIGNIFICANCE: Our observations provide evidence for a novel signaling mechanism involving HSP70, TAK1, and NF-κB in the response of IL-1β cytocidal effects. This research also provides insight into mechanisms by which HSP70 exerts its cytoprotective action upon toxic stimuli in tumor cells.

  14. The HSP70 chaperone machines of Chlamydomonas are induced by cold stress.

    Science.gov (United States)

    Maikova, Anna; Zalutskaya, Zhanneta; Lapina, Tatiana; Ermilova, Elena

    2016-10-01

    The responses of Chlamydomonas reinhardtii cells to low temperatures have not been extensively studied compared with other stresses. Like other organisms, this green alga has heat shock protein 70s (HSP70s) that are located in chloroplast, mitochondrion and cytosol. To test whether temperature downshifts affected HSP70s synthesis, we used real-time PCR and protein gel blot analysis. C. reinhardtii cells exposed to cold stress show increased HSP70s mRNA levels. Genes encoding other components of HSP70 chaperone machines (e.g. CGE1, CDJ1, HSP90C and HSP90A) are also up-regulated in response to decreased temperature. We demonstrated that the accumulation of all analyzed mRNA occur more slowly and with reduced amplitude in cells exposed to cold than in cells treated with heat. Furthermore, C. reinhardtii cells display the splicing of the CGE1 transcript that was dependent on low temperature. Finally, the transcription regulator of C. reinhardtii HSF1 is also cold-responsive, suggesting its role in the transcriptional regulation of HSP genes at low temperature. Copyright © 2016 Elsevier GmbH. All rights reserved.

  15. Plasmodium falciparum Hsp70-z, an Hsp110 homologue, exhibits independent chaperone activity and interacts with Hsp70-1 in a nucleotide-dependent fashion.

    Science.gov (United States)

    Zininga, Tawanda; Achilonu, Ikechukwu; Hoppe, Heinrich; Prinsloo, Earl; Dirr, Heini W; Shonhai, Addmore

    2016-05-01

    The role of molecular chaperones, among them heat shock proteins (Hsps), in the development of malaria parasites has been well documented. Hsp70s are molecular chaperones that facilitate protein folding. Hsp70 proteins are composed of an N-terminal nucleotide binding domain (NBD), which confers them with ATPase activity and a C-terminal substrate binding domain (SBD). In the ADP-bound state, Hsp70 possesses high affinity for substrate and releases the folded substrate when it is bound to ATP. The two domains are connected by a conserved linker segment. Hsp110 proteins possess an extended lid segment, a feature that distinguishes them from canonical Hsp70s. Plasmodium falciparum Hsp70-z (PfHsp70-z) is a member of the Hsp110 family of Hsp70-like proteins. PfHsp70-z is essential for survival of malaria parasites and is thought to play an important role as a molecular chaperone and nucleotide exchange factor of its cytosolic canonical Hsp70 counterpart, PfHsp70-1. Unlike PfHsp70-1 whose functions are fairly well established, the structure-function features of PfHsp70-z remain to be fully elucidated. In the current study, we established that PfHsp70-z possesses independent chaperone activity. In fact, PfHsp70-z appears to be marginally more effective in suppressing protein aggregation than its cytosol-localized partner, PfHsp70-1. Furthermore, based on coimmunoaffinity chromatography and surface plasmon resonance analyses, PfHsp70-z associated with PfHsp70-1 in a nucleotide-dependent fashion. Our findings suggest that besides serving as a molecular chaperone, PfHsp70-z could facilitate the nucleotide exchange function of PfHsp70-1. These dual functions explain why it is essential for parasite survival.

  16. The molecular chaperone HSP70 binds to and stabilizes NOD2, an important protein involved in Crohn disease.

    Science.gov (United States)

    Mohanan, Vishnu; Grimes, Catherine Leimkuhler

    2014-07-04

    Microbes are detected by the pathogen-associated molecular patterns through specific host pattern recognition receptors. Nucleotide-binding oligomerization domain-containing protein 2 (NOD2) is an intracellular pattern recognition receptor that recognizes fragments of the bacterial cell wall. NOD2 is important to human biology; when it is mutated it loses the ability to respond properly to bacterial cell wall fragments. To determine the mechanisms of misactivation in the NOD2 Crohn mutants, we developed a cell-based system to screen for protein-protein interactors of NOD2. We identified heat shock protein 70 (HSP70) as a protein interactor of both wild type and Crohn mutant NOD2. HSP70 has previously been linked to inflammation, especially in the regulation of anti-inflammatory molecules. Induced HSP70 expression in cells increased the response of NOD2 to bacterial cell wall fragments. In addition, an HSP70 inhibitor, KNK437, was capable of decreasing NOD2-mediated NF-κB activation in response to bacterial cell wall stimulation. We found HSP70 to regulate the half-life of NOD2, as increasing the HSP70 level in cells increased the half-life of NOD2, and down-regulating HSP70 decreased the half-life of NOD2. The expression levels of the Crohn-associated NOD2 variants were less compared with wild type. The overexpression of HSP70 significantly increased NOD2 levels as well as the signaling capacity of the mutants. Thus, our study shows that restoring the stability of the NOD2 Crohn mutants is sufficient for rescuing the ability of these mutations to signal the presence of a bacterial cell wall ligand. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  17. The Molecular Chaperone HSP70 Binds to and Stabilizes NOD2, an Important Protein Involved in Crohn Disease*

    Science.gov (United States)

    Mohanan, Vishnu; Grimes, Catherine Leimkuhler

    2014-01-01

    Microbes are detected by the pathogen-associated molecular patterns through specific host pattern recognition receptors. Nucleotide-binding oligomerization domain-containing protein 2 (NOD2) is an intracellular pattern recognition receptor that recognizes fragments of the bacterial cell wall. NOD2 is important to human biology; when it is mutated it loses the ability to respond properly to bacterial cell wall fragments. To determine the mechanisms of misactivation in the NOD2 Crohn mutants, we developed a cell-based system to screen for protein-protein interactors of NOD2. We identified heat shock protein 70 (HSP70) as a protein interactor of both wild type and Crohn mutant NOD2. HSP70 has previously been linked to inflammation, especially in the regulation of anti-inflammatory molecules. Induced HSP70 expression in cells increased the response of NOD2 to bacterial cell wall fragments. In addition, an HSP70 inhibitor, KNK437, was capable of decreasing NOD2-mediated NF-κB activation in response to bacterial cell wall stimulation. We found HSP70 to regulate the half-life of NOD2, as increasing the HSP70 level in cells increased the half-life of NOD2, and down-regulating HSP70 decreased the half-life of NOD2. The expression levels of the Crohn-associated NOD2 variants were less compared with wild type. The overexpression of HSP70 significantly increased NOD2 levels as well as the signaling capacity of the mutants. Thus, our study shows that restoring the stability of the NOD2 Crohn mutants is sufficient for rescuing the ability of these mutations to signal the presence of a bacterial cell wall ligand. PMID:24790089

  18. KADAR MDA DAN HSP 70 PADA PLASENTA PENDERITA PREEKLAMPSIA

    Directory of Open Access Journals (Sweden)

    Arleni

    2008-12-01

    Full Text Available The MDA and HSP70 Concentration in Preeclamptic Patient Placenta’s. Objective: Preeclampsia is a disease in pregnancy and characterized by hypertension and proteinuria. Preeclampsia and eclampsia are the most causes of maternal and fetal mortality and morbidity in Indonesia. Placental and systemic oxidative stress caused endothelial cell dysfunction and injury. Placental oxidative stress also linked to fetal growth restriction. HSP70 is essential for cellular recovery, survival and maintenance of homeostasis. The purpose of this study was to compare the MDA, a marker for oxidative stress and HSP70 production in placental of severe preeclampsia, mild preeclampsia and normotensive pregnant women. Placenta were collected after delivery from normotensive pregnancies (N=10, severe preeclampsia (N=10 and mild preeclampsia (N=10. Placenta was cultured in RPMI and 20% FBS, and supernatant were collected in day 3. MDA was measured using spectrophotometer and absorbance read in 530nm. HSP70 was measured using enzyme-linked immunosorbent assay. The mean MDA concentration did not differ significantly between patients with severe preeclampsia (7.13+5.36 nmol/ml and mild preeclampsia (4.82+2.47 nmol/ml when compared with normotensive pregnancies (4.57+2.4 nmol/ml. The mean HSP70 concentration in mild preeclampsia is highest (10.15+12.39 nmo/ml when compared with severe preeclampsia (3.78 +3.07 nmol/ml and normotensive pregnant women (3.76+4.65nmol/ml, but the difference was not significant. Although the difference was not significant, is indicates homeostasis response in mild preclampsia women is relative good. This response was abated in severe preeclampstic women. Although MDA and HSP70 concentration did not differ significantly between groups, however the high HSP70 concentration is indicates homeostasis response relatively good in mild preeclamptic women.

  19. AAV-sBTLA facilitates HSP70 vaccine-triggered prophylactic antitumor immunity against a murine melanoma pulmonary metastasis model in vivo.

    Science.gov (United States)

    Han, Lingfei; Wang, Wei; Lu, Jiahong; Kong, Fanfei; Ma, Ge; Zhu, Yiping; Zhao, Dong; Zhu, Jianlong; Shuai, Wen; Zhou, Qian; Chen, Ping; Ye, Lei; Tao, Jie; Ahmad, Sarfraz; Li, Fang; Sun, Jing

    2014-11-28

    Activation of the BTLA-HVEM co-inhibitory signaling pathway impairs antitumor immunity. Our previous study demonstrated that the extracellular domain of murine BTLA (the soluble form of BTLA) can facilitate HSP70 vaccine-triggered antitumor immunity by blocking BTLA-HVEM interactions in a murine TC-1 non-metastatic tumor model. However, it is unknown whether this strategy has beneficial effects on highly malignant metastatic tumors, such as melanoma. To address this question, we expressed the soluble form of BTLA (sBTLA) in combination with HSP70 vaccine and examined the resulting antitumor activity in a melanoma pulmonary metastasis model. A recombinant adeno-associated virus (AAV) vector was used for the sBTLA gene delivery because of its high transfection efficiency and low toxicity. In vitro expression of AAV-sBTLA enhanced lymphocyte activation and induced specific cytotoxicity against B16F1 murine melanoma cells, while in vivo administration of AAV-sBTLA plus HSP70 vaccine by tail vein injection exerted a limited, late-stage antitumor effect against the existing B16F1 cells. However, the combination treatment generated a potent prophylactic antitumor response in the melanoma lung metastasis model in B6 mice. In this case, most of the metastatic foci were inhibited, and mouse survival was prolonged. Furthermore, the Th1 cytokines IL-2 and IFN-γ were up-regulated, while the negative regulatory molecules IL-10 and TGF-β were down-regulated. The number of regulatory T cells also decreased in the tumor environment. Therefore, AAV-sBTLA plus HSP70 vaccine may have therapeutic potential for the prevention of metastatic melanoma. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  20. Induction of hsp70 in the fungus Rhizopus nigricans.

    Science.gov (United States)

    Cernila, B; Cresnar, B; Breskvar, K

    1999-11-19

    We characterised two positive clones, RnH20/2 and RnH2/3, isolated from the cDNA library prepared from the fungus Rhizopus nigricans previously exposed to heat shock. Nucleotide sequences of both cDNA clones contained open reading frames for polypeptides with a significant amino acid identity to each other and to cytoplasmic members of Hsp70s from different eukaryotic organisms. Northern blot analysis, using a fragment of RnH2/3 as a probe, revealed that in the fungus R. nigricans the Hsp70 transcripts were inducible with deoxycorticosterone and testosterone as well as with heat stress, ethanol, CuSO(4), and H(2)O(2). This is the first report on isolation of cDNAs encoding Hsp70s from the fungal phylum Zygomycota. Copyright 1999 Academic Press.

  1. Metallothionein and Hsp70 trade-off against one another in Daphnia magna cross-tolerance to cadmium and heat stress

    Energy Technology Data Exchange (ETDEWEB)

    Haap, Timo, E-mail: timo.haap@gmx.de; Schwarz, Simon; Köhler, Heinz-R.

    2016-01-15

    Highlights: • Cadmium acclimation of two Daphnia magna clones which differed in Cd sensitivity and Hsp70 levels. • Two distinct metal-handling strategies regarding Hsp70 and MT expression were observed. • High Hsp70 levels did not confer an increase in Cd and heat stress tolerance. • Our results indicate a trade-off between Hsp70 and MT. - Abstract: The association between the insensitivity of adapted ecotypes of invertebrates to environmental stress, such as heavy metal pollution, and overall low Hsp levels characterizing these organisms has been attracting attention in various studies. The present study seeks to induce and examine this phenomenon in Daphnia magna by multigenerational acclimation to cadmium in a controlled laboratory setting. In this experiment, interclonal variation was examined: two clones of D. magna that have previously been characterized to diverge regarding their cadmium resistance and levels of the stress protein Hsp70, were continuously exposed to a sublethal concentration of Cd over four generations to study the effects of acclimation on Hsp70, metallothionein (MT), reproduction and cross-tolerance to heat stress. The two clones differed in all the measured parameters in a characteristic way, clone T displaying Cd and heat resistance, lower Hsp70 levels and offspring numbers on the one hand and higher MT expression on the other hand, clone S the opposite for all these parameters. We observed only slight acclimation-induced changes in constitutive Hsp70 levels and reproductive output. The differences in MT expression between clones as well as between acclimated organisms and controls give evidence for MT accounting for the higher Cd tolerance of clone T. Overall high Hsp70 levels of clone S did not confer cross tolerance to heat stress, contrary to common expectations. Our results suggest a trade-off between the efforts to limit the proteotoxic symptoms of Cd toxicity by Hsp70 induction and those to sequester and detoxify Cd by

  2. Hsp 70/Hsp 90 organizing protein as a nitrosylation target in cystic fibrosis therapy

    Science.gov (United States)

    Marozkina, Nadzeya V.; Yemen, Sean; Borowitz, Molly; Liu, Lei; Plapp, Melissa; Sun, Fei; Islam, Rafique; Erdmann-Gilmore, Petra; Townsend, R. Reid; Lichti, Cheryl F.; Mantri, Sneha; Clapp, Phillip W.; Randell, Scott H.; Gaston, Benjamin; Zaman, Khalequz

    2010-01-01

    The endogenous signaling molecule S-nitrosoglutathione (GSNO) and other S-nitrosylating agents can cause full maturation of the abnormal gene product ΔF508 cystic fibrosis (CF) transmembrane conductance regulator (CFTR). However, the molecular mechanism of action is not known. Here we show that Hsp70/Hsp90 organizing protein (Hop) is a critical target of GSNO, and its S-nitrosylation results in ΔF508 CFTR maturation and cell surface expression. S-nitrosylation by GSNO inhibited the association of Hop with CFTR in the endoplasmic reticulum. This effect was necessary and sufficient to mediate GSNO-induced cell-surface expression of ΔF508 CFTR. Hop knockdown using siRNA recapitulated the effect of GSNO on ΔF508 CFTR maturation and expression. Moreover, GSNO acted additively with decreased temperature, which promoted mutant CFTR maturation through a Hop-independent mechanism. We conclude that GSNO corrects ΔF508 CFTR trafficking by inhibiting Hop expression, and that combination therapies—using differing mechanisms of action—may have additive benefits in treating CF. PMID:20534503

  3. Hsp 70/Hsp 90 organizing protein as a nitrosylation target in cystic fibrosis therapy.

    Science.gov (United States)

    Marozkina, Nadzeya V; Yemen, Sean; Borowitz, Molly; Liu, Lei; Plapp, Melissa; Sun, Fei; Islam, Rafique; Erdmann-Gilmore, Petra; Townsend, R Reid; Lichti, Cheryl F; Mantri, Sneha; Clapp, Phillip W; Randell, Scott H; Gaston, Benjamin; Zaman, Khalequz

    2010-06-22

    The endogenous signaling molecule S-nitrosoglutathione (GSNO) and other S-nitrosylating agents can cause full maturation of the abnormal gene product DeltaF508 cystic fibrosis (CF) transmembrane conductance regulator (CFTR). However, the molecular mechanism of action is not known. Here we show that Hsp70/Hsp90 organizing protein (Hop) is a critical target of GSNO, and its S-nitrosylation results in DeltaF508 CFTR maturation and cell surface expression. S-nitrosylation by GSNO inhibited the association of Hop with CFTR in the endoplasmic reticulum. This effect was necessary and sufficient to mediate GSNO-induced cell-surface expression of DeltaF508 CFTR. Hop knockdown using siRNA recapitulated the effect of GSNO on DeltaF508 CFTR maturation and expression. Moreover, GSNO acted additively with decreased temperature, which promoted mutant CFTR maturation through a Hop-independent mechanism. We conclude that GSNO corrects DeltaF508 CFTR trafficking by inhibiting Hop expression, and that combination therapies--using differing mechanisms of action--may have additive benefits in treating CF.

  4. The 70 kD heat shock protein (hsp 70) in soil invertebrates: a possible tool for monitoring environmental toxicants.

    Science.gov (United States)

    Köhler, H R; Triebskorn, R; Stöcker, W; Kloetzel, P M; Alberti, G

    1992-04-01

    The expression of hsp 70 after heat shock or exposure to heavy metals/molluscicides was investigated by fluorography or immunoblot in three diplopods (Glomeris marginata, Cylindroiulus punctatus, Tachypodoiulus niger), two slugs (Deroceras reticulatum, Arion ater), and one isopod (Oniscus asellus). In O. asellus, hsp 70 expression occurred after heat shock and also after lead treatment, whereby a solution of 100 mg/kg Pb2+ was sufficient. Animals of the same species taken from a heavy metal polluted site in the vicinity of a lead/zinc smelter also showed the presence of hsp 70. The comparison of laboratory and field experiments demonstrated the suitability of O. asellus for monitoring tests. In contrast, the blot pattern after contamination with 1,000 mg/kg Pb2+ (in the mentioned diplopods) or different concentrations of the molluscicide Cloethocarb (BASF, FRG) (slugs) showed no differences compared to the respective control group.

  5. Methyl CpG level at distal part of heat-shock protein promoter HSP70 exhibits epigenetic memory for heat stress by modulating recruitment of POU2F1-associated nucleosome-remodeling deacetylase (NuRD) complex.

    Science.gov (United States)

    Kisliouk, Tatiana; Cramer, Tomer; Meiri, Noam

    2017-05-01

    Depending on its stringency, exposure to heat in early life leads to either resilience or vulnerability to heat stress later in life. We hypothesized that epigenetic alterations in genes belonging to the cell proteostasis pathways are attributed to long-term responses to heat stress. Epigenetic regulation of the mRNA expression of the molecular chaperone heat-shock protein (HSP) 70 (HSPA2) was evaluated in the chick hypothalamus during the critical period of thermal-control establishment on day 3 post-hatch and during heat challenge on day 10. Both the level and duration of HSP70 expression during heat challenge a week after heat conditioning were more pronounced in chicks conditioned under harsh versus mild temperature. Analyzing different segments of the promoter in vitro indicated that methylation of a distal part altered its transcriptional activity. In parallel, DNA-methylation level of this segment in vivo was higher in harsh- compared to mild-heat-conditioned chicks. Hypermethylation of the HSP70 promoter in high-temperature-conditioned chicks was accompanied by a reduction in both POU Class 2 Homeobox 1 (POU2F1) binding and recruitment of the nucleosome remodeling deacetylase (NuRD) chromatin-remodeling complex. As a result, histone H3 acetylation levels at the HSP70 promoter were higher in harsh-temperature-conditioned chicks than in their mild-heat-conditioned counterparts. These results suggest that methylation level of a distal part of the HSP70 promoter and POU2F1 recruitment may reflect heat-stress-related epigenetic memory and may be useful in differentiating between individuals that are resilient or vulnerable to stress. © 2017 International Society for Neurochemistry.

  6. Non-invasive assessment of animal exercise stress: real-time PCR of GLUT4, COX2, SOD1 and HSP70 in avalanche military dog saliva.

    Science.gov (United States)

    Diverio, S; Guelfi, G; Barbato, O; Di Mari, W; Egidi, M G; Santoro, M M

    2015-01-01

    Exercise has been shown to increase mRNA expression of a growing number of genes. The aim of this study was to assess if mRNA expression of the metabolism- and oxidative stress-related genes GLUT4 (glucose transporter 4), COX2 (cyclooxygenase 2), SOD1 (superoxide dismutase 1) and HSP70 (heat shock protein 70) in saliva changes following acute exercise stress in dogs. For this purpose, 12 avalanche dogs of the Italian Military Force Guardia di Finanza were monitored during simulation of a search for a buried person in an artificial avalanche area. Rectal temperature (RT) and saliva samples were collected the day before the trial (T0), immediately after the descent from a helicopter at the onset of a simulated avalanche search and rescue operation (T1), after the discovery of the buried person (T2) and 2 h later (T3). Expressions of GLUT4, SOD1, COX2 and HSP70 were measured by real-time PCR. The simulated avalanche search and rescue operation was shown to exert a significant effect on RT, as well as on the expression of all metabolism- and oxidative stress-related genes investigated, which peaked at T2. The observed expression patterns indicate an acute exercise stress-induced upregulation, as confirmed by the reductions in expression at T3. Moreover, our findings indicate that saliva is useful for assessing metabolism- and oxidative stress-related genes without the need for restraint, which could affect working dog performance.

  7. Function of SSA subfamily of Hsp70 within and across species varies widely in complementing Saccharomyces cerevisiae cell growth and prion propagation.

    Directory of Open Access Journals (Sweden)

    Deepak Sharma

    2009-08-01

    Full Text Available The cytosol of most eukaryotic cells contains multiple highly conserved Hsp70 orthologs that differ mainly by their spatio-temporal expression patterns. Hsp70s play essential roles in protein folding, transport or degradation, and are major players of cellular quality control processes. However, while several reports suggest that specialized functions of Hsp70 orthologs were selected through evolution, few studies addressed systematically this issue.We compared the ability of Ssa1p-Ssa4p from Saccharomyces cerevisiae and Ssa5p-Ssa8p from the evolutionary distant yeast Yarrowia lipolytica to perform Hsp70-dependent tasks when expressed as the sole Hsp70 for S. cerevisiae in vivo. We show that Hsp70 isoforms (i supported yeast viability yet with markedly different growth rates, (ii influenced the propagation and stability of the [PSI(+] and [URE3] prions, but iii did not significantly affect the proteasomal degradation rate of CFTR. Additionally, we show that individual Hsp70 orthologs did not induce the formation of different prion strains, but rather influenced the aggregation properties of Sup35 in vivo. Finally, we show that [URE3] curing by the overexpression of Ydj1p is Hsp70-isoform dependent.Despite very high homology and overlapping functions, the different Hsp70 orthologs have evolved to possess distinct activities that are required to cope with different types of substrates or stress situations. Yeast prions provide a very sensitive model to uncover this functional specialization and to explore the intricate network of chaperone/co-chaperone/substrates interactions.

  8. Proteotoxicity is not the reason for the dependence of cancer cells on the major chaperone Hsp70.

    Science.gov (United States)

    Colvin, Teresa A; Gabai, Vladimir L; Sherman, Michael Y

    2014-01-01

    Several years ago a hypothesis was proposed that the survival of cancer cells depend on elevated expression of molecular chaperones because these cells are prone to proteotoxic stress. A critical prediction of this hypothesis is that depletion of chaperones in cancer cells should lead to proteotoxicity. Here, using the major chaperone Hsp70 as example, we demonstrate that its depletion does not trigger proteotoxic stress, thus refuting the model. Accordingly, other functions of chaperones, e.g., their role in cell signaling, might define the requirements for chaperones in cancer cells, which is critical for rational targeting Hsp70 in cancer treatment.

  9. Isolation, partial length sequence and expression of steroid inducible hps 70 gene from Rhizopus nigricans.

    Science.gov (United States)

    Cernila, B; Cresnar, B; Breskvar, K

    2000-01-01

    Previous studies have shown that filamentous fungus Rhizopus nigricans responds to addition of different steroids into growth medium with induction of hydroxylation system and that some steroids provoke stress response. The purpose of this study was to investigate whether those steroids provoke induction of Hsp70 gene(s), well studied markers of stress response in different cells and organisms. The expression studies of fungal Hsp70 gene(s) using Northern blot analysis showed that fungal hsp70 mRNA was upregulated after treatment of mycelia with deoxycorticosterone and testosterone, but not after exposure to progesterone. In addition, expression of fungal Hsp70 mRNA was elevated after exposure of mycelia to heat shock (32 degrees C), ethanol, heavy metal (CuSO4), and oxidative stressor (H2O2), whereas treatment of mycelia with osmotic stressor (KCl) didn't have any influence on stress protein expression. The partial nucleotide sequence and deduced amino acid sequence homology search revealed that the cDNA clone (lambda hs20/2), isolated from cDNA library prepared from heat shock treated fungal mycelia, contained Hsp70 gene of DnaK subfamily.

  10. 70 kD stress protein (Hsp70) analysis in living shallow-water benthic foraminifera

    Digital Repository Service at National Institute of Oceanography (India)

    Heinz, P.; Marten, R.A; Linshy, V.N; Haap, T.; Geslin, E.; Kohler, H-R.

    Hsp70 is a phylogenetically highly conserved protein family present in all eukaryotic organisms tested so far. Its synthesis is induced by proteotoxic stress. The detection of Hsp70 in foraminifera is presented here. We introduce a standard...

  11. The preparation of HL-60 cells vaccine expressing BCG heat shock protein 70 and detection of its immunogenicity in vitro.

    Science.gov (United States)

    Li, Xiao-Ling; Zhao, Yan-Xia; Sun, Li-Rong; Yang, Jing; Xu, Hui-Juan

    2012-10-01

    Gene-modified cell vaccines are the best way to achieve the immunotherapy for all types of acute leukemia. In this study, the recombinant eukaryotic expression vector (pDisplay-HSP70) of heat shock protein 70 (HSP70) of Bacille Calmette-Guérin (BCG) was constructed by amplifying the whole BCG HSP70 gene using polymerase chain reaction (PCR) and sub-cloning into the polyclone endonuclease sites in pDisplay. Then the HL-60 cell vaccine expressing the protein onto the cell surface was prepared by lipofectamine transfection and its anti-tumor effect and mechanism were further studied. Results showed that the fragment of BCG HSP70 was consistent with Mycobacterium tuberculosis HSP70 gene published in GeneBank. DNA sequencing showed that the recombinant vector was correctly constructed and named pDisplay-HSP70. After BCG HSP70 gene transfection, the yellow-green fluorescence on the HL-60 cells surface was observed under a fluorescence microscope. The immunogenicity of HSP70-transfected HL-60 cells exhibited upregulated proliferation of lymphocytes, increased cytokine secretion (IFN-γ) and enhanced killing activity. These results suggested that gene transfection of BCG HSP70 could significantly enhance the immunogenicity of HL-60 cells. It may be used as a suitable candidate gene-modified cell vaccine for cancer immunotherapy.

  12. Metabolic fates and effects of nitrite in brown trout under normoxic and hypoxic conditions: blood and tissue nitrite metabolism and interactions with branchial NOS, Na+/K+-ATPase and hsp70 expression

    DEFF Research Database (Denmark)

    Jensen, Frank Bo; Gerber, Lucie; Hansen, Marie Niemann

    2015-01-01

    Nitrite secures essential nitric oxide (NO) bioavailability in hypoxia at low endogenous concentrations, whereas it becomes toxic at high concentrations. We exposed brown trout to normoxic and hypoxic water in the absence and presence of added ambient nitrite to decipher the cellular metabolism...... and effects of nitrite at basal and elevated concentrations under different oxygen regimes. We also tested hypotheses concerning the influence of nitrite on branchial nitric oxide synthase (NOS), Na+/K+-ATPase (nka) and heat shock protein (hsp70) mRNA expression. Basal plasma and erythrocyte nitrite levels...... were higher in hypoxia than normoxia, suggesting increased NOS activity. Nitrite exposure strongly elevated nitrite concentrations in plasma, erythrocytes, heart tissue and white muscle, which was associated with an extensive metabolism of nitrite to nitrate and to iron-nitrosylated and S...

  13. The role of Hsp70 in oxi-inflamm-aging and its use as a potential biomarker of lifespan.

    Science.gov (United States)

    Martínez de Toda, I; De la Fuente, M

    2015-12-01

    The heat-shock protein 70 (HSPA1A or Hsp70) acts as a cellular defense mechanism its expression being induced under stressful conditions. Aging has been related to an impairment in this induction. However, an extended longevity has been associated with its increased expression. According to the oxidation-inflammation theory of aging, chronic oxidative stress and inflammatory stress situations (with higher levels of oxidant and inflammatory compounds and lower antioxidant and anti-inflammatory defenses) are the basis of the age-related alterations of body cells. Since oxidation and inflammation are interlinked processes, and Hsp70 has been shown to confer protection against the harmful effects of oxidative stress as well as modulating the inflammatory status, it could play a role as a regulator of the rate of aging. This role may be different in mitotic and post-mitotic tissues due to the differences in their age-related mechanisms of response, such as apoptosis. Mechanisms affected by Hsp70 that can interfere with the deleterious effects of excessive oxidative stress and chronic low-grade inflammation and that are closely related to the aging process have been detailed. In addition, the potential use of the basal levels (with their differences in post-mitotic and mitotic tissues), the inducible levels, as well as the extracellular levels of Hsp70 as possible biomarkers of the rate of aging and lifespan, have also been discussed.

  14. Reactive oxygen species generated by a heat shock protein (Hsp) inducing product contributes to Hsp70 production and Hsp70-mediated protective immunity in Artemia franciscana against pathogenic vibrios.

    Science.gov (United States)

    Baruah, Kartik; Norouzitallab, Parisa; Linayati, Linayati; Sorgeloos, Patrick; Bossier, Peter

    2014-10-01

    The cytoprotective role of heat shock protein (Hsp70) described in a variety of animal disease models, including vibriosis in farmed aquatic animals, suggests that new protective strategies relying upon the use of compounds that selectively turn on Hsp genes could be developed. The product Tex-OE® (hereafter referred to as Hspi), an extract from the skin of the prickly pear fruit, Opuntia ficus indica, was previously shown to trigger Hsp70 synthesis in a non-stressful situation in a variety of animals, including in a gnotobiotically (germ-free) cultured brine shrimp Artemia franciscana model system. This model system offers great potential for carrying out high-throughput, live-animal screens of compounds that have health benefit effects. By using this model system, we aimed to disclose the underlying cause behind the induction of Hsp70 by Hspi in the shrimp host, and to determine whether the product affects the shrimp in inducing resistance towards pathogenic vibrios. We provide unequivocal evidences indicating that during the pretreatment period with Hspi, there is an initial release of reactive oxygen species (hydrogen peroxide and/or superoxide anion), generated by the added product, in the rearing water and associated with the host. The reactive molecules generated are the triggering factors responsible for causing Hsp70 induction within Artemia. We have also shown that Hspi acts prophylactically at an optimum dose regimen to confer protection against pathogenic vibrios. This salutary effect was associated with upregulation of two important immune genes, prophenoloxidase and transglutaminase of the innate immune system. These findings suggest that inducers of stress protein (e.g. Hsp70) are potentially important modulator of immune responses and might be exploited to confer protection to cultured shrimp against Vibrio infection. Copyright © 2014 Elsevier Ltd. All rights reserved.

  15. Variation in Hsp70 levels after cold shock: signs of evolutionary responses to thermal selection among Leptinotarsa decemlineata populations.

    Directory of Open Access Journals (Sweden)

    Anne Lyytinen

    Full Text Available Individuals of widely spread species are expected to show local adaption in temperature tolerance as they encounter a range of thermal conditions. We tracked thermal adaptations of the Colorado potato beetle (Leptinotarsa decemlineata that invaded Europe within the last 100 years. It has occupied various conditions although, like the majority of invasive species, it lost a measurable amount of neutral genetic variation due to bottleneck effect when it invaded Europe. We exposed diapausing beetles originated from three different latitudes (54°N, 59°N, 60°N to cold shock (-5°C, 1.5 hrs in order to test if beetles from the northern populations express differential levels of cold-induced and constitutive Hsp70 compared to the beetles from milder temperature regime. The level of cold-induced Hsp70 was lowest in the northernmost beetle populations while the level of constitutive Hsp70 did not differ with the population. Moreover, the southernmost beetles were more plastic in their response to cold shock than the northernmost beetles. These results suggest that physiological adaptation, like the synthesis of Hsp70, can evolve very quickly.

  16. Immunogenicity of HSP-70, KMP-11 and PFR-2 leishmanial antigens in the experimental model of canine visceral leishmaniasis.

    Science.gov (United States)

    Carrillo, Eugenia; Crusat, Martín; Nieto, Javier; Chicharro, Carmen; Thomas, Maria del Carmen; Martínez, Enrique; Valladares, Basilio; Cañavate, Carmen; Requena, Jose María; López, Manuel Carlos; Alvar, Jorge; Moreno, Javier

    2008-03-28

    Zoonotic visceral leishmaniasis (ZVL) is a parasitic disease caused by Leishmania infantum/L. chagasi that is emerging as an important medical and veterinary problem. Dogs are the domestic reservoir for this parasite and, therefore, the main target for controlling the transmission to humans. In the present work, we have evaluated the immunogenicity of the Leishmania infantum heat shock protein (HSP)-70, paraflagellar rod protein (PFR)-2 and kinetoplastida membrane protein (KMP)-11 recombinant proteins in dogs experimentally infected with the parasite. We have shown that peripheral blood mononuclear cells (PBMC) from experimentally infected dogs proliferated in response to these recombinant antigens and against the soluble leishmanial antigen (SLA). We have also quantified the mRNA expression level of the cytokines induced in PBMC upon stimulation with the HSP-70, PFR-2 and KMP-11 proteins. These recombinant proteins induced an up-regulation of IFN-gamma. HSP-70 and PFR-2 also produced an increase of the TNF-alpha transcripts abundance. No measurable induction of IL-10 was observed and low levels of IL-4 mRNA were produced in response to the three mentioned recombinant antigens. Serum levels of specific antibodies against HSP-70, PFR-2 and KMP-11 recombinant proteins were also determined in these animals. Our study showed that HSP-70, KMP-11 and PFR-2 proteins are recognized by infected canines. Furthermore, these antigens produce a Th1-type immune response, suggesting that they may be involved in protection. The identification as vaccine candidates of Leishmania antigens that elicit appropriate immune responses in the canine model is a key step in the rational approach to generate a vaccine for canine visceral leishmaniasis.

  17. Genetic modification of cold-preserved renal grafts using HSP70 or bcl-2 HVJ-liposome method.

    Science.gov (United States)

    Kita, Junji; Kobayashi, Eiji; Hishinuma, Akira; Kaneda, Yasufumi

    2003-01-01

    We tested the hypothesis that the best time for genetic modification is while the cell viability of the graft is reduced for long-term preservation. The hemagglutinating virus of Japan (HVJ)-liposome method, a nonviral gene transfer technique, was used with a luciferase gene to test the efficacy of protein induction under the critical preservation time. Furthermore, we tested this genetic modification with heat shock protein (HSP) 70 or bcl-2 genes to prevent primary nonfunction (PNF) after long-term preservation. Orthotopic rat renal transplantation (RT) was performed using the cuff technique in the syngeneic combination of Lew (major histocompatible complex, haplotype: RT1(l)). Rat kidney grafts were preserved for 24 or 48 h in University of Wisconsin (UW) or Ringer's lactate solution using HVJ method with the luciferase gene. Rats with gene-transfected kidneys were re-laparotomized 48 h after transplantation to estimate the lack of arterial flow in the graft and killed for histological evaluation of the degree of PNF luciferase intensity assay. Then, two functional genes (HSP70 or bcl-2) were tested for the occurrence of PNF and histological and immunohistochemical analysis of the grafted kidneys preserved for 48 h in the UW solution. In the kidneys preserved for 24 h, 50% of the Ringer's lactate group had PNF; but all of the UW group had sufficient blood flow. The graft viability was well corrected by the degree of luciferase intensity. The PNF rate was significantly suppressed in the bcl-2 gene-transfer group, and tended to be reduced in the HSP70 group. The HVJ-liposome method effectively induced the foreign gene for kidney grafts even in the cold-preservation solution. Induction of bcl-2 or the HSP70 gene reduced the occurrence of PNF in the rat renal graft. The results suggest that gene transfer not only maintains graft viability, but also graft activation.

  18. The Hsp70/Hsp90 Chaperone Machinery in Neurodegenerative Diseases

    Science.gov (United States)

    Lackie, Rachel E.; Maciejewski, Andrzej; Ostapchenko, Valeriy G.; Marques-Lopes, Jose; Choy, Wing-Yiu; Duennwald, Martin L.; Prado, Vania F.; Prado, Marco A. M.

    2017-01-01

    The accumulation of misfolded proteins in the human brain is one of the critical features of many neurodegenerative diseases, including Alzheimer's disease (AD). Assembles of beta-amyloid (Aβ) peptide—either soluble (oligomers) or insoluble (plaques) and of tau protein, which form neurofibrillary tangles, are the major hallmarks of AD. Chaperones and co-chaperones regulate protein folding and client maturation, but they also target misfolded or aggregated proteins for refolding or for degradation, mostly by the proteasome. They form an important line of defense against misfolded proteins and are part of the cellular quality control system. The heat shock protein (Hsp) family, particularly Hsp70 and Hsp90, plays a major part in this process and it is well-known to regulate protein misfolding in a variety of diseases, including tau levels and toxicity in AD. However, the role of Hsp90 in regulating protein misfolding is not yet fully understood. For example, knockdown of Hsp90 and its co-chaperones in a Caenorhabditis elegans model of Aβ misfolding leads to increased toxicity. On the other hand, the use of Hsp90 inhibitors in AD mouse models reduces Aβ toxicity, and normalizes synaptic function. Stress-inducible phosphoprotein 1 (STI1), an intracellular co-chaperone, mediates the transfer of clients from Hsp70 to Hsp90. Importantly, STI1 has been shown to regulate aggregation of amyloid-like proteins in yeast. In addition to its intracellular function, STI1 can be secreted by diverse cell types, including astrocytes and microglia and function as a neurotrophic ligand by triggering signaling via the cellular prion protein (PrPC). Extracellular STI1 can prevent Aβ toxic signaling by (i) interfering with Aβ binding to PrPC and (ii) triggering pro-survival signaling cascades. Interestingly, decreased levels of STI1 in C. elegans can also increase toxicity in an amyloid model. In this review, we will discuss the role of intracellular and extracellular STI1 and the

  19. The Hsp70/Hsp90 Chaperone Machinery in Neurodegenerative Diseases

    Directory of Open Access Journals (Sweden)

    Rachel E. Lackie

    2017-05-01

    Full Text Available The accumulation of misfolded proteins in the human brain is one of the critical features of many neurodegenerative diseases, including Alzheimer's disease (AD. Assembles of beta-amyloid (Aβ peptide—either soluble (oligomers or insoluble (plaques and of tau protein, which form neurofibrillary tangles, are the major hallmarks of AD. Chaperones and co-chaperones regulate protein folding and client maturation, but they also target misfolded or aggregated proteins for refolding or for degradation, mostly by the proteasome. They form an important line of defense against misfolded proteins and are part of the cellular quality control system. The heat shock protein (Hsp family, particularly Hsp70 and Hsp90, plays a major part in this process and it is well-known to regulate protein misfolding in a variety of diseases, including tau levels and toxicity in AD. However, the role of Hsp90 in regulating protein misfolding is not yet fully understood. For example, knockdown of Hsp90 and its co-chaperones in a Caenorhabditis elegans model of Aβ misfolding leads to increased toxicity. On the other hand, the use of Hsp90 inhibitors in AD mouse models reduces Aβ toxicity, and normalizes synaptic function. Stress-inducible phosphoprotein 1 (STI1, an intracellular co-chaperone, mediates the transfer of clients from Hsp70 to Hsp90. Importantly, STI1 has been shown to regulate aggregation of amyloid-like proteins in yeast. In addition to its intracellular function, STI1 can be secreted by diverse cell types, including astrocytes and microglia and function as a neurotrophic ligand by triggering signaling via the cellular prion protein (PrPC. Extracellular STI1 can prevent Aβ toxic signaling by (i interfering with Aβ binding to PrPC and (ii triggering pro-survival signaling cascades. Interestingly, decreased levels of STI1 in C. elegans can also increase toxicity in an amyloid model. In this review, we will discuss the role of intracellular and extracellular

  20. 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.

  1. Unraveling the CHIP:Hsp70 complex as an information processor for protein quality control.

    Science.gov (United States)

    VanPelt, Jamie; Page, Richard C

    2017-02-01

    The CHIP:Hsp70 complex stands at the crossroads of the cellular protein quality control system. Hsp70 facilitates active refolding of misfolded client proteins, while CHIP directs ubiquitination of misfolded client proteins bound to Hsp70. The direct competition between CHIP and Hsp70 for the fate of misfolded proteins leads to the question: how does the CHIP:Hsp70 complex execute triage decisions that direct misfolded proteins for either refolding or degradation? The current body of literature points toward action of the CHIP:Hsp70 complex as an information processor that takes inputs in the form of client folding state, dynamics, and posttranslational modifications, then outputs either refolded or ubiquitinated client proteins. Herein we examine the CHIP:Hsp70 complex beginning with the structure and function of CHIP and Hsp70, followed by an examination of recent studies of the interactions and dynamics of the CHIP:Hsp70 complex. Copyright © 2016 Elsevier B.V. All rights reserved.

  2. Human skeletal muscle HSP70 response to physical training depends on exercise intensity.

    Science.gov (United States)

    Liu, Y; Lormes, W; Baur, C; Opitz-Gress, A; Altenburg, D; Lehmann, M; Steinacker, J M

    2000-07-01

    We have previously reported that HSP70 in human skeletal muscle could be induced by training. However, whether HSP70 induction is dependent upon exercise volume or exercise intensity remains unknown. The aim of the present study was to investigate the relationship between HSP70 and training intensity in rowers. Fourteen well-trained male rowers were divided into two groups (group A, n = 6; group B, n = 8). Group A performed higher intensity exercise during 1st phase, whereas group B performed higher intensity exercise during 2nd training phase. Training volume in 2nd phase increased in both groups. Both training intensity and volume were reduced in 3rd phase. Muscle samples were taken from m. vastus lateralis by fine needle biopsy before training, at the end of the 1st, 2nd and 3rd training phases. HSP70 was quantitatively determined using SDS-PAGE with silver stain. In group A, HSP70 increased significantly from 38 +/- 12 etag before training to 59 +/- 16 etag at the end of the lst training phase (loaded total protein 2.5microg), and decreased afterwards. In group B, HSP70 increase (from 36 +/- 11 etag to 50 +/- 13 etag) in the 1st phase was significantly smaller, there was a further increase of HSP70 in the 2nd phase (60 +/- 14 etag). At the end of the training, HSP70 decreased in both groups. Thus, HSP70 response to training seems to be dependent upon exercise intensity.

  3. Fine particulate matter potentiates type 2 diabetes development in high-fat diet-treated mice: stress response and extracellular to intracellular HSP70 ratio analysis.

    Science.gov (United States)

    Goettems-Fiorin, Pauline Brendler; Grochanke, Bethânia Salamoni; Baldissera, Fernanda Giesel; Dos Santos, Analu Bender; Homem de Bittencourt, Paulo Ivo; Ludwig, Mirna Stela; Rhoden, Claudia Ramos; Heck, Thiago Gomes

    2016-12-01

    Exposure to fine particulate matter (PM2.5) air pollution is a risk factor for type 2 diabetes (T2DM). We argue whether the potentiating effect of PM2.5 over the development of T2DM in high-fat diet (HFD)-fed mice would be related to modification in cell stress response, particularly in antioxidant defenses and 70-kDa heat shock proteins (HSP70) status. Male mice were fed standard chow or HFD for 12 weeks and then randomly exposed to daily nasotropic instillation of PM2.5 for additional 12 weeks under the same diet schedule, divided into four groups (n = 14-15 each): Control, PM2.5, HFD, and HFD + PM2.5 were evaluated biometric and metabolic profiles of mice, and cellular stress response (antioxidant defense and HSP70 status) of metabolic tissues. Extracellular to intracellular HSP70 ratio ([eHSP72]/[iHSP70]), viz. H-index, was then calculated. HFD + PM2.5 mice presented a positive correlation between adiposity, increased body weight and glucose intolerance, and increased glucose and triacylglycerol plasma levels. Pancreas exhibited lower iHSP70 expression, accompanied by 3.7-fold increase in the plasma to pancreas [eHSP72]/[iHSP70] ratio. Exposure to PM2.5 markedly potentiated metabolic dysfunction in HFD-treated mice and promoted relevant alteration in cell stress response assessed by [eHSP72]/[iHSP70], a relevant biomarker of chronic low-grade inflammatory state and T2DM risk.

  4. Quantitative assessment of hsp70, IL-1β and TNF-α in the spinal cord of dogs with E40K SOD1-associated degenerative myelopathy.

    Science.gov (United States)

    Lovett, M C; Coates, J R; Shu, Y; Oglesbee, M J; Fenner, W; Moore, S A

    2014-05-01

    Inflammation is involved in the pathogenesis of many neurodegenerative diseases. Canine degenerative myelopathy (DM) is a progressive adult-onset neurodegenerative disease commonly associated with an E40K missense mutation in the SOD1 gene. DM has many similarities to some familial forms of human amyotrophic lateral sclerosis (ALS) and may serve as an important disease model for therapy development. Pro-inflammatory mediators such as interleukin (IL)-1β, tumor necrosis factor (TNF)-α and heat shock protein (hsp) 70 play a role in the pathogenesis of ALS. The focus of the current work was to determine whether an inflammatory phenotype is present in canine DM as defined by IL-1β, TNF-α, and hsp70 responses in cerebrospinal fluid (CSF) and spinal cord tissue. Concentrations of hsp70, IL-1β and TNF-α were below the limits of detection by ELISA in the CSF of both normal and DM-affected dogs. Immunohistochemical staining for hsp70 was significantly increased in ependymal cells lining the spinal cord central canal of DM-affected dogs (P = 0.003). This was not associated with increased IL-1β or TNF-α staining, but was associated with increased CD18 staining in the gray matter of DM-affected dogs. These results suggest that hsp70 in spinal cord tissue is a potential inflammatory signature in canine DM. Copyright © 2014 Elsevier Ltd. All rights reserved.

  5. Congenital sideroblastic anemia due to mutations in the mitochondrial HSP70 homologue HSPA9.

    Science.gov (United States)

    Schmitz-Abe, Klaus; Ciesielski, Szymon J; Schmidt, Paul J; Campagna, Dean R; Rahimov, Fedik; Schilke, Brenda A; Cuijpers, Marloes; Rieneck, Klaus; Lausen, Birgitte; Linenberger, Michael L; Sendamarai, Anoop K; Guo, Chaoshe; Hofmann, Inga; Newburger, Peter E; Matthews, Dana; Shimamura, Akiko; Snijders, Pieter J L M; Towne, Meghan C; Niemeyer, Charlotte M; Watson, Henry G; Dziegiel, Morten H; Heeney, Matthew M; May, Alison; Bottomley, Sylvia S; Swinkels, Dorine W; Markianos, Kyriacos; Craig, Elizabeth A; Fleming, Mark D

    2015-12-17

    The congenital sideroblastic anemias (CSAs) are relatively uncommon diseases characterized by defects in mitochondrial heme synthesis, iron-sulfur (Fe-S) cluster biogenesis, or protein synthesis. Here we demonstrate that mutations in HSPA9, a mitochondrial HSP70 homolog located in the chromosome 5q deletion syndrome 5q33 critical deletion interval and involved in mitochondrial Fe-S biogenesis, result in CSA inherited as an autosomal recessive trait. In a fraction of patients with just 1 severe loss-of-function allele, expression of the clinical phenotype is associated with a common coding single nucleotide polymorphism in trans that correlates with reduced messenger RNA expression and results in a pseudodominant pattern of inheritance. © 2015 by The American Society of Hematology.

  6. Modulation of polyglutamine inclusion formation by the Hsp70 chaperone machine

    NARCIS (Netherlands)

    Rujano, M. A.; Kampinga, H. H.; Salomons, F. A.

    2007-01-01

    Components of the Hsp70 chaperone machine have been implied in protection against polyglutamine (poly-Q) pathologies. Yet, little is known about specific mechanisms and the rate-limiting components that account for this protective effect. Here, we examined the effects of an Hsp70 chaperone family

  7. Chaperone proteostasis in Parkinson's disease : stabilization of the Hsp70/alpha-synuclein complex by Hip

    NARCIS (Netherlands)

    Roodveldt, Cintia; Bertoncini, Carlos W.; Andersson, August; van der Goot, Annemieke T.; Hsu, Shang-Te; Fernandez-Montesinos, Rafael; de Jong, Jannie; van Ham, Tjakko J.; Nollen, Ellen A.; Pozo, David; Christodoulou, John; Dobson, Christopher M.

    2009-01-01

    The ATP-dependent protein chaperone heat-shock protein 70 (Hsp70) displays broad anti-aggregation functions and has a critical function in preventing protein misfolding pathologies. According to in vitro and in vivo models of Parkinson's disease (PD), loss of Hsp70 activity is associated with

  8. The Role of Hsp70 in the Regulation of Autophagy in Gametogenesis, Pregnancy, and Parturition.

    Science.gov (United States)

    Witkin, Steven S; Kanninen, Tomi T; Sisti, Giovanni

    2017-01-01

    Induction of the 70 kDa heat shock protein (hsp70) and autophagy are two major mechanisms that promote cell homeostasis during the rapid cell growth and differentiation characteristic of reproduction. Hsp70 insures proper assembly, conformation, and intracellular transport of nascent proteins. Autophagy removes from the cytoplasm proteins, other macromolecules, and organelles that are no longer functional or needed and recycles their components for synthesis of new products under nutritionally limiting conditions. Hsp70 inhibits autophagy and so a proper balance between these two processes is essential for optimal germ cell production and survival and pregnancy progression. A marked inhibition in autophagy and a concomitant increase in hsp70 at term is a trigger for parturition. Excessive external or endogenous stress that induces a high level of hsp70 production can lead to a non-physiological inhibition of autophagy, resulting in altered spermatogenesis, premature ovarian failure, and complications of pregnancy including preeclampsia, intrauterine growth restriction, and preterm birth.

  9. Heat-shock protein (HSP70-2) allelic frequencies in three distinct Mexican populations.

    Science.gov (United States)

    Zúñiga, J; Vargas-Alarcón, G; Osnaya, N; Martínez-Tripp, S; Rodríguez-Reyna, T S; Hernández-Martinez, B; Hesiquio, R; Hernández-Pacheco, G; Gamboa, R; Juárez, F; Salgado, N; Granados, J

    1999-09-01

    The major histocompatibility complex (MHC) genes are highly polymorphic and therefore have been useful in population genetics and disease association studies. We analyzed restriction fragment length polymorphism of HSP70-2 alleles in healthy unrelated Mestizo, Mazatecan and Nahua populations. Both Indian groups, Mazatecans and Nahuas, were in Hardy-Weinberg equilibrium, while Mestizos were in disequilibrium (chi 2 = 0.399; P < 0.05). The Mazatecan Indians presented a high frequency of BB homozygosity (17.35%) compared to Mestizos (5%) (P = 0.01). Mexican ethnic groups present differences in distribution of BB genotype. The low frequency of BB genotype in Mestizos may be the result of a negative selection process.

  10. Serum levels of HSP70 and other DAMP proteins can aid in patient diagnosis after traumatic injury.

    Science.gov (United States)

    Ren, Biqiong; Zou, Guoying; Huang, Yiran; Xu, Guofeng; Xu, Fei; He, Junyu; Zhu, Haowen; Yu, Ping

    2016-07-01

    Danger-associated molecular patterns (DAMPs) are activated by endogenous signals that originate from stressed, injured, or necrotic cells, signifying "danger" to the host. In this study, we evaluated the expression of the DAMP heat shock protein 70 (HSP70) in trauma patients with and without secondary infections. Levels of glucose (GLU), procalcitonin (PCT), total cholesterol (T-Chol), and white blood cell (WBC) counts were also evaluated at three time stages after trauma. Our analysis showed that the levels of serum HSP70 in patients with minor, moderate, and severe injuries were significantly higher than in healthy patients at each time point post-injury (P < 0.01), and levels of serum HSP70 in the severe injury group were significantly higher than in the minor injury group at 1-6 h after trauma (P = 0.047). HSP70 was correlated with GLU and was negatively correlated with T-Chol in the period 1-6 h after injury (P = 0.008/0.032). WBC and GLU were elevated after trauma, with mutual positive correlation (P < 0.001). PCT levels increased later than WBC counts and GLU levels; these levels were correlated at the two later time periods, 24-48 h and 60-90 h (P = 0.008/0.041). PCT continued to rise in patients with secondary infection, but PCT dropped at the third time period in patients without secondary infection. In summary, our results suggest that danger and stress theory can be used to predict severity of trauma.

  11. Allicin protects traumatic spinal cord injury through regulating the HSP70/Akt/iNOS pathway in mice.

    Science.gov (United States)

    Wang, Shunyi; Ren, Dongliang

    2016-10-01

    Allicin is a major component of garlic, extracted as an oily liquid. The present study was designed to investigate the beneficial effects of allicin on traumatic spinal cord injury (TSCI) in mice, and whether the effects are mediated via regulation of the heat shock protein 70 (HSP70), v‑akt murine thymoma viral oncogene homolog 1 (Akt) and inducible nitric oxide synthase (iNOS) pathways. Adult BALB/c mice (30‑40 g) received a laminectomy at the T9 vertebral level as a model of TSCI. In the present study, treatment of the TSCI mice with allicin significantly increased their Basso, Beattie and Bresnahan (BBB) scores (Pallicin increased the protein levels of HSP70, increased the phosphorylation of Akt and reduced the iNOS protein expression levels in TSCI mice. Additionally, treatment with allicin significantly reduced the levels of ROS and enhanced the NADH levels in TSCI mice. Collectively, these data demonstrate that the effects of allicin on TSCI are mediated via regulation of the HSP70, Akt and iNOS pathways in mice.

  12. Effects of rapid temperature changes on HK, PK and HSP70 of Litopenaeus vannamei in different seasons

    Science.gov (United States)

    Guo, Biao; Wang, Fang; Dong, Shuanglin; Hou, Chunqiang

    2010-09-01

    Activities of hexokinase (HK), pyruvate kinase (PK) and levels of HSP70 were measured to evaluate the response of Litopenaeus vannamei to rapid temperature changes under controlled laboratory conditions. Shrimps were subjected to a quick temperature change from 27°C to 17°C for the summer case (Cold temperature treatment), or from 17°C to 27°C for the winter case (Warm temperature treatment). After 0.5, 1, 3, 6, 12, 24, 48, and 72 h of exposure time, shrimps were sampled and prepared for further analysis. The results showed that the effect of acute temperature changes on activities of HK was significant. Patterns of variations of the two glycolytic enzymes suggested that enzymes in the glycolysis cycle could adjust their activities to meet the acute temperature change. The HSP70 level increased in both cold and warm temperature treatments, suggesting that the rapid temperature changes activated the process of body’s self-protection. But the difference in expression peak of HSP70 might be related to the different body size and the higher thermal sensitivity to temperature increase than to temperature decrease of L. vannamei.

  13. Construction and prokaryotic expression of the fusion gene PRRSV ...

    African Journals Online (AJOL)

    ajl4

    2013-07-24

    Jul 24, 2013 ... pathology mechanisms of autoimmune diseases (Koets et al., 1999; Wong, 1999; Atay et al., 2009; ... Amplification of target gene PRRSV GP5 and construction of. pMD18-GP5 plasmid. Virus RNA was ... Construction of fusion expressed plasmid pET32-GP5-Hsp70. pMD18-GP5 and pET-32(α+) plasmids ...

  14. Interleukin-17A-Deficient Mice Are Highly Susceptible to Toxoplasma gondii Infection Due to Excessively Induced T. gondii HSP70 and Interferon Gamma Production.

    Science.gov (United States)

    Moroda, Masataka; Takamoto, Masaya; Iwakura, Yoichiro; Nakayama, Jun; Aosai, Fumie

    2017-12-01

    Interleukin17A (IL-17A) is known to be involved in the host defense against pathogens and the pathogenesis of autoimmune diseases. Previously, we showed that excessive amounts of interferon gamma (IFN-γ) play an important role in the pathogenesis of the lethal effects of Toxoplasma gondii by inducing anaphylactic responses. In the study described in this report, we examined the effects of IL-17A deficiency on murine host defense against oral T. gondii infection. IL-17A-deficient C57BL/6 (B6) mice exhibited higher rates of mortality than wild-type (WT) mice during the acute phase of T. gondii infection. CD4+ T cells in the mesenteric lymph nodes (mLNs) and ileum of T. gondii-infected IL-17A-deficient mice produced higher levels of IFN-γ than did those of WT mice. In addition, the level of T. gondii HSP70 (T.gHSP70) expression was also significantly increased in the ileum, mLNs, liver, and spleen of infected IL-17A-deficient mice compared with that in WT mice. These elevated levels of expression of T.gHSP70 and IFN-γ in infected IL-17A-deficient mice were presumably linked to the IL-17A defect since they decreased to WT levels after treatment with recombinant IL-17A. Furthermore, IL-17A-deficient mice were highly susceptible to the anaphylactic effect of T.gHSP70, and the survival of IL-17A-deficient mice during the acute phase was improved by treatment with an anti-T.gHSP70 monoclonal antibody. These results suggest that IL-17A plays an important role in host survival against T. gondii infection by protecting the host from an anaphylactic reaction via the downregulation of T.gHSP70 and IFN-γ production. Copyright © 2017 American Society for Microbiology.

  15. Broadening the functionality of a J-protein/Hsp70 molecular chaperone system.

    Directory of Open Access Journals (Sweden)

    Brenda A Schilke

    2017-10-01

    Full Text Available By binding to a multitude of polypeptide substrates, Hsp70-based molecular chaperone systems perform a range of cellular functions. All J-protein co-chaperones play the essential role, via action of their J-domains, of stimulating the ATPase activity of Hsp70, thereby stabilizing its interaction with substrate. In addition, J-proteins drive the functional diversity of Hsp70 chaperone systems through action of regions outside their J-domains. Targeting to specific locations within a cellular compartment and binding of specific substrates for delivery to Hsp70 have been identified as modes of J-protein specialization. To better understand J-protein specialization, we concentrated on Saccharomyces cerevisiae SIS1, which encodes an essential J-protein of the cytosol/nucleus. We selected suppressors that allowed cells lacking SIS1 to form colonies. Substitutions changing single residues in Ydj1, a J-protein, which, like Sis1, partners with Hsp70 Ssa1, were isolated. These gain-of-function substitutions were located at the end of the J-domain, suggesting that suppression was connected to interaction with its partner Hsp70, rather than substrate binding or subcellular localization. Reasoning that, if YDJ1 suppressors affect Ssa1 function, substitutions in Hsp70 itself might also be able to overcome the cellular requirement for Sis1, we carried out a selection for SSA1 suppressor mutations. Suppressing substitutions were isolated that altered sites in Ssa1 affecting the cycle of substrate interaction. Together, our results point to a third, additional means by which J-proteins can drive Hsp70's ability to function in a wide range of cellular processes-modulating the Hsp70-substrate interaction cycle.

  16. Broadening the functionality of a J-protein/Hsp70 molecular chaperone system.

    Science.gov (United States)

    Schilke, Brenda A; Ciesielski, Szymon J; Ziegelhoffer, Thomas; Kamiya, Erina; Tonelli, Marco; Lee, Woonghee; Cornilescu, Gabriel; Hines, Justin K; Markley, John L; Craig, Elizabeth A

    2017-10-01

    By binding to a multitude of polypeptide substrates, Hsp70-based molecular chaperone systems perform a range of cellular functions. All J-protein co-chaperones play the essential role, via action of their J-domains, of stimulating the ATPase activity of Hsp70, thereby stabilizing its interaction with substrate. In addition, J-proteins drive the functional diversity of Hsp70 chaperone systems through action of regions outside their J-domains. Targeting to specific locations within a cellular compartment and binding of specific substrates for delivery to Hsp70 have been identified as modes of J-protein specialization. To better understand J-protein specialization, we concentrated on Saccharomyces cerevisiae SIS1, which encodes an essential J-protein of the cytosol/nucleus. We selected suppressors that allowed cells lacking SIS1 to form colonies. Substitutions changing single residues in Ydj1, a J-protein, which, like Sis1, partners with Hsp70 Ssa1, were isolated. These gain-of-function substitutions were located at the end of the J-domain, suggesting that suppression was connected to interaction with its partner Hsp70, rather than substrate binding or subcellular localization. Reasoning that, if YDJ1 suppressors affect Ssa1 function, substitutions in Hsp70 itself might also be able to overcome the cellular requirement for Sis1, we carried out a selection for SSA1 suppressor mutations. Suppressing substitutions were isolated that altered sites in Ssa1 affecting the cycle of substrate interaction. Together, our results point to a third, additional means by which J-proteins can drive Hsp70's ability to function in a wide range of cellular processes-modulating the Hsp70-substrate interaction cycle.

  17. Expression of Selected Ginkgo biloba Heat Shock Protein Genes After Cold Treatment Could Be Induced by Other Abiotic Stress

    Science.gov (United States)

    Cao, Fuliang; Cheng, Hua; Cheng, Shuiyuan; Li, Linling; Xu, Feng; Yu, Wanwen; Yuan, Honghui

    2012-01-01

    Heat shock proteins (HSPs) play various stress-protective roles in plants. In this study, three HSP genes were isolated from a suppression subtractive hybridization (SSH) cDNA library of Ginkgo biloba leaves treated with cold stress. Based on the molecular weight, the three genes were designated GbHSP16.8, GbHSP17 and GbHSP70. The full length of the three genes were predicted to encode three polypeptide chains containing 149 amino acids (Aa), 152 Aa, and 657 Aa, and their corresponding molecular weights were predicted as follows: 16.67 kDa, 17.39 kDa, and 71.81 kDa respectively. The three genes exhibited distinctive expression patterns in different organs or development stages. GbHSP16.8 and GbHSP70 showed high expression levels in leaves and a low level in gynoecia, GbHSP17 showed a higher transcription in stamens and lower level in fruit. This result indicates that GbHSP16.8 and GbHSP70 may play important roles in Ginkgo leaf development and photosynthesis, and GbHSP17 may play a positive role in pollen maturation. All three GbHSPs were up-regulated under cold stress, whereas extreme heat stress only caused up-regulation of GbHSP70, UV-B treatment resulted in up-regulation of GbHSP16.8 and GbHSP17, wounding treatment resulted in up-regulation of GbHSP16.8 and GbHSP70, and abscisic acid (ABA) treatment caused up-regulation of GbHSP70 primarily. PMID:22754330

  18. Changing of the HSP70 Content in the Baikal Endemic Sponges Lubomirskiidae Under Conditions of Hyperthermia

    Directory of Open Access Journals (Sweden)

    Itskovich V.B.

    2015-12-01

    Full Text Available Baikal endemic sponges (Lubomirskiidae make up the bulk of the benthos biomass of the lake. For the first time the changes in the content of HSP70 in response to elevated environment temperature were analyzed in three endemic species of Baikal sponges: Baikalospongia bacillifera (Dybowski, 1880, B. intermedia (Dybowski, 1880 and Swartschewskia papyracea (Dybowski, 1880. Interspecific variability of constitutive HSP70 level was revealed for representatives of the three analyzed Lubomirskiidae species. After exposure at 13 °С for 3 and 7 days opposite changes were noted in the amount of HSP70. Under conditions of hyperthermia the protein level decrease at Baikalospongia species, while at the S. papyracea HSP70 content slightly increased. The differences in the mechanisms of stress adaptation probably affect the thermal resistance of the species, as well as are evidence supporting their specific status.

  19. The exported chaperone Hsp70-x supports virulence functions for Plasmodium falciparum blood stage parasites.

    Directory of Open Access Journals (Sweden)

    Sarah C Charnaud

    Full Text Available Malaria is caused by five different Plasmodium spp. in humans each of which modifies the host erythrocyte to survive and replicate. The two main causes of malaria, P. falciparum and P. vivax, differ in their ability to cause severe disease, mainly due to differences in the cytoadhesion of infected erythrocytes (IE in the microvasculature. Cytoadhesion of P. falciparum in the brain leads to a large number of deaths each year and is a consequence of exported parasite proteins, some of which modify the erythrocyte cytoskeleton while others such as PfEMP1 project onto the erythrocyte surface where they bind to endothelial cells. Here we investigate the effects of knocking out an exported Hsp70-type chaperone termed Hsp70-x that is present in P. falciparum but not P. vivax. Although the growth of Δhsp70-x parasites was unaffected, the export of PfEMP1 cytoadherence proteins was delayed and Δhsp70-x IE had reduced adhesion. The Δhsp70-x IE were also more rigid than wild-type controls indicating changes in the way the parasites modified their host erythrocyte. To investigate the cause of this, transcriptional and translational changes in exported and chaperone proteins were monitored and some changes were observed. We propose that PfHsp70-x is not essential for survival in vitro, but may be required for the efficient export and functioning of some P. falciparum exported proteins.

  20. Co-delivery of doxorubicin and recombinant plasmid pHSP70-Plk1-shRNA by bacterial magnetosomes for osteosarcoma therapy

    Science.gov (United States)

    Cheng, Li; Ke, Youqun; Yu, Shuisheng; Jing, Juehua

    2016-01-01

    To explore a novel combination of chemotherapy, gene therapy, and thermotherapy for osteosarcoma, a targeted heat-sensitive co-delivery system based on bacterial magnetosomes (BMs) was developed. The optimal culture conditions of magnetotactic bacteria (MTB) AMB-1 and characterization of BMs were achieved. A recombinant eukaryotic plasmid heat shock protein 70-polo-like kinase 1-short hairpin RNA (pHSP70-Plk1-shRNA) under transcriptional control of a thermosensitive promoter (human HSP70 promoter) was constructed for gene therapy. Doxorubicin (DOX) and pHSP70-Plk1-shRNA were included in the targeted thermosensitive co-delivery system, and in vitro DOX release activity, targeted gene silencing efficiency and in vitro antitumor efficacy were investigated. The results showed that the optimal culture conditions of MTB AMB-1 are an oxygen concentration of 4.0%, a pH value of 7.0, 20 μmol/L of ferrous sulfate, 800 mg/L of sodium nitrate, and 200 mg/L of succinic acid. The temperature of BMs reached 43°C within 3 minutes and could be maintained for 30 minutes by adjusting the magnitude of the alternating magnetic field (AMF). The diameters of BMs, BM-DOX, BM-recombinant eukaryotic plasmid pHSP70-Plk1-shRNA (shPlk1), and BM-DOX-shPlk1 were 43.7±4.6, 79.2±5.4, 88.9±7.8, and 133.5±11.4 nm, respectively. The zeta potentials of BMs, BM-DOX, BM-shPlk1, and BM-DOX-shPlk1 were −29.4±6.9, −9.5±5.6, −16.7±4.8, and −10.3±3.1 mV, respectively. Besides, the system exhibited good release behavior. DOX release rate from BM-DOX-shPlk1 was 54% after incubation with phosphate-buffered saline at 43°C and 37% after incubation with 50% fetal bovine serum, which was significantly higher than that at 37°C (P<0.05). In addition, the expressions of Plk1 mRNA and protein were significantly suppressed in cells treated with BM-DOX-shPlk1 following hyperthermia treatment under the influence of an AMF compared to other groups (P<0.05). Furthermore, evaluation of the effect of in

  1. Hsp 70 and 90 proteins as bio indicators of stress and protein damage in human lymphocytes exposed to neutrons; Proteinas Hsp 70 y 90 como bioindicadores de estres y dano proteico en linfocitos humanos expuestos a neutrones

    Energy Technology Data Exchange (ETDEWEB)

    Delgado, C. E.; Letechipia de L, C.; Vega C, H. R.; Sanchez R, S. H., E-mail: rmfranccesco00223@hotmail.com [Universidad Autonoma de Zacatecas, Unidad Academica de Estudios Nucleares, Cipres No. 10, Fracc. La Penuela, 98068 Zacatecas, Zac. (Mexico)

    2016-09-15

    Neutrons, when interacting with the cells of the body produce free radicals, so that exposure to high doses of ionizing radiation can cause different damage to the body that can cause cell death, therefore, these effects will depend on the amount of dose, time and individual factors such as gender, age, health status and nutrition. Therefore, knowledge of cellular responses to radiation exposure is critical for developing predictive markers useful for assessing people's exposure to radiation. The purpose of this study was to estimate the cellular protein damage through the Hsp 70 and 90 proteins exposed to neutrons in human lymphocytes from clinically healthy subjects. The cell tissue was obtained by venipuncture, the lymphocytes were separated by Ficoll-Paque concentration gradient, the experimental batches were formed, thus having 5 duplicate samples, subjected to neutron irradiation in a {sup 242}Am-Be at doses of 0.25, 0.50, 0.75, 1 and 1.25 μGy at three distances 20, 21.5 and 23 cm. As a positive control, a sample exposed to heat (40 degrees Celsius) was used for 40 min. The proteins of the experimental batch were analyzed by Western-Blot and protein quantification was analyzed by densitometry, on the other hand the oxidative stress was quantified by Oxi-Blot. Was found that the neutrons at doses of 0.25 and 0.5 μGy over expressed the Hsp-70 proteins, but for Hsp-90 no over-dose expressed, there was no protein damage at the exposure doses that were established. It can be estimated that Hsp-70 proteins can serve as bio indicators of cell stress by exposure doses of 0.25 and 0.5 μGy of neutrons. (Author)

  2. 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 homeostasis in goats.

  3. Monitoring the change of mitochondrial morphology and its metabolism of the breast cancer cells with the treatment of Hsp70 inhibitor during heat shock by fluorescence imaging

    Science.gov (United States)

    Yu, Biying; Yang, Hongqin; Zhang, Xiaoman; Li, Hui

    2016-10-01

    Heat shock (HS) is one of the best-studied exogenous cellular stresses, and all cellular compartments and metabolic processes are involved in HS response. The heat shock proteins (Hsps) expression enhanced during HS mainly localized in subcellular compartments, such as cytosol, endoplasmic reticulum and mitochandria. The major inducible heat shock protein 70 (Hsp70) modulate cellular homeostasis and promote cellular survival by blocking a caspase independent cell death through its association with apoptosis inducing factor. Mitochondria as the critical elements of HS response that participate in key metabolic reactions, and the changes in mitochonrial morphology may impact on mitochondrial metabolism. In this paper, the changes of mitorchondrial morphology in breast cancer cell have been monitored in real time after heat shock (43 °) by the fluorescence imaging, and the influence of Hsp70 inhibitor on mitochandrial structures have also been investigated. Then the information of mitochondrial metabolism which can be characterized by the level of the mitochondrial membrane potential has also been obtained wihout/with the treatment of Hsp70 inhibitor. Our data indicated that the mitochandrial morphology were related with the mitochandrial membrane potential, and the mitochandrial membrane potential was influenced significantly with the treatment of Hsp70 inhibitor during HS.

  4. Temperature effects on gene expression and morphological development of European eel, Anguilla anguilla larvae

    DEFF Research Database (Denmark)

    Politis, Sebastian Nikitas; Mazurais, David; Servili, Arianna

    2017-01-01

    to first-feeding, and the linked expression of targeted genes [heat shock proteins (hsp), growth hormone (gh) and insulin-like growth factors (igf)] associated to larval performance of European eel, Anguilla anguilla. Temperature effects on larval morphology and gene expression were investigated throughout......Temperature is important for optimization of rearing conditions in aquaculture, especially during the critical early life history stages of fish. Here, we experimentally investigated the impact of temperature (16, 18, 20, 22 and 24°C) on thermally induced phenotypic variability, from larval hatch...... affected by temperature. In real time, increasing temperature from 16 to 22°C accelerated larval development, while larval gene expression patterns (hsp70, hsp90, gh and igf-1) were delayed at cold temperatures (16°C) or accelerated at warm temperatures (20-22°C). All targeted genes (hsp70, hsp90, gh, igf...

  5. Hsp70/J-protein machinery from Glossina morsitans morsitans, vector of African trypanosomiasis.

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    Stephen J Bentley

    Full Text Available Tsetse flies (Glossina spp. are the sole vectors of the protozoan parasites of the genus Trypanosoma, the causative agents of African Trypanosomiasis. Species of Glossina differ in vector competence and Glossina morsitans morsitans is associated with transmission of Trypanosoma brucei rhodesiense, which causes an acute and often fatal form of African Trypanosomiasis. Heat shock proteins are evolutionarily conserved proteins that play critical roles in proteostasis. The activity of heat shock protein 70 (Hsp70 is regulated by interactions with its J-protein (Hsp40 co-chaperones. Inhibition of these interactions are emerging as potential therapeutic targets. The assembly and annotation of the G. m. morsitans genome provided a platform to identify and characterize the Hsp70s and J-proteins, and carry out an evolutionary comparison to its well-studied eukaryotic counterparts, Drosophila melanogaster and Homo sapiens, as well as Stomoxys calcitrans, a comparator species. In our study, we identified 9 putative Hsp70 proteins and 37 putative J-proteins in G. m. morsitans. Phylogenetic analyses revealed three evolutionarily distinct groups of Hsp70s, with a closer relationship to orthologues from its blood-feeding dipteran relative Stomoxys calcitrans. G. m. morsitans also lacked the high number of heat inducible Hsp70s found in D. melanogaster. The potential localisations, functions, domain organisations and Hsp70/J-protein partnerships were also identified. A greater understanding of the heat shock 70 (Hsp70 and J-protein (Hsp40 families in G. m. morsitans could enhance our understanding of the cell biology of the tsetse fly.

  6. Curcuma purpurascens BI. rhizome accelerates rat excisional wound healing: involvement of Hsp70/Bax proteins, antioxidant defense, and angiogenesis activity

    Science.gov (United States)

    Rouhollahi, Elham; Moghadamtousi, Soheil Zorofchian; Hajiaghaalipour, Fatemeh; Zahedifard, Maryam; Tayeby, Faezeh; Awang, Khalijah; Abdulla, Mahmood Ameen; Mohamed, Zahurin

    2015-01-01

    Purpose Curcuma purpurascens BI. is a member of Zingiberaceae family. The purpose of this study is to investigate the wound healing properties of hexane extract of C. purpurascens rhizome (HECP) against excisional wound healing in rats. Materials and methods Twenty four rats were randomly divided into 4 groups: A) negative control (blank placebo, acacia gum), B) low dose of HECP, C) high dose of HECP, and D) positive control, with 6 rats in each group. Full-thickness incisions (approximately 2.00 cm) were made on the neck area of each rat. Groups 1–4 were treated two-times a day for 20 days with blank placebo, HECP (100 mg/kg), HECP (200 mg/kg), and intrasite gel as a positive control, respectively. After 20 days, hematoxylin and eosin and Masson’s trichrome stainings were employed to investigate the histopathological alterations. Protein expressions of Bax and Hsp70 were examined in the wound tissues using immunohistochemistry analysis. In addition, levels of enzymatic antioxidants and malondialdehyde representing lipid peroxidation were measured in wound tissue homogenates. Results Macroscopic evaluation of wounds showed conspicuous elevation in wound contraction after topical administration of HECP at both doses. Moreover, histopathological analysis revealed noteworthy reduction in the scar width correlated with the enhanced collagen content and fibroblast cells, accompanied by a reduction of inflammatory cells in the granulation tissues. At the molecular level, HECP facilitates wound-healing process by downregulating Bax and upregulating Hsp70 protein at the wound site. The formation of new blood vessel was observed in Masson’s trichrome staining of wounds treated with HECP (100 and 200 mg/kg). In addition, HECP administration caused a significant surge in enzymatic antioxidant activities and a decline in lipid peroxidation. Conclusion These findings suggested that HECP accelerated wound-healing process in rats via antioxidant activity, angiogenesis

  7. Expression of Selected Ginkgo biloba Heat Shock Protein Genes After Cold Treatment Could Be Induced by Other Abiotic Stress

    Directory of Open Access Journals (Sweden)

    Feng Xu

    2012-05-01

    Full Text Available Heat shock proteins (HSPs play various stress-protective roles in plants. In this study, three HSP genes were isolated from a suppression subtractive hybridization (SSH cDNA library of Ginkgo biloba leaves treated with cold stress. Based on the molecular weight, the three genes were designated GbHSP16.8, GbHSP17 and GbHSP70. The full length of the three genes were predicted to encode three polypeptide chains containing 149 amino acids (Aa, 152 Aa, and 657 Aa, and their corresponding molecular weights were predicted as follows: 16.67 kDa, 17.39 kDa, and 71.81 kDa respectively. The three genes exhibited distinctive expression patterns in different organs or development stages. GbHSP16.8 and GbHSP70 showed high expression levels in leaves and a low level in gynoecia, GbHSP17 showed a higher transcription in stamens and lower level in fruit. This result indicates that GbHSP16.8 and GbHSP70 may play important roles in Ginkgo leaf development and photosynthesis, and GbHSP17 may play a positive role in pollen maturation. All three GbHSPs were up-regulated under cold stress, whereas extreme heat stress only caused up-regulation of GbHSP70, UV-B treatment resulted in up-regulation of GbHSP16.8 and GbHSP17, wounding treatment resulted in up-regulation of GbHSP16.8 and GbHSP70, and abscisic acid (ABA treatment caused up-regulation of GbHSP70 primarily.

  8. Diverse roles of C-terminal Hsp70-interacting protein (CHIP) in tumorigenesis.

    Science.gov (United States)

    Sun, Chao; Li, Hai-Long; Shi, Mei-Lin; Liu, Qing-Hua; Bai, Jin; Zheng, Jun-Nian

    2014-02-01

    The carboxyl terminus of Hsp70-interacting protein (CHIP) is a member of E3 ubiquitin ligase, functioning as a link between the chaperone (heat shock protein 70/90) and proteasome systems, playing a vital role in maintaining the protein homeostasis in the cytoplasm. CHIP has been demonstrated to be involved in tumorigenesis, proliferation and invasion in several malignancies, regulating a number of oncogenic proteins. However, CHIP has also been implicated in the modulation of tumor suppressor proteins. The pathogenic mechanism of CHIP expression in human malignancy is not yet clear, and a number of studies have suggested that CHIP may have opposing roles in different cancers. Therefore, many studies have focused on the relationship between CHIP and carcinoma. A literature search focusing on regulation network, biological function and clinical significance of CHIP in connection with its role in cancer development was performed on the MEDLINE databases. CHIP may be a potential diagnostic biomarker and therapeutic target for human cancer, and may play different roles in different human cancers. This inconsistence might be induced by the diversity of CHIP downstream targeting proteins. Therefore, the phenotypes determined by CHIP should be dependent on the function of its specific targets in a specific type of cancer cells. Whether CHIP contributes to tumor progression or suppression in various human cancers remains unclear, suggesting the necessity of further extensive investigation of its role in tumorigenesis.

  9. Clathrin Coat Disassembly Illuminates the Mechanisms of Hsp70 Force Generation

    Science.gov (United States)

    Liao, Hsien-Shun; Cuéllar, Jorge; Jin, Suping; Valpuesta, Jose M.; Jin, Albert J.; Lafer, Eileen M.

    2016-01-01

    Hsp70s use ATP hydrolysis to disrupt protein:protein associations or move macromolecules. One example is Hsc70-mediated disassembly of clathrin coats that form on vesicles during endocytosis. We exploit the exceptional features of these coats to test three models—Brownian ratchet, power-stroke and entropic pulling—proposed to explain how Hsp70s transform their substrates. Our data rule out the ratchet and power-stroke models, and instead support a collision pressure mechanism whereby collisions between clathrin coat walls and Hsc70s drive coats apart. Collision pressure is the complement to the pulling force described in the entropic pulling model. We also find that self-association can augment collision pressure to allow disassembly of clathrin lattices predicted to resist disassembly. These results illuminate how Hsp70s generate the forces that transform their substrates. PMID:27478930

  10. Structural insights into a unique Hsp70-Hsp40 interaction in the eukaryotic ribosome-associated complex.

    Science.gov (United States)

    Weyer, Felix Alexander; Gumiero, Andrea; Gesé, Genís Valentín; Lapouge, Karine; Sinning, Irmgard

    2017-02-01

    Cotranslational chaperones assist de novo folding of nascent polypeptides, prevent them from aggregating and modulate translation. The ribosome-associated complex (RAC) is unique in that the Hsp40 protein Zuo1 and the atypical Hsp70 chaperone Ssz1 form a stable heterodimer, which acts as a cochaperone for the Hsp70 chaperone Ssb. Here we present the structure of the Chaetomium thermophilum RAC core comprising Ssz1 and the Zuo1 N terminus. We show how the conserved allostery of Hsp70 proteins is abolished and this Hsp70-Hsp40 pair is molded into a functional unit. Zuo1 stabilizes Ssz1 in trans through interactions that in canonical Hsp70s occur in cis. Ssz1 is catalytically inert and cannot adopt the closed conformation, but the substrate binding domain β is completed by Zuo1. Our study offers insights into the coupling of a special Hsp70-Hsp40 pair, which evolved to link protein folding and translation.

  11. High Resolution Melting Analysis Targeting hsp70 as a Fast and Efficient Method for the Discrimination of Leishmania Species.

    Science.gov (United States)

    Zampieri, Ricardo Andrade; Laranjeira-Silva, Maria Fernanda; Muxel, Sandra Marcia; Stocco de Lima, Ana Carolina; Shaw, Jeffrey Jon; Floeter-Winter, Lucile Maria

    2016-02-01

    Protozoan parasites of the genus Leishmania cause a large spectrum of clinical manifestations known as Leishmaniases. These diseases are increasingly important public health problems in many countries both within and outside endemic regions. Thus, an accurate differential diagnosis is extremely relevant for understanding epidemiological profiles and for the administration of the best therapeutic protocol. Exploring the High Resolution Melting (HRM) dissociation profiles of two amplicons using real time polymerase chain reaction (real-time PCR) targeting heat-shock protein 70 coding gene (hsp70) revealed differences that allowed the discrimination of genomic DNA samples of eight Leishmania species found in the Americas, including Leishmania (Leishmania) infantum chagasi, L. (L.) amazonensis, L. (L.) mexicana, L. (Viannia) lainsoni, L. (V.) braziliensis, L. (V.) guyanensis, L. (V.) naiffi and L. (V.) shawi, and three species found in Eurasia and Africa, including L. (L.) tropica, L. (L.) donovani and L. (L.) major. In addition, we tested DNA samples obtained from standard promastigote culture, naturally infected phlebotomines, experimentally infected mice and clinical human samples to validate the proposed protocol. HRM analysis of hsp70 amplicons is a fast and robust strategy that allowed for the detection and discrimination of all Leishmania species responsible for the Leishmaniases in Brazil and Eurasia/Africa with high sensitivity and accuracy. This method could detect less than one parasite per reaction, even in the presence of host DNA.

  12. High Resolution Melting Analysis Targeting hsp70 as a Fast and Efficient Method for the Discrimination of Leishmania Species.

    Directory of Open Access Journals (Sweden)

    Ricardo Andrade Zampieri

    2016-02-01

    Full Text Available Protozoan parasites of the genus Leishmania cause a large spectrum of clinical manifestations known as Leishmaniases. These diseases are increasingly important public health problems in many countries both within and outside endemic regions. Thus, an accurate differential diagnosis is extremely relevant for understanding epidemiological profiles and for the administration of the best therapeutic protocol.Exploring the High Resolution Melting (HRM dissociation profiles of two amplicons using real time polymerase chain reaction (real-time PCR targeting heat-shock protein 70 coding gene (hsp70 revealed differences that allowed the discrimination of genomic DNA samples of eight Leishmania species found in the Americas, including Leishmania (Leishmania infantum chagasi, L. (L. amazonensis, L. (L. mexicana, L. (Viannia lainsoni, L. (V. braziliensis, L. (V. guyanensis, L. (V. naiffi and L. (V. shawi, and three species found in Eurasia and Africa, including L. (L. tropica, L. (L. donovani and L. (L. major. In addition, we tested DNA samples obtained from standard promastigote culture, naturally infected phlebotomines, experimentally infected mice and clinical human samples to validate the proposed protocol.HRM analysis of hsp70 amplicons is a fast and robust strategy that allowed for the detection and discrimination of all Leishmania species responsible for the Leishmaniases in Brazil and Eurasia/Africa with high sensitivity and accuracy. This method could detect less than one parasite per reaction, even in the presence of host DNA.

  13. Ekspresi Human Leukocyte Antigen-G (HLA-G dan Heat-Shock Protein-70 (Hsp-70 pada Pertumbuhan Janin Terhambat

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    Sri Sulistyowati

    2014-03-01

    Full Text Available Intra uterine growth retardation (IUGR is one of the leading causes of higher morbidity and mortality in perinatal. Immune maladaptation affects trophoblast invasion and spiralis arteria remodeling that will cause placental tissue hypoxia. This research aimed to analyze human leukocyte antigen-G (HLA-G and heat-shock protein-70 (Hsp-70 expression on the IUGR trophoblast and normal pregnancy, by applying analytical observational method and cross sectional approach. This research was conducted at the Obstetric and Gynecology Department of Dr. Moewardi Hospital Surakarta from November to December 2011. The total samples were 30, divided into two groups. There were 15 samples trophoblast on IUGR and 15 samples trophoblast from normal pregnancy. All samples were tested for HLA-G and Hsp-70 using immunohistochemistry. The data were analyzed by using t-test. The mean of HLA-G expression on the IUGR groups was 32.42±8.90 and on the normal pregnancy groups was 43.92±14.91 (p=0.016. Heat-shock protein70 expression on the IUGR groups was 2.4355+0.26647 and on the normal pregnancy groups was 1.5920+0.17142 with p=0.008. In conclusion, in IUGR, the HLA-G expression is lower and the Hsp-70 expression is higher than in normal pregnancy.

  14. Regulation of human heme oxygenase-1 gene expression under thermal stress.

    Science.gov (United States)

    Okinaga, S; Takahashi, K; Takeda, K; Yoshizawa, M; Fujita, H; Sasaki, H; Shibahara, S

    1996-06-15

    Heme oxygenase-1 is an essential enzyme in heme catabolism, and its human gene promoter contains a putative heat shock element (HHO-HSE). This study was designed to analyze the regulation of human heme oxygenase-1 gene expression under thermal stress. The amounts of heme oxygenase-1 protein were not increased by heat shock (incubation at 42 degrees C) in human alveolar macrophages and in a human erythroblastic cell line, YN-1-0-A, whereas heat shock protein 70 (HSP70) was noticeably induced. However, heat shock factor does bind in vitro to HHO-HSE and the synthetic HHO-HSE by itself is sufficient to confer the increase in the transient expression of a reporter gene upon heat shock. The deletion of the sequence, located downstream from HHO-HSE, resulted in the activation of a reporter gene by heat shock. These results suggest that HHO-HSE is potentially functional but is repressed in vivo. Interestingly, heat shock abolished the remarkable increase in the levels of heme oxygenase-1 mRNA in YN-1-0-A cells treated with hemin or cadmium, in which HSP70 mRNA was noticeably induced. Furthermore, transient expression assays showed that heat shock inhibits the cadmium-mediated activation of the heme oxygenase-1 promoter, whereas the HSP70 gene promoter was activated upon heat shock. Such regulation of heme oxygenase-1 under thermal stress may be of physiologic significance in erythroid cells.

  15. Leishmania infantum HSP70-II null mutant as candidate vaccine against leishmaniasis: a preliminary evaluation

    Directory of Open Access Journals (Sweden)

    Fresno Manuel

    2011-07-01

    Full Text Available Abstract Background Visceral leishmaniasis is the most severe form of leishmaniasis and no effective vaccine exists. The use of live attenuated vaccines is emerging as a promising vaccination strategy. Results In this study, we tested the ability of a Leishmania infantum deletion mutant, lacking both HSP70-II alleles (ΔHSP70-II, to provide protection against Leishmania infection in the L. major-BALB/c infection model. Administration of the mutant line by either intraperitoneal, intravenous or subcutaneous route invariably leads to the production of high levels of NO and the development in mice of type 1 immune responses, as determined by analysis of anti-Leishmania IgG subclasses. In addition, we have shown that ΔHSP70-II would be a safe live vaccine as immunodeficient SCID mice, and hamsters (Mesocricetus auratus, infected with mutant parasites did not develop any sign of pathology. Conclusions The results suggest that the ΔHSP70-II mutant is a promising and safe vaccine, but further studies in more appropriate animal models (hamsters and dogs are needed to appraise whether this attenuate mutant would be useful as vaccine against visceral leishmaniasis.

  16. Congenital sideroblastic anemia due to mutations in the mitochondrial HSP70 homologue HSPA9

    DEFF Research Database (Denmark)

    Schmitz-Abe, Klaus; Ciesielski, Szymon J.; Schmidt, Paul J.

    2015-01-01

    The congenital sideroblastic anemias (CSAs) are relatively uncommon diseases characterized by defects in mitochondrial heme synthesis, iron-sulfur (Fe-S) cluster biogenesis, or protein synthesis. Here we demonstrate that mutations in HSPA9, a mitochondrial HSP70 homolog located in the chromosome 5q...

  17. Leishmania infantum HSP70-II null mutant as candidate vaccine against leishmaniasis: a preliminary evaluation.

    Science.gov (United States)

    Carrión, Javier; Folgueira, Cristina; Soto, Manuel; Fresno, Manuel; Requena, Jose M

    2011-07-27

    Visceral leishmaniasis is the most severe form of leishmaniasis and no effective vaccine exists. The use of live attenuated vaccines is emerging as a promising vaccination strategy. In this study, we tested the ability of a Leishmania infantum deletion mutant, lacking both HSP70-II alleles (ΔHSP70-II), to provide protection against Leishmania infection in the L. major-BALB/c infection model. Administration of the mutant line by either intraperitoneal, intravenous or subcutaneous route invariably leads to the production of high levels of NO and the development in mice of type 1 immune responses, as determined by analysis of anti-Leishmania IgG subclasses. In addition, we have shown that ΔHSP70-II would be a safe live vaccine as immunodeficient SCID mice, and hamsters (Mesocricetus auratus), infected with mutant parasites did not develop any sign of pathology. The results suggest that the ΔHSP70-II mutant is a promising and safe vaccine, but further studies in more appropriate animal models (hamsters and dogs) are needed to appraise whether this attenuate mutant would be useful as vaccine against visceral leishmaniasis.

  18. An Hsp70 peptide initiates NK cell killing of leukemic blasts after stem cell transplantation

    NARCIS (Netherlands)

    Gross, Catharina; Holler, Ernst; Stangl, Stefan; Dickinson, Anne; Pockley, A. Graham; Asea, Alexzander A.; Mallappa, Nagaraj; Multhoff, Gabriele

    In contrast to solid tumors, leukemic blasts frequently present both Hsp70 and HLA-E on their cell Surface and thereby present activating and inhibitory signals to CD94(+) NK cells. In the first 12 months after stem cell trail splantation (SCT) CD94(+) NK cells clearly dominate over

  19. Design, synthesis and biological evaluation of novel HSP70 inhibitors: N, N'-disubstituted thiourea derivatives.

    Science.gov (United States)

    Zeng, Yan-Qun; Cao, Rui-Yuan; Yang, Jian-Ling; Li, Xing-Zhou; Li, Song; Zhong, Wu

    2016-08-25

    As novel heat shock protein 70 (HSP70) inhibitors, N, N'-disubstituted thiourea derivatives were designed and synthesized based on the X-ray structure of the ATPase domain (nucleotide binding domain, NBD). An ATPase activity inhibition assay revealed that these compounds effectively inhibited HSP70 ATPase activity. The results revealed that the compounds 370/371/374/379/380//392/394/397/404/405 and 407 can inhibit the HSP70 ATPase turnover with high percentages of inhibition: 50.42, 38.46, 50.45, 44.12, 47.13, 50.50, 40.95, 65.36, 46.23, 35.78, and 58.37 in 200 μM, respectively. Significant synergies with lapatinib were observed for compound 379 and compound 405 in the BT474 breast cancer cell line. A structure-function analysis revealed that most of the thiourea derivatives exhibited cooperative action with lapatinib in the BT474 cancer cell line and the BT/Lap(R)1.0 lapatinib-resistant cell line. HSP70 inhibitors may be developed as synergetic drugs in drug-resistant cancer therapy. Copyright © 2016 The Authors. Published by Elsevier Masson SAS.. All rights reserved.

  20. Identification and characterization of cytosolic Hansenula polymorpha proteins belonging to the Hsp70 protein family

    NARCIS (Netherlands)

    Titorenko, Vladimir I.; Evers, Melchior E.; Diesel, Andre; Samyn, Bart; Beeumen, Josef van; Roggenkamp, Rainer; Kiel, Jan A.K.W.; Klei, Ida J. van der; Veenhuis, Marten

    We have isolated two members of the Hsp70 protein family from the yeast Hansenula polymorpha using affinity chromatography. Both proteins were located in the cytoplasm. One of these, designated Hsp72, was inducible in nature (e.g. by heat shock). The second protein (designated Hsc74) was

  1. Genome-Wide Characterization of Heat-Shock Protein 70s from Chenopodium quinoa and Expression Analyses of Cqhsp70s in Response to Drought Stress.

    Science.gov (United States)

    Liu, Jianxia; Wang, Runmei; Liu, Wenying; Zhang, Hongli; Guo, Yaodong; Wen, Riyu

    2018-01-23

    Heat-shock proteins (HSPs) are ubiquitous proteins with important roles in response to biotic and abiotic stress. The 70-kDa heat-shock genes (Hsp70s) encode a group of conserved chaperone proteins that play central roles in cellular networks of molecular chaperones and folding catalysts across all the studied organisms including bacteria, plants and animals. Several Hsp70s involved in drought tolerance have been well characterized in various plants, whereas no research on Chenopodium quinoa HSPs has been completed. Here, we analyzed the genome of C. quinoa and identified sixteen Hsp70 members in quinoa genome. Phylogenetic analysis revealed the independent origination of those Hsp70 members, with eight paralogous pairs comprising the Hsp70 family in quinoa. While the gene structure and motif analysis showed high conservation of those paralogous pairs, the synteny analysis of those paralogous pairs provided evidence for expansion coming from the polyploidy event. With several subcellular localization signals detected in CqHSP70 protein paralogous pairs, some of the paralogous proteins lost the localization information, indicating the diversity of both subcellular localizations and potential functionalities of those HSP70s. Further gene expression analyses revealed by quantitative polymerase chain reaction (qPCR) analysis illustrated the significant variations of Cqhsp70s in response to drought stress. In conclusion, the sixteen Cqhsp70s undergo lineage-specific expansions and might play important and varied roles in response to drought stress.

  2. Association of plasma IL-6 and Hsp70 with HRV at different levels of PAHs metabolites.

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    Jian Ye

    Full Text Available Exposure to polycyclic aromatic hydrocarbons (PAHs is associated with reduced heart rate variability (HRV, a strong predictor of cardiovascular diseases, but the mechanism is not well understood.We hypothesized that PAHs might induce systemic inflammation and stress response, contributing to altered cardiac autonomic function.HRV indices were measured using a 3-channel digital Holter monitor in 800 coke oven workers. Plasma levels of interleukin-6 (IL-6 and heat shock protein 70 (Hsp70 were determined using ELISA. Twelve urinary PAHs metabolites (OH-PAHs were measured by gas chromatography-mass spectrometry.We found that significant dose-dependent relationships between four urinary OH-PAHs and IL-6 (all Ptrend<0.05; and an increase in quartiles of IL-6 was significantly associated with a decrease in total power (TP and low frequency (LF (Ptrend = 0.014 and 0.006, respectively. In particular, elevated IL-6 was associated in a dose-dependent manner with decreased TP and LF in the high-PAHs metabolites groups (all Ptrend<0.05, but not in the low-PAHs metabolites groups. No significant association between Hsp70 and HRV in total population was found after multivariate adjustment. However, increased Hsp70 was significantly associated with elevated standard deviation of NN intervals (SDNN, TP and LF in the low-PAHs metabolites groups (all Ptrend<0.05. We also observed that both IL-6 and Hsp70 significantly interacted with multiple PAHs metabolites in relation to HRV.In coke oven workers, increased IL-6 was associated with a dose-response decreased HRV in the high-PAHs metabolites groups, whereas increase of Hsp70 can result in significant dose-related increase in HRV in the low-PAHs metabolites groups.

  3. Exercise attenuates neurological deficits by stimulating a critical HSP70/NF-κB/IL-6/synapsin I axis in traumatic brain injury rats.

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    Chio, Chung-Ching; Lin, Hung-Jung; Tian, Yu-Feng; Chen, Yu-Chieh; Lin, Mao-Tsun; Lin, Cheng-Hsien; Chang, Ching-Ping; Hsu, Chien-Chin

    2017-04-24

    Despite previous evidence for a potent inflammatory response after a traumatic brain injury (TBI), it is unknown whether exercise preconditioning (EP) improves outcomes after a TBI by modulating inflammatory responses. We performed quantitative real-time PCR (qPCR) to quantify the genes encoding 84 cytokines and chemokines in the peripheral blood and used ELISA to determine both the cerebral and blood levels of interleukin-6 (IL-6). We also performed the chromatin immunoprecipitation (ChIP) assay to evaluate the extent of nuclear factor kappa-B (NF-κB) binding to the DNA elements in the IL-6 promoter regions. Also, we adopted the Western blotting assay to measure the cerebral levels of heat shock protein (HSP) 70, synapsin I, and β-actin. Finally, we performed both histoimmunological and behavioral assessment to measure brain injury and neurological deficits, respectively. We first demonstrated that TBI upregulated nine pro-inflammatory and/or neurodegenerative messenger RNAs (mRNAs) in the peripheral blood such as CXCL10, IL-18, IL-16, Cd-70, Mif, Ppbp, Ltd, Tnfrsf 11b, and Faslg. In addition to causing neurological injuries, TBI also upregulated the following 14 anti-inflammatory and/or neuroregenerative mRNAs in the peripheral blood such as Ccl19, Ccl3, Cxcl19, IL-10, IL-22, IL-6, Bmp6, Ccl22, IL-7, Bmp7, Ccl2, Ccl17, IL-1rn, and Gpi. Second, we observed that EP inhibited both neurological injuries and six pro-inflammatory and/or neurodegenerative genes (Cxcl10, IL-18, IL-16, Cd70, Mif, and Faslg) but potentiated four anti-inflammatory and/or neuroregenerative genes (Bmp6, IL-10, IL-22, and IL-6). Prior depletion of cerebral HSP70 with gene silence significantly reversed the beneficial effects of EP in reducing neurological injuries and altered gene profiles after a TBI. A positive Pearson correlation exists between IL-6 and HSP70 in the peripheral blood or in the cerebral levels. In addition, gene silence of cerebral HSP70 significantly reduced the

  4. HSP70-2 (HSPA1B) is associated with noncognitive symptoms in late-onset Alzheimer's disease.

    Science.gov (United States)

    Clarimón, Jordi; Bertranpetit, Jaume; Boada, Mercè; Tàrraga, Lluís; Comas, David

    2003-09-01

    Neuropsychiatric manifestations are common in Alzheimer's disease (AD) and their phenotypic expression might be related to physiopathological and genetic causes. Multiple studies have implicated oxidative stress to the pathogenesis and possible etiology of AD. One of the mechanisms to protect cells from oxidative stress is the expression of heat-shock proteins (HSP). HSPA1B (alternatively known as HSP70-2) has been related to AD pathophysiology. In the present analysis, 77 AD patients were classified according to their cognitive status with the Neuropsychiatric Inventory and were genotyped for an insertion/deletion (A1/A2) polymorphism. The A2 allele conferred a significant increase of psychiatric morbidity in an allele-dose manner (P noncognitive symptoms in AD.

  5. Hsp70 Forms Antiparallel Dimers Stabilized by Post-translational Modifications to Position Clients for Transfer to Hsp90

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    Nina Morgner

    2015-05-01

    Full Text Available Protein folding in cells is regulated by networks of chaperones, including the heat shock protein 70 (Hsp70 system, which consists of the Hsp40 cochaperone and a nucleotide exchange factor. Hsp40 mediates complex formation between Hsp70 and client proteins prior to interaction with Hsp90. We used mass spectrometry (MS to monitor assemblies formed between eukaryotic Hsp90/Hsp70/Hsp40, Hop, p23, and a client protein, a fragment of the glucocorticoid receptor (GR. We found that Hsp40 promotes interactions between the client and Hsp70, and facilitates dimerization of monomeric Hsp70. This dimerization is antiparallel, stabilized by post-translational modifications (PTMs, and maintained in the stable heterohexameric client-loading complex Hsp902Hsp702HopGR identified here. Addition of p23 to this client-loading complex induces transfer of GR onto Hsp90 and leads to expulsion of Hop and Hsp70. Based on these results, we propose that Hsp70 antiparallel dimerization, stabilized by PTMs, positions the client for transfer from Hsp70 to Hsp90.

  6. In contrast to conventional inactivated influenza vaccines, 4xM2e.HSP70c fusion protein fully protected mice against lethal dose of H1, H3 and H9 influenza A isolates circulating in Iran

    Energy Technology Data Exchange (ETDEWEB)

    Ebrahimi, Seyyed Mahmoud, E-mail: smebrahimi@shirazu.ac.ir [Applied Biotechnology Research Center, Baqiyatallah University of Medical Sciences, P.O. Box 14155-3651,Tehran (Iran, Islamic Republic of); Research Center of Virus and Vaccine, Baqiyatallah University of Medical Science, P.O.Box 14155-3651, Tehran (Iran, Islamic Republic of); Dabaghian, Mehran [Department of Pathobiology, University of Tehran, Faculty of Veterinary Medicine, P.O. Box 14155-6453, Tehran (Iran, Islamic Republic of); Tebianian, Majid [Department of Biotechnology, Razi Vaccine and Serum Research Institute (RVSRI), P.O. Box 31975/148, Karaj, Tehran (Iran, Islamic Republic of); Zabeh Jazi, Mohammad Hossein [Department of Pathobiology, University of Tehran, Faculty of Veterinary Medicine, P.O. Box 14155-6453, Tehran (Iran, Islamic Republic of)

    2012-08-15

    Ideal vaccines against influenza viruses should elicit not only a humoral response, but also a cellular response. Mycobacterium tuberculosis HSP70 (mHSP70) have been found to promote immunogenic APCs function, elicit a strong cytotoxic T lymphocyte (CTL) response, and prevent the induction of tolerance. Moreover, it showed linkage of antigens to the C-terminus of mHSP70 (mHSP70c) can represent them as vaccines resulted in more potent, protective antigen specific responses in the absence of adjuvants or complex formulations. Hence, recombinant fusion protein comprising C-terminus of mHSP70 genetically fused to four tandem repeats of the ectodomain of the conserved influenza matrix protein M2 (M2e) was expressed in Escherichia coli, purified under denaturing condition, refolding, and then confirmed by SDS-PAGE, respectively. The recombinant fusion protein, 4xM2e.HSP70c, retained its immunogenicity and displayed the protective epitope of M2e by ELISA and FITC assays. A prime-boost administration of 4xM2e.HSP70c formulated in F105 buffer by intramuscular route in mice (Balb/C) provided full protection against lethal dose of mouse-adapted H1N1, H3N2, or H9N2 influenza A isolates from Iran compared to 0-33.34% survival rate of challenged unimmunized and immunized mice with the currently in use conventional vaccines designated as control groups. However, protection induced by immunization with 4xM2e.HSP70c failed to prevent weight loss in challenged mice; they experienced significantly lower weight loss, clinical symptoms and higher lung viral clearance in comparison with protective effects of conventional influenza vaccines in challenged mice. These data demonstrate that C-terminal domain of mHSP70 can be a superior candidate to deliver the adjuvant function in M2e-based influenza A vaccine in order to provide significant protection against multiple influenza A virus strains.

  7. [The role of extracellular chaperone Hsp70 in creating antitumor immunity in rat rhabdomyosarcoma RA-2 model].

    Science.gov (United States)

    Guzhova, I V; Komarova, E Iu; Pimenova, A A; Bakhtin, Iu B; Kaminskaia, E V; Margulis, B A

    2008-01-01

    Immunization of experimental animals with extract or membranes of rat rhabdomyosarcoma RA-2 in combination with pure Hsp70 did not offer any significant effect of protection from subsequent RA-2 cells-stimulated tumor growth. By contrast, immunization with preparations of pure Hsp70 led to a significant decrease in number and size of tumors as well as elevation of concentrations of antibodies against RA-2 cells. Also, enhanced blood levels of Hsp70 involved delayed tumor growth. In vitro tests Hsp70 incubation with RA-2 cells was followed by a 30-35% rise in cytotoxic lymphocytes levels. An ability of pure Hsp70 preparations to stimulate humoral and antitumor response was demonstrated. Hence, they may be used in developing vaccine formulas.

  8. The endosymbiotic bacterium Holospora obtusa enhances heat-shock gene expression of the host Paramecium caudatum.

    Science.gov (United States)

    Hori, Manabu; Fujishima, Masahiro

    2003-01-01

    The bacterium Holospora obtusa is a macronuclear-specific symbiont of the ciliate Paramecium caudatum. H. obtusa-bearing paramecia could survive even after the cells were quickly heated from 25 degrees C to 35 degrees C. To determine whether infection with H. obtusa confers heat shock resistance on its host, we isolated genes homologous to the heat shock protein genes hsp60 and hsp70 from P. caudatum. The deduced amino acid sequences of both cDNAs were highly homologous to hsp family sequences from other eukaryotes. Competitive PCR showed that H. obtusa-free paramecia expressed only trace amounts of hsp60 and hsp70 mRNA at 25 degrees C, but that expression of hsp70 was enhanced immediately after the cells were transferred to 35 degrees C. H. obtusa-bearing paramecia expressed high levels of hsp7O mRNA even at 25 degrees C and the level was further enhanced when the cells were incubated at 35 degrees C. In contrast, the expression pattern of hsp60 mRNA was the same in H. obtusa-bearing as in H. obtusa-free paramecia. These results indicate that infection with its endosymbiont can confer a heat-shock resistant nature on its host cells.

  9. The Malarial Exported PFA0660w Is an Hsp40 Co-Chaperone of PfHsp70-x.

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    Michael O Daniyan

    Full Text Available Plasmodium falciparum, the human pathogen responsible for the most dangerous malaria infection, survives and develops in mature erythrocytes through the export of proteins needed for remodelling of the host cell. Molecular chaperones of the heat shock protein (Hsp family are prominent members of the exportome, including a number of Hsp40s and a Hsp70. PFA0660w, a type II Hsp40, has been shown to be exported and possibly form a complex with PfHsp70-x in the infected erythrocyte cytosol. However, the chaperone properties of PFA0660w and its interaction with human and parasite Hsp70s are yet to be investigated. Recombinant PFA0660w was found to exist as a monomer in solution, and was able to significantly stimulate the ATPase activity of PfHsp70-x but not that of a second plasmodial Hsp70 (PfHsp70-1 or a human Hsp70 (HSPA1A, indicating a potential specific functional partnership with PfHsp70-x. Protein binding studies in the presence and absence of ATP suggested that the interaction of PFA0660w with PfHsp70-x most likely represented a co-chaperone/chaperone interaction. Also, PFA0660w alone produced a concentration-dependent suppression of rhodanese aggregation, demonstrating its chaperone properties. Overall, we have provided the first biochemical evidence for the possible role of PFA0660w as a chaperone and as co-chaperone of PfHsp70-x. We propose that these chaperones boost the chaperone power of the infected erythrocyte, enabling successful protein trafficking and folding, and thereby making a fundamental contribution to the pathology of malaria.

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

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    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.

  11. The effect of glutamine infusion on the inflammatory response and HSP70 during human experimental endotoxaemia

    DEFF Research Database (Denmark)

    Andreasen, Anne Sofie; Pedersen-Skovsgaard, Theis; Mortensen, Ole Hartvig

    2009-01-01

    INTRODUCTION: Glutamine supplementation has beneficial effects on morbidity and mortality in critically ill patients, possibly in part through an attenuation of the proinflammatory cytokine response and a stimulation of heat shock protein (HSP)70. We infused either alanine-glutamine or saline...... an infusion of alanine-glutamine at a rate of 0.025 g/(kg body weight x hour) or saline for 10 hours. After 2 hours, an intravenous bolus of Escherichia coli endotoxin (0.3 ng/kg) was administered. Blood samples were collected hourly for the following 8 hours. HSP70 protein content in isolated blood...... mononuclear cells (BMNCs) was measured by Western blotting. RESULTS: Plasma glutamine increased during alanine-glutamine infusion. Endotoxin reduced plasma glutamine during both trials, but plasma glutamine levels remained above baseline with alanine-glutamine supplementation. Endotoxin injection...

  12. Mechanics of Hsp70 chaperones enables differential interaction with client proteins.

    Science.gov (United States)

    Schlecht, Rainer; Erbse, Annette H; Bukau, Bernd; Mayer, Matthias P

    2011-03-01

    Hsp70 chaperones interact with a wide spectrum of substrates ranging from unfolded to natively folded and aggregated proteins. Structural evidence suggests that bound substrates are entirely enclosed in a β-sheet cavity covered by a helical lid, which requires structural rearrangements including lid opening to allow substrate access. We analyzed the mechanics of the lid movement of bacterial DnaK by disulfide fixation of lid elements to the β-sheet and by electron paramagnetic resonance spectroscopy using spin labels in the lid and β-sheet. Our results indicate that the lid-forming helix B adopts at least three conformational states and, notably, does not close over bound proteins, implying that DnaK does not only bind to extended peptide stretches of protein substrates but can also accommodate regions with substantial tertiary structure. This flexible binding mechanism provides a basis for the broad spectrum of substrate conformers of Hsp70s.

  13. Genetic evaluation of the HSP70 protein in the Japanese quail (Coturnix japonica).

    Science.gov (United States)

    Gaviol, H C T; Gasparino, E; Prioli, A J; Soares, M A M

    2008-02-12

    Heat stress is one of the main problems in modern aviculture, since it affects birds especially in the final phase of rearing, causing bird mortality and economic losses to the aviculturist. The quail, as most birds, has difficulties in dissipating heat. However, little is known about the mechanism that controls the responses of the organism to stressor agents. Therefore, the study of heat shock proteins (HSPs) in these birds is important. A 960-bp portion of HSP70 was amplified using oligonucleotide primers specific for chickens. The fragment was sequenced, since it was the same protein, although some modifications have been observed. It showed 98% homology with HSP70 stress protein in Gallus gallus and 99% homology with Numida meleageris.

  14. Differences in salinity tolerance and gene expression between two populations of Atlantic cod (Gadus morhua) in response to salinity stress

    DEFF Research Database (Denmark)

    Larsen, Peter Foged; Eg Nielsen, Einar; Meier, Kristian

    2012-01-01

    in salinity tolerance and gene expression among Atlantic cod (Gadus morhua) from two populations distributed across a steep salinity gradient, we observed high mortality (45% North Sea cod and 80% Baltic Sea cod) in a reciprocal common garden setup. Quantitative RT-PCR assays for expression of hsp70 and Na....... The findings strongly suggest that Atlantic cod are adapted to local saline conditions, despite relatively low levels of neutral genetic divergence between populations...

  15. Identification of a Hsp70 recognition domain within the rubisco small subunit transit peptide.

    Science.gov (United States)

    Ivey, R A; Subramanian, C; Bruce, B D

    2000-04-01

    The interaction between SStp, the transit peptide of the precursor protein to the small subunit of Rubisco (prSSU) and two Hsp70 molecular chaperones, Escherichia coli DnaK and pea (Pisum sativum) CSS1, was investigated in detail. Two statistical analyses were developed and used to investigate and predict regions of SStp recognized by DnaK. Both algorithms suggested that DnaK would have high affinity for the N terminus of SStp, moderate affinity for the central region, and low affinity for the C terminus. Furthermore, both algorithms predicted this affinity pattern for >75% of the transit peptides analyzed in the chloroplast transit peptide (CHLPEP) database. In vitro association between SStp and these Hsp70s was confirmed by three independent assays: limited trypsin resistance, ATPase stimulation, and native gel shift. Finally, synthetic peptides scanning the length of SStp and C-terminal deletion mutants of SStp were used to experimentally map the region of greatest DnaK affinity to the N terminus. CSS1 displayed a similar affinity for the N terminus of SStp. The major stromal Hsp70s affinity for the N terminus of SStp and other transit peptides supports a molecular motor model in which the chaperone functions as an ATP-dependent translocase, committing chloroplast precursor proteins to unidirectional movement across the envelope.

  16. Graphene oxide based electrochemical label free immunosensor for rapid and highly sensitive determination of tumor marker HSP70.

    Science.gov (United States)

    Özcan, Burcu; Sezgintürk, Mustafa Kemal

    2016-11-01

    In this study, it is aimed to design a label free immunosensor for determination of HSP70 (heat shock protein 70). Glassy carbon electrode was used as a working electrode. Graphene oxide was covered on the working electrode surface. AntiHSP70 as a biorecognition element of the biosensor was covalently immobilized onto the graphene oxide layer by using EDC/NHS chemistry. The immobilization of antiHSP70 and binding of HSP70 protein onto the electrode surface were monitored by cyclic voltammetry and electrochemical impedance spectroscopy. Single frequency technique was also utilized to monitor binding characterization of HSP70 and antiHSP70. Surface morphology was defined by using scanning electron microscopy. All important fabrication steps of the biosensor were optimized to prepare an ultrasensitive biosensor. Under optimum conditions, analytical studies such as linearity, repeatability, and reproducibility were also experienced. A linear detection range of HSP70 was determined between 12 and 144fg/mL. Moreover, Kramer's Kronig transform was applied on impedance data. Finally, the biosensor was applied with real human blood serum samples and hopeful results were obtained. Copyright © 2016 Elsevier B.V. All rights reserved.

  17. Cysteine string protein promotes proteasomal degradation of the cystic fibrosis transmembrane conductance regulator (CFTR) by increasing its interaction with the C terminus of Hsp70-interacting protein and promoting CFTR ubiquitylation.

    Science.gov (United States)

    Schmidt, Béla Z; Watts, Rebecca J; Aridor, Meir; Frizzell, Raymond A

    2009-02-13

    Cysteine string protein (Csp) is a J-domain-containing protein whose overexpression blocks the exit of cystic fibrosis transmembrane conductance regulator (CFTR) from the endoplasmic reticulum (ER). Another method of blocking ER exit, the overexpression of Sar1-GTP, however, yielded twice as much immature CFTR compared with Csp overexpression. This finding suggested that Csp not only inhibits CFTR ER exit but also facilitates the degradation of immature CFTR. This was confirmed by treatment with a proteasome inhibitor, which returned the level of immature CFTR to that found in cells expressing Sar1-GTP only. CspH43Q, which does not interact with Hsc70/Hsp70 efficiently, did not promote CFTR degradation, suggesting that the pro-degradative effect of Csp requires Hsc70/Hsp70 binding/activation. In agreement with this, Csp overexpression increased the amount of Hsc70/Hsp70 co-immunoprecipitated with CFTR, whereas overexpression of CspH43Q did not. The Hsc70/Hsp70 binding partner C terminus of Hsp70-interacting protein (CHIP) can target CFTR for proteasome-mediated degradation. Csp overexpression also increased the amount of CHIP co-immunoprecipitated with CFTR. In addition, CHIP interacted directly with Csp, which was confirmed by in vitro binding experiments. Csp overexpression also increased CFTR ubiquitylation and reduced the half-life of immature CFTR. These findings indicate that Csp not only regulates the exit of CFTR from the ER, but that this action is accompanied by Hsc70/Hsp70 and CHIP-mediated CFTR degradation.

  18. In vivo generation of neurotoxic prion protein: role for hsp70 in accumulation of misfolded isoforms.

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    Pedro Fernandez-Funez

    2009-06-01

    Full Text Available Prion diseases are incurable neurodegenerative disorders in which the normal cellular prion protein (PrP(C converts into a misfolded isoform (PrP(Sc with unique biochemical and structural properties that correlate with disease. In humans, prion disorders, such as Creutzfeldt-Jakob disease, present typically with a sporadic origin, where unknown mechanisms lead to the spontaneous misfolding and deposition of wild type PrP. To shed light on how wild-type PrP undergoes conformational changes and which are the cellular components involved in this process, we analyzed the dynamics of wild-type PrP from hamster in transgenic flies. In young flies, PrP demonstrates properties of the benign PrP(C; in older flies, PrP misfolds, acquires biochemical and structural properties of PrP(Sc, and induces spongiform degeneration of brain neurons. Aged flies accumulate insoluble PrP that resists high concentrations of denaturing agents and contains PrP(Sc-specific conformational epitopes. In contrast to PrP(Sc from mammals, PrP is proteinase-sensitive in flies. Thus, wild-type PrP rapidly converts in vivo into a neurotoxic, protease-sensitive isoform distinct from prototypical PrP(Sc. Next, we investigated the role of molecular chaperones in PrP misfolding in vivo. Remarkably, Hsp70 prevents the accumulation of PrP(Sc-like conformers and protects against PrP-dependent neurodegeneration. This protective activity involves the direct interaction between Hsp70 and PrP, which may occur in active membrane microdomains such as lipid rafts, where we detected Hsp70. These results highlight the ability of wild-type PrP to spontaneously convert in vivo into a protease-sensitive isoform that is neurotoxic, supporting the idea that protease-resistant PrP(Sc is not required for pathology. Moreover, we identify a new role for Hsp70 in the accumulation of misfolded PrP. Overall, we provide new insight into the mechanisms of spontaneous accumulation of neurotoxic PrP and uncover

  19. Hsp70 May Be a Molecular Regulator of Schistosome Host Invasion.

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    Kenji Ishida

    2016-09-01

    Full Text Available Schistosomiasis is a debilitating disease that affects over 240 million people worldwide and is considered the most important neglected tropical disease following malaria. Free-swimming freshwater cercariae, one of the six morphologically distinct schistosome life stages, infect humans by directly penetrating through the skin. Cercariae identify and seek the host by sensing chemicals released from human skin. When they reach the host, they burrow into the skin with the help of proteases and other contents released from their acetabular glands and transform into schistosomula, the subsequent larval worm stage upon skin infection. Relative to host invasion, studies have primarily focused on the nature of the acetabular gland secretions, immune response of the host upon exposure to cercariae, and cercaria-schistosomulum transformation methods. However, the molecular signaling pathways involved from host-seeking through the decision to penetrate skin are not well understood. We recently observed that heat shock factor 1 (Hsf1 is localized to the acetabular glands of infectious schistosome cercariae, prompting us to investigate a potential role for heat shock proteins (HSPs in cercarial invasion. In this study, we report that cercarial invasion behavior, similar to the behavior of cercariae exposed to human skin lipid, is regulated through an Hsp70-dependent process, which we show by using chemical agents that target Hsp70. The observation that biologically active protein activity modulators can elicit a direct and clear behavioral change in parasitic schistosome larvae is itself interesting and has not been previously observed. This finding suggests a novel role for Hsp70 to act as a switch in the cercaria-schistosomulum transformation, and it allows us to begin elucidating the pathways associated with cercarial host invasion. In addition, because the Hsp70 protein and its structure/function is highly conserved, the model that Hsp70 acts as a

  20. Conformational changes in human Hsp70 induced by high hydrostatic pressure produce oligomers with ATPase activity but without chaperone activity.

    Science.gov (United States)

    Araujo, Thaís L S; Borges, Julio Cesar; Ramos, Carlos H; Meyer-Fernandes, José Roberto; Oliveira Júnior, Reinaldo S; Pascutti, Pedro G; Foguel, Debora; Palhano, Fernando L

    2014-05-13

    We investigated the folding of the 70 kDa human cytosolic inducible protein (Hsp70) in vitro using high hydrostatic pressure as a denaturing agent. We followed the structural changes in Hsp70 induced by high hydrostatic pressure using tryptophan fluorescence, molecular dynamics, circular dichroism, high-performance liquid chromatography gel filtration, dynamic light scattering, ATPase activity, and chaperone activity. Although monomeric, Hsp70 is very sensitive to hydrostatic pressure; after pressure had been removed, the protein did not return to its native sate but instead formed oligomeric species that lost chaperone activity but retained ATPase activity.

  1. Interpopulation differences in expression of candidate genes for salinity tolerance in winter migrating anadromous brown trout ( Salmo trutta L.)

    DEFF Research Database (Denmark)

    Larsen, Peter Foged; Eg Nielsen, Einar; Koed, Anders

    2008-01-01

    saline conditions and quantified expression of the hsp70 and Na/K-ATPases alpha 1b genes following acclimation to freshwater and full-strength seawater at 2 degrees C and 10 degrees C. An interaction effect of low temperature and high salinity on expression of both hsp70 and Na/K-ATPase alpha 1b......Background: Winter migration of immature brown trout (Salmo trutta) into freshwater rivers has been hypothesized to result from physiologically stressful combinations of high salinity and low temperature in the sea. Results: We sampled brown trout from two Danish populations entering different...... was found in trout from the river entering high saline conditions, while a temperature independent up-regulation of both genes in full-strength seawater was found for trout entering marine conditions with lower salinities. Conclusion: Overall our results support the hypothesis that physiologically stressful...

  2. Physiological and pharmacological inductors of HSP70 enhance the antioxidative defense mechanisms of the liver and pancreas in diabetic rats.

    Science.gov (United States)

    Dimitrovska, Maja; Dervisevik, Mirsada; Cipanovska, Natasa; Gerazova, Katerina; Dinevska-Kjovkarovska, Suzana; Miova, Biljana

    2018-02-01

    Heat preconditioning (HP) is a powerful adaptive and protective phenomenon and the heat stress proteins (HSPs) it produces are an important determinant for the development of diabetic complications. Aspirin has been reported to modulate heat shock response in different organisms through increased induction of HSPs and is also known to exert antioxidative and radical scavenging effects in diabetes. We estimated the effect of physiological (heat stress: 45 min at 41 ± 0.5 °C) and pharmacological (aspirin treatment) induction of HSP70 on several parameters of oxidative state in the pancreas and liver of diabetic rats. Diabetes increased HSP70 level and decreased poly(ADP) ribose polymerase (PARP), glutathione (GSH), and glutathione peroxidase (GPx) activities in the pancreas. In the liver, there was reduction of HSP70 level, GSH concentration, and CAT activity, while GPx and GR activity were enhanced. HP of diabetic rats caused an additional increase of HSP70, GSH, and antioxidant enzymes in both organs. Pre-treatment of HP-diabetic animals with aspirin led to an additional increase of PARP and HSP70. Both HP and aspirin, as physiological and pharmacological inductors of HSP70, respectively, enhanced the antioxidative defense mechanisms of the liver and pancreas in diabetic rats.

  3. Anaplasma phagocytophilum MSP4 and HSP70 Proteins Are Involved in Interactions with Host Cells during Pathogen Infection

    Directory of Open Access Journals (Sweden)

    Marinela Contreras

    2017-07-01

    Full Text Available Anaplasma phagocytophilum transmembrane and surface proteins play a role during infection and multiplication in host neutrophils and tick vector cells. Recently, A. phagocytophilum Major surface protein 4 (MSP4 and Heat shock protein 70 (HSP70 were shown to be localized on the bacterial membrane, with a possible role during pathogen infection in ticks. In this study, we hypothesized that A. phagocytophilum MSP4 and HSP70 have similar functions in tick-pathogen and host-pathogen interactions. To address this hypothesis, herein we characterized the role of these bacterial proteins in interaction and infection of vertebrate host cells. The results showed that A. phagocytophilum MSP4 and HSP70 are involved in host-pathogen interactions, with a role for HSP70 during pathogen infection. The analysis of the potential protective capacity of MSP4 and MSP4-HSP70 antigens in immunized sheep showed that MSP4-HSP70 was only partially protective against pathogen infection. This limited protection may be associated with several factors, including the recognition of non-protective epitopes by IgG in immunized lambs. Nevertheless, these antigens may be combined with other candidate protective antigens for the development of vaccines for the control of human and animal granulocytic anaplasmosis. Focusing on the characterization of host protective immune mechanisms and protein-protein interactions at the host-pathogen interface may lead to the discovery and design of new effective protective antigens.

  4. Involvement of serum HSP 70 in Guillain-Barré Syndrome: An exploratory study and a review of current literature

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    Aida Loshaj-Shala

    2015-01-01

    Full Text Available The evolutionary conserved family of heat shock proteins (HSP is responsible for protecting cells against different types of stress. Although the levels of HSP can be readily measured in serum, the levels of HSP 70 in patients Guillain-Barre Syndrome (GBS have not been studied before. To this aim we investigate whether patients with GBS (n=21 had altered serum HSP 70 levels compared to healthy controls (HC, n=9 and to patients affected by other immune disorders such as multifocal motor neuropathy (MMN, n=4 and chronic inflammatory demyelinating polyneuropathy (CIDP, n=6. The highest HSP 70 value (15.78 ± 1.72 ng/mL was found in one patient in the GBS group, although we have found that serum HSP70 levels were significantly higher in 2 out of the 21 GBS patients (9.5%. Hence, it is of interest to underline that the patient with the highest HSP70 level, had also the best recovery rate. Моrе extensive research is required in order to support the hypothesis that HSP 70 serum concentration may be a useful biomarker for the prediction of remission outcome for GBS patients.

  5. Mekanisme Peningkatan Heat Shock Protein-70 pada Kanker Payudara Tikus yang Diradiasi, Pascapemberian Ekstrak Meniran (Phyllanthus niruri (MECHANISM OF INCREASING OF HSP-70 ON IRRADIATED RAT BREAST CANCER, DUE TO APPLICATION OF EXTRACT OF PHYLLANTHUS NIR

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    Bambang Soeprijanto

    2014-10-01

    Full Text Available The used of radiation as a cancer treatment is also proven giving the side effect of damaging thenormal tissue. The extract of Phyllanthus niruri L plant has already been known have an ability to modulatethe immune system. Heat Shock Protein 70 (HSP70 is a protein which can protect other proteins from anydamages. The transcription factor, such as Nuclear factor kappa-light-chain-enhancer of activated B cells(NfkB,is required for protein synthesis. This research was intended to analyze the mechanism, of the immunecompetent cells in expressing the HSP70,through the increase of Nf-kB, at the rat breast cancer tissueunder radiation, due to the application of the extract of P.niruri L plant. An experimental study wasperformed by using the randomized separate pre-test post test controlled group design. The female whiterat (Rattus norvegicus strain Sprague Dawleystrain, undergoing breast cancer due to the application ofcarcinogen materials 7,12-dimethylbenz(aantrasen(DMBA at 20 mg/kg.b.wt, and then the externalradiation at 6Gy given. The treatment group was given the aqueous extract of P.niruri L plant per oral at250 mg/kg b.wt.By immunohistochemistry and t-test study we observe a significant increases in thenumber of cells expressing Nf-kB (p<0.05 and a significant increases in the number of cells expressingHSP70 (p<0.05 at the treatment group. At the regression test, there found to be a stronger influence of NfkBto HSP70 at the treatment group. It is concluded that the mechanism of cell increase expressing theHSP70 at the rat breast cancer tissue under radiation due to the application of the aqueous extract ofP.niruri L plant per oral, is through the increase of cells expressing the Nf-kB.

  6. Both Hsp70 chaperone and Clp protease plastidial systems are required for protection against oxidative stress.

    Science.gov (United States)

    Pulido, Pablo; Llamas, Ernesto; Rodriguez-Concepcion, Manuel

    2017-03-04

    Environmental stress conditions such as high light, extreme temperatures, salinity or drought trigger oxidative stress and eventually protein misfolding in plants. In chloroplasts, chaperone systems refold proteins after stress, while proteases degrade misfolded and aggregated proteins that cannot be refolded. We observed that reduced activity of chloroplast Hsp70 chaperone or Clp protease systems both prevented growth of Arabidopsis thaliana seedlings after treatment with the oxidative agent methyl viologen. Besides showing a role for these particular protein quality control components on the protection against oxidative stress, we provide evidence supporting the existence of a yet undiscovered pathway for Clp-mediated degradation of the damaged proteins.

  7. Prognostic implication of HSPA (HSP70) in breast cancer patients treated with neoadjuvant anthracycline-based chemotherapy.

    Science.gov (United States)

    Nadin, Silvina B; Sottile, Mayra L; Montt-Guevara, Maria M; Gauna, Gisel V; Daguerre, Pedro; Leuzzi, Marcela; Gago, Francisco E; Ibarra, Jorge; Cuello-Carrión, F Darío; Ciocca, Daniel R; Vargas-Roig, Laura M

    2014-07-01

    Neoadjuvant chemotherapy is used in patients with locally advanced breast cancer to reduce tumor size before surgery. Unfortunately, resistance to chemotherapy may arise from a variety of mechanisms. Heat shock proteins (HSPs), which are highly expressed in mammary tumor cells, have been implicated in anticancer drug resistance. In spite of the widely described value of HSPs as molecular markers in cancer, their implications in breast tumors treated with anthracycline-based neoadjuvant chemotherapy has been poorly explored. In this study, we have evaluated, by immunohistochemistry, the expression of HSP27 (HSPB1) and HSP70 (HSPA) in serial biopsies from locally advanced breast cancer patients (n = 60) treated with doxorubicin (DOX)- or epirubicin (EPI)-based monochemotherapy. Serial biopsies were taken at days 1, 3, 7, and 21, and compared with prechemotherapy and surgical biopsies. After surgery, the patients received additional chemotherapy with cyclophosphamide, methotrexate, and 5-fluorouracil. High nuclear HSPB1 and HSPA expressions were found in invasive cells after DOX/EPI administration (P 31 % of the cells) and cytoplasmic HSPA expressions (>11 % of the tumor cells) were associated with better DFS (P = 0.0348 and P = 0.0118, respectively). We conclude that HSPA expression may be a useful prognostic marker in breast cancer patients treated with neoadjuvant DOX/EPI chemotherapy indicating the need to change the administered drugs after surgery for overcoming drug resistance.

  8. Effects of Structurally Related Flavonoids on hsp Gene Expression in Human Promyeloid Leukaemia Cells

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    Herwig O. Gutzeit

    2002-01-01

    Full Text Available Quercetin is a known specific inhibitor of hsp70 synthesis and thus might be a potent agent for enhancing the selective cytotoxicity of heat on tumour cells. A comparative analysis of the effects of quercetin and five structurally related flavonoids on hsp90α, hsp70A, hsp60 and hsp27 gene expression was carried out using human myeloid leukaemia cells (HL-60. The cells were preincubated with 50 μM quercetin, kaempferol, myricetin, taxifolin, isorhamnetin, methylquercetagetin or 0.1 % DMSO (controls for 24 h at 37 °C before heat shock treatment (43 °C for 30 min. Total RNA was isolated from heat-stressed and unstressed cells and analysed by RT PCR. Hsp27 gene expression was inhibited by flavonoids more strongly than other hsp genes investigated in heat stressed as well as in unstressed cells. Among the hsp genes tested, only hsp60 was expressed above control level under the influence of taxifolin. Members of the hsp70 and hsp27 families are highly expressed in breast and lung cancer and leukaemias and they play a role in the acquired resistance to chemotherapy or radiation therapy combined with hyperthermia. Therefore, hsps present potential targets for cancer diagnosis and treatment. The present structure/activity study indicates that position, number and substitution of hydroxyl groups of the B ring and saturation of the C2-C3 bond are important factors affecting flavonoid activity on hsp gene expression. This study could help provide a basis for further design of specific inhibitors of hsp gene expression.

  9. 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.

  10. 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.

  11. Pharmacophore modeling, virtual screening and molecular docking of ATPase inhibitors of HSP70.

    Science.gov (United States)

    Sangeetha, K; Sasikala, R P; Meena, K S

    2017-10-01

    Heat shock protein 70 is an effective anticancer target as it influences many signaling pathways. Hence the study investigated the important pharmacophore feature required for ATPase inhibitors of HSP70 by generating a ligand based pharmacophore model followed by virtual based screening and subsequent validation by molecular docking in Discovery studio V4.0. The most extrapolative pharmacophore model (hypotheses 8) consisted of four hydrogen bond acceptors. Further validation by external test set prediction identified 200 hits from Mini Maybridge, Drug Diverse, SCPDB compounds and Phytochemicals. Consequently, the screened compounds were refined by rule of five, ADMET and molecular docking to retain the best competitive hits. Finally Phytochemical compounds Muricatetrocin B, Diacetylphiladelphicalactone C, Eleutheroside B and 5-(3-{[1-(benzylsulfonyl)piperidin-4-yl]amino}phenyl)- 4-bromo-3-(carboxymethoxy)thiophene-2-carboxylic acid were obtained as leads to inhibit the ATPase activity of HSP70 in our findings and thus can be proposed for further in vitro and in vivo evaluation. Copyright © 2017 Elsevier Ltd. All rights reserved.

  12. Hsp70 Isoforms Are Essential for the Formation of Kaposi's Sarcoma-Associated Herpesvirus Replication and Transcription Compartments.

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    Belinda Baquero-Pérez

    2015-11-01

    Full Text Available Kaposi's sarcoma-associated herpesvirus (KSHV is an oncogenic herpesvirus associated with various AIDS-related malignancies. Like other herpesviruses, multiple processes required for KSHV lytic replication, including viral transcription, viral DNA synthesis and capsid assembly occur in virus-induced intranuclear structures, termed replication and transcription compartments (RTCs. Here we utilised a novel methodology, combining subcellular fractionation and quantitative proteomics, to identify cellular proteins which are recruited to KSHV-induced RTCs and thus play a key role in KSHV lytic replication. We show that several isoforms of the HSP70 chaperone family, Hsc70 and iHsp70, are redistributed from the cytoplasm into the nucleus coinciding with the initial formation of KSHV-induced RTCs. We demonstrate that nuclear chaperone foci are dynamic, initially forming adjacent to newly formed KSHV RTCs, however during later time points the chaperones move within KSHV RTCs and completely co-localise with actively replicating viral DNA. The functional significance of Hsp70 isoforms recruitment into KSHV RTCs was also examined using the specific Hsp70 isoform small molecule inhibitor, VER-155008. Intriguingly, results highlight an essential role of Hsp70 isoforms in the KSHV replication cycle independent of protein stability and maturation. Notably, inhibition of Hsp70 isoforms precluded KSHV RTC formation and RNA polymerase II (RNAPII relocalisation to the viral genome leading to the abolishment of global KSHV transcription and subsequent viral protein synthesis and DNA replication. These new findings have revealed novel mechanisms that regulate KSHV lytic replication and highlight the potential of HSP70 inhibitors as novel antiviral agents.

  13. Survival rate and expression of Heat-shock protein 70 and Frost genes after temperature stress in Drosophila melanogaster lines that are selected for recovery time from temperature coma.

    Science.gov (United States)

    Udaka, Hiroko; Ueda, Chiaki; Goto, Shin G

    2010-12-01

    In this study, we investigated the physiological mechanisms underlying temperature tolerance using Drosophila melanogaster lines with rapid, intermediate, or slow recovery from heat or chill coma that were established by artificial selection or by free recombination without selection. Specifically, we focused on the relationships among their recovery from heat or chill coma, survival after severe heat or cold, and survival enhanced by rapid cold hardening (RCH) or heat hardening. The recovery time from heat coma was not related to the survival rate after severe heat. The line with rapid recovery from chill coma showed a higher survival rate after severe cold exposure, and therefore the same mechanisms are likely to underlie these phenotypes. The recovery time from chill coma and survival rate after severe cold were unrelated to RCH-enhanced survival. We also examined the expression of two genes, Heat-shock protein 70 (Hsp70) and Frost, in these lines to understand the contribution of these stress-inducible genes to intraspecific variation in recovery from temperature coma. The line showing rapid recovery from heat coma did not exhibit higher expression of Hsp70 and Frost. In addition, Hsp70 and Frost transcription levels were not correlated with the recovery time from chill coma. Thus, Hsp70 and Frost transcriptional regulation was not involved in the intraspecific variation in recovery from temperature coma. Copyright © 2010 Elsevier Ltd. All rights reserved.

  14. Generation of the first TCR transgenic mouse with CD4+ T cells recognizing an anti-inflammatory regulatory T cell-inducing HSP70 peptide

    Directory of Open Access Journals (Sweden)

    Manon AA Jansen

    2016-03-01

    Full Text Available Antigen-specific regulatory T cells (Tregs directed at self-antigens are difficult to study since suitable specific tools to isolate and characterize these cells are lacking. A T cell receptor (TCR-transgenic mouse would generate possibilities to study such antigen-specific T cells. As was shown previously, immunization with the mycobacterial heat shock protein 70 derived peptide B29 and its mouse homologues mB29a and mB29b induced anti-inflammatory responses. Furthermore, B29 induced antigen-specific regulatory T cells (Tregs in vivo. To study mB29b specific Tregs, we isolated the TCR from T cell hybridomas generated against mB29b and produced a TCR transgenic mouse that expresses a MHC-class II restricted mB29b-specific TCR. These TCR transgenic CD4+ T cells were found to cross react with the B29 epitope as identified with peptide induced proliferation and IL-2 production. Thus, we have successfully generated a novel mouse model with antigen-specific CD4+ T cells that recognize self and bacterial heat shock protein (Hsp 70 derived peptides. With this novel mouse model, it will be possible to study primary antigen specific T cells with specificity for a regulatory HSP70 T cell epitope. This will enable the isolation and characterization CD4+CD25+ Tregs with a proven specificity. This will provide useful knowledge of the induction, activation and mode of action of Hsp70- specific Tregs, for instance during experimental arthritis.

  15. Antibody to Heat Shock Protein 70 (HSP70 Inhibits Human T-Cell Lymphoptropic Virus Type I (HTLV-I Production by Transformed Rabbit T-Cell Lines

    Directory of Open Access Journals (Sweden)

    Hanan Fallouh

    2012-09-01

    Full Text Available Adult T cell leukemia is a fatal malignant transformation caused by the human T-cell lymphoptropic virus type I (HTLV-I. HTLV-I is only associated with the development of this disease in a small percentage of infected individuals. Using two rabbit transformed T-cell lines; RH/K30 (asymptomatic and RH/K34 (leukemogenic, we have investigated the expression of heat shock proteins (HSP 90 and 70 and the role of anti-HSPs antibodies on virus production. HSPs surface expression was higher on RH/K34 than RH/K30 cells. Heat treatment of cells increased the expression of HSPs proteins and virus production; HSPs augmentation was stabilized after 12 h and virus production reached a maximum between 8 h–12 h then returned to normal level after 24 h of culture. Incubation of cells only with rabbit anti-HSP 70 antibodies prevented virus production specifically in the leukemogenic cell line. The results indicate a relationship between HSP 70 and virus production.

  16. Effect of acute exposure to cadmium on the expression of heat-shock and hormone-nuclear receptor genes in the aquatic midge Chironomus riparius

    Energy Technology Data Exchange (ETDEWEB)

    Planello, R.; Martinez-Guitarte, J.L. [Grupo de Biologia y Toxicologia Ambiental, Facultad de Ciencias, Universidad Nacional de Educacion a Distancia, UNED, Senda del Rey 9, 28040 Madrid (Spain); Morcillo, G., E-mail: gmorcillo@ccia.uned.es [Grupo de Biologia y Toxicologia Ambiental, Facultad de Ciencias, Universidad Nacional de Educacion a Distancia, UNED, Senda del Rey 9, 28040 Madrid (Spain)

    2010-03-01

    Cadmium is a widespread and highly toxic pollutant of particular ecotoxicological relevance for aquatic ecosystems where it accumulates. To identify biomarkers for ecotoxicity monitoring, the effect of cadmium on the expression of different genes related to the stress response as well as to the ecdysone hormone-signalling pathway was studied in the aquatic larvae of Chironomus riparius (Diptera, Chironomidae), a standard test organism in aquatic toxicology testing. Reverse Transcription Polymerase Chain Reaction (RT-PCR) was used to evaluate the effects of acute and short-term cadmium exposures (10 mM CdCl{sub 2}, 12 h and 24 h) on the expression of hsp70, hsc70, hsp90 and hsp40 genes, as well as on that of the ecdysone hormonal-receptor genes (EcR and usp). A significant 3-fold increase in the level of hsp70 gene transcripts was induced by the treatment, whereas neither the other stress genes tested (hsp90 and hsp40) nor the constitutive form of hsp70, hsc70, was affected in the larvae exposed to cadmium. These results show that hsp70 is differentially activated to other environmentally regulated heat-shock genes, and constitutes a biomarker of exposure to this toxic metal. In addition, we also found that cadmium is able to alter the expression of the ecdysone receptor gene (EcR), whose mRNA level is significantly increased whereas usp levels remained unaltered. This finding, evidenced for the first time in invertebrates, supports the view that cadmium has the ability to mimic the effect of the hormone by the activation of the ecdysone nuclear receptor, which may partly explain the endocrine disruption capability that has been previously suggested for this toxic metal. Our research adds to the growing evidence implicating heavy metals, and cadmium in particular, as potential endocrine disruptive agents and may have significant implications for ecological risk assessment of endocrine-disrupting compounds in invertebrates.

  17. Computational analysis of the human HSPH/HSPA/DNAJ family and cloning of a human HSPH/HSPA/DNAJ expression library

    NARCIS (Netherlands)

    Hageman, Jurre; Kampinga, Harm H.

    In this manuscript, we describe the generation of a gene library for the expression of HSP110/HSPH, HSP70/HSPA and HSP40/DNAJ members. First, the heat shock protein (HSP) genes were collected from the gene databases and the gene families were analyzed for expression patterns, heat inducibility,

  18. Evaluation of the toxic potential of calcium carbide in the third instar larvae of transgenic Drosophila melanogaster (hsp70-lacZ)Bg(9).

    Science.gov (United States)

    Danish, Mohd; Fatima, Ambreen; Khanam, Saba; Jyoti, Smita; Rahul; Ali, Fahad; Naz, Falaq; Siddique, Yasir Hasan

    2015-11-01

    In the present study the toxic potential of calcium carbide (CaC2) was studied on the third instar larvae of transgenic Drosophila melanogaster (hsp70-lacZ)Bg(9). The third instar larvae were exposed to 2, 4, 8, 16 and 32×10(-3)g/ml of CaC2 in diet for 24h. The results reveal that the dose 2×10(-3)g/ml was not toxic but the remaining doses showed a dose dependent significant increase in the hsp70 expression, β-galactosidase activity, tissue damage, oxidative stress markers (lipid peroxidation and protein carbonyl content), glutathione-S-transferase activity, expression of Caspase 3 and 9, apoptotic index and DNA damage (midgut cells). A significant reduction as compared to control group in total protein, glutathione content and acetylcholinesterase activity was also observed. The Inductively Coupled Plasma Atomic Emission Spectroscopy analysis (ICPAES) reveals the presence of copper, iron, sodium, aluminium, manganese, calcium, nickel and mercury. The toxic effects of CaC2 in the present study may be attributed to the impurities present in it. Copyright © 2015 Elsevier Ltd. All rights reserved.

  19. Significance of Immunohistochemical Studies of Heat Shock Proteins of the HSP70 Family in the Investigation of Postresuscitative Brain Changes

    Directory of Open Access Journals (Sweden)

    I. V. Ostrova

    2007-01-01

    Full Text Available Objective: to evaluate the immunological responsiveness of various neuronal populations of heat shock proteins of the HSP 70 family and to reveal their association with the magnitude of postresuscitative neuronal morphological changes.Materials and methods. The hypoxia-highly sensitive neuronal populations of pyramidal neurons in the layer of the fifth cerebral sensomotor cortex and hippocampal sectors CA1 and CA4 throughout the postresuscitative period after 12-minute cardiac arrest were subject to a complex immunocytochemical and morphometric study. Their immunological responsiveness to the thermal shock proteins was determined by the indirect peroxidase-antiperoxidase test using the polyclonal antibodies to HSP70. The density and composition of neuronal populations were ascertained by the morphometric assay.Results. The baseline immunological responsiveness of neuronal populations has been demonstrated to be an important factor of its resistance to ischemic lesions. The authors have determined a trend in postresuscitative changes in the immunological responsiveness of neuronal populations to HSP70 and established its association with the development of nerve cell dystrophic changes. They discuss whether complex immunocytochemical and morphometric studies are promising in investigating the mechanisms of postresuscitative brain abnormality.Conclusion. By and large, the findings suggest that the role of HSP70 in maintaining the homeostasis of neuronal populations is more complex and multifactorial, as earlier considered, and it is unlikely to be restricted itself to only merely neuroprotective properties. The high baseline content of HSP70 makes a considerable contribution to the resistance of neuronal populations to ischemia-reperfusion; and the changes in the immunological responsiveness to HSP70 are closely associated with the development of pathological nerve cell changes during a postresuscitative process. 

  20. Detection and identification of Leishmania spp.: application of two hsp70-based PCR-RFLP protocols to clinical samples from the New World.

    Science.gov (United States)

    Montalvo, Ana M; Fraga, Jorge; Tirado, Dídier; Blandón, Gustavo; Alba, Annia; Van der Auwera, Gert; Vélez, Iván Darío; Muskus, Carlos

    2017-07-01

    Leishmaniasis is highly prevalent in New World countries, where several methods are available for detection and identification of Leishmania spp. Two hsp70-based PCR protocols (PCR-N and PCR-F) and their corresponding restriction fragment length polymorphisms (RFLP) were applied for detection and identification of Leishmania spp. in clinical samples recruited in Colombia, Guatemala, and Honduras. A total of 93 cases were studied. The samples were classified into positive or suspected of leishmaniasis according to parasitological criteria. Molecular amplification of two different hsp70 gene fragments and further RFLP analysis for identification of Leishmania species was done. The detection in parasitologically positive samples was higher using PCR-N than PCR-F. In the total of samples studied, the main species identified were Leishmania panamensis, Leishmania braziliensis, and Leishmania infantum (chagasi). Although RFLP-N was more efficient for the identification, RFLP-F is necessary for discrimination between L. panamensis and Leishmania guyanesis, of great importance in Colombia. Unexpectedly, one sample from this country revealed an RFLP pattern corresponding to Leishmania naiffi. Both molecular variants are applicable for the study of clinical samples originated in Colombia, Honduras, and Guatemala. Choosing the better tool for each setting depends on the species circulating. More studies are needed to confirm the presence of L. naiffi in Colombian territory.

  1. Roles of Intramolecular and Intermolecular Interactions in Functional Regulation of the Hsp70 J-protein Co-Chaperone Sis1

    Energy Technology Data Exchange (ETDEWEB)

    Yu, Hyun Young; Ziegelhoffer, Thomas; Osipiuk, Jerzy; Ciesielski, Szymon J.; Baranowski, Maciej; Zhou, Min; Joachimiak, Andrzej; Craig, Elizabeth A.

    2015-04-01

    Unlike other Hsp70 molecular chaperones, those of the eukaryotic cytosol have four residues, EEVD, at heir C-termini. EEVD(Hsp70) binds adaptor proteins of the Hsp90 chaperone system and mitochondrial membrane preprotein receptors, thereby facilitating processing of Hsp70-bound clients through protein folding and translocation pathways. Among J-protein co-chaperones functioning in these pathways, Sis1 is unique, as it also binds the EEVD(Hsp70) motif. However, little is known about the role of the Sis1:EEVD(Hsp70) interaction. We found that deletion of EEVD(Hsp70) abolished the ability of Sis1, but not the ubiquitous J-protein Ydj1, to partner with Hsp70 in in vitro protein refolding. Sis1 co-chaperone activity with Hsp70ΔEEVD was restored upon substitution of a glutamic acid of the J-domain. Structural analysis revealed that this key glutamic acid, which is not present in Ydj1, forms a salt bridge with an arginine of the immediately adjacent glycine-rich region. Thus, restoration of Sis1 in vitro activity suggests that intramolecular interactions between the J-domain and glycine-rich region control co-chaperone activity, which is optimal only when Sis1 interacts with the EEVD(Hsp70) motif. However, we found that disruption of the Sis1:EEVD(Hsp70) interaction enhances the ability of Sis1 to substitute for Ydj1 in vivo. Our results are consistent with the idea that interaction of Sis1 with EEVD(Hsp70) minimizes transfer of Sis1-bound clients to Hsp70s that are primed for client transfer to folding and translocation pathways by their preassociation with EEVD binding adaptor proteins. These interactions may be one means by which cells triage Ydj1- and Sis1-bound clients to productive and quality control pathways, respectively.

  2. Protective effect of capsaicin against methyl methanesulphonate induced toxicity in the third instar larvae of transgenic Drosophila melanogaster (hsp70-lacZ)Bg9.

    Science.gov (United States)

    Khanam, Saba; Fatima, Ambreen; Jyoti, Rahul Smita; Ali, Fahad; Naz, Falaq; Shakya, Barkha; Siddique, Yasir Hasan

    2017-04-01

    Capsaicin (trans-8-methyl-N-vanillyl-6-nonenamide) is the main component in hot peppers, including red chili peppers, jalapenos, and habanero, belonging to the genus Capsicum. Capsaicin is a potent antioxidant that interferes with free radical activities. In the present study, the possible protective effect of capsaicin was studied against methyl methanesulphonate (MMS) induced toxicity in third instar larvae of transgenic Drosophila melanogaster (hsp70-lacZ)Bg 9 . The third instar was allowed to feed on the diet having different doses of capsaicin and MMS separately and in combination. The results suggested that the exposure of third instar larvae to the diet having MMS alone showed significant hsp70 expression as well as tissue DNA and oxidative damage, whereas the larvae feed on the diet having MMS and capsaicin showed a decrease in the toxic effects for 48-h of exposure. In conclusion, capsaicin showed a dose-dependent decrease in the toxic effects induced by MMS in the third instar larvae of transgenic Drosophila melanogaster. Copyright © 2017 China Pharmaceutical University. Published by Elsevier B.V. All rights reserved.

  3. 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

    mature PR complexes. In the present study we observe that a monoclonal antibody specific for p60 can, on the one hand, inhibit formation of mature PR complexes containing heat shock protein 90 (hsp90), p23, and immunophilins and, on the other, enhance recovery of early PR complexes containing hsp70...

  4. Evolutionary Conservation and Emerging Functional Diversity of the Cytosolic Hsp70:J Protein Chaperone Network of Arabidopsis thaliana.

    Science.gov (United States)

    Verma, Amit K; Diwan, Danish; Raut, Sandeep; Dobriyal, Neha; Brown, Rebecca E; Gowda, Vinita; Hines, Justin K; Sahi, Chandan

    2017-06-07

    Heat shock proteins of 70 kDa (Hsp70s) partner with structurally diverse Hsp40s (J proteins), generating distinct chaperone networks in various cellular compartments that perform myriad housekeeping and stress-associated functions in all organisms. Plants, being sessile, need to constantly maintain their cellular proteostasis in response to external environmental cues. In these situations, the Hsp70:J protein machines may play an important role in fine-tuning cellular protein quality control. Although ubiquitous, the functional specificity and complexity of the plant Hsp70:J protein network has not been studied. Here, we analyzed the J protein network in the cytosol of Arabidopsis thaliana and, using yeast genetics, show that the functional specificities of most plant J proteins in fundamental chaperone functions are conserved across long evolutionary timescales. Detailed phylogenetic and functional analysis revealed that increased number, regulatory differences, and neofunctionalization in J proteins together contribute to the emerging functional diversity and complexity in the Hsp70:J protein network in higher plants. Based on the data presented, we propose that higher plants have orchestrated their "chaperome," especially their J protein complement, according to their specialized cellular and physiological stipulations. Copyright © 2017 Verma et al.

  5. Not all J domains are created equal: Implications for the specificity of Hsp40–Hsp70 interactions

    National Research Council Canada - National Science Library

    Hennessy, Fritha; Nicoll, William S; Zimmermann, Richard; Cheetham, Michael E; Blatch, Gregory L

    2005-01-01

    Heat shock protein 40s (Hsp40s) and heat shock protein 70s (Hsp70s) form chaperone partnerships that are key components of cellular chaperone networks involved in facilitating the correct folding of a broad range of client proteins...

  6. Morphological characterization and HSP70-, IGS-based phylogenetic analysis of two microsporidian parasites isolated from Antheraea pernyi.

    Science.gov (United States)

    Wang, Yong; Ru, Yutao; Liu, Wei; Wang, Deyi; Zhou, Jinglin; Jiang, Yiren; Shi, Shenglin; Qin, Li

    2017-03-01

    Two microsporidian isolates were extracted from single infected egg-laying tussah silk moth (Antheraea pernyi) in Liaoning Province, China. The microsporidia were subsequently grown in silk moth larvae, isolated, and subjected to morphological characterization (by light and transmission electron microscopy) and phylogenetic analysis (based on conserved genes). One type of spore was long-axis-oval in shape, measuring 4.71 × 1.95 μm, and the other type was short-axis-oval, measuring 3.64 × 2.17 μm. These dimensions were markedly different from those reported in the spores of the common microsporidia infecting A. pernyi, namely, Nosema pernyi (4.36 × 1.49 μm). A neighbor-joining phylogenetic tree based on HSP70 indicated that these microsporidia belonged to Nosema species and were closely related with Nosema bombycis and Nosema ceranae. Furthermore, in the phylogenetic tree based on the intergenic spacer (IGS) region, the long-axis-oval isolates were closely related and tended to form a clade away from the short-axis-oval isolates and N. pernyi isolates. The microsporidia isolated from A. pernyi clustered in one group. Nosema bombycis, Nosema spodopterae, and Endoreticulatus spp. appeared to be genetically distant from N. pernyi. The two isolates from A. pernyi fell in the Nosema group, but their spores differed from those of the spores of the common A. pernyi parasite N. pernyi, both in morphological and genetic aspects. The two isolates were designated Nosema sp. Ap (L) and Nosema sp. Ap (S). IGS was found to be informative in ascertaining phylogenetic relationships among species, and even closely related strains, of microsporidia.

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

    Science.gov (United States)

    Tomasello, G.; Sciumè, C.; Rappa, F.; Rodolico, V.; Zerilli, M.; Martorana, A.; Cicero, G.; De Luca, R.; Damiani, P.; Accardo, F.M.; Romeo, M.; Farina, F.; Bonaventura, G.; Modica, G.; Zummo, G.; Conway de Macario, E.; Macario, A.J.L.; Cappello, F.

    2011-01-01

    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). PMID:22297444

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

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    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.

  9. Analysis of rice ER-resident J-proteins reveals diversity and functional differentiation of the ER-resident Hsp70 system in plants

    Science.gov (United States)

    Takaiwa, Fumio

    2013-01-01

    The heat shock protein 70 (Hsp70) chaperone system participates in protein folding and quality control of unfolded proteins. To examine the roles of co-chaperones in the rice Hsp70 chaperone system in the endoplasmic reticulum (ER), the functions of six ER-resident J-proteins (OsP58A, OsP58B, OsERdj2, OsERdj3A, OsERdj3B, and OsERdj7) in rice were investigated. The expression of OsP58B, OsERdj3A, and OsERdj3B was predominantly up-regulated in roots subjected to ER stress. This response was mediated by signalling through ATF6 orthologues such as OsbZIP39 and OsbZIP60, but not through the IRE1/OsbZIP50 pathway. A co-immunoprecipitation assay demonstrated that OsP58A, OsP58B, and OsERdj3B preferentially interact with the major OsBiP, OsBiP1, while OsERdj3A interacts preferentially with OsBiP5, suggesting that there are different affinities between OsBiPs and J-proteins. In the endosperm tissue, OsP58A, OsP58B, and OsERdj2 were mainly localized in the ER, whereas OsERdj2 was localized around the outer surfaces of ER-derived protein bodies (PB-Is). Furthermore, OsERdj3A was not expressed in wild-type seeds but was up-regulated in transgenic seeds accumulating human interleukin-7 (hIL-7). Since ERdj3A–green fluorescent protein (GFP) was also detected in vacuoles of callus cells under ER stress conditions, OsERdj3A is a bona fide vacuole-localized protein. OsP58A, OsP58B and OsERdj3A were differentially accumulated in transgenic plants expressing various recombinant proteins. These results reveal the functional diversity of the rice ER-resident Hsp70 system. PMID:24153418

  10. Ubiquitinylation of α-Synuclein by Carboxyl Terminus Hsp70-Interacting Protein (CHIP) Is Regulated by Bcl-2-Associated Athanogene 5 (BAG5)

    Science.gov (United States)

    Chau, Hien; Lozano, Andres M.; Hyman, Bradley T.; McLean, Pamela J.

    2011-01-01

    Parkinson's disease (PD) is a common neurodegenerative condition in which abnormalities in protein homeostasis, or proteostasis, may lead to accumulation of the protein α-synuclein (α-syn). Mutations within or multiplications of the gene encoding α-syn are known to cause genetic forms of PD and polymorphisms in the gene are recently established risk factors for idiopathic PD. α-syn is a major component of Lewy bodies, the intracellular proteinaceous inclusions which are pathological hallmarks of most forms of PD. Recent evidence demonstrates that α-syn can self associate into soluble oligomeric species and implicates these α-syn oligomers in cell death. We have previously shown that carboxyl terminus of Hsp70-interacting protein (CHIP), a co-chaperone molecule with E3 ubiquitin ligase activity, may reduce the levels of toxic α-syn oligomers. Here we demonstrate that α-syn is ubiquitinylated by CHIP both in vitro and in cells. We find that the products from ubiquitinylation by CHIP include both monoubiquitinylated and polyubiquitinylated forms of α-syn. We also demonstrate that CHIP and α-syn exist within a protein complex with the co-chaperone bcl-2-associated athanogene 5 (BAG5) in brain. The interaction of CHIP with BAG5 is mediated by Hsp70 which binds to the tetratricopeptide repeat domain of CHIP and the BAG domains of BAG5. The Hsp70-mediated association of BAG5 with CHIP results in inhibition of CHIP E3 ubiquitin ligase activity and subsequently reduces α-syn ubiquitinylation. Furthermore, we use a luciferase-based protein-fragment complementation assay of α-syn oligomerization to investigate regulation of α-syn oligomers by CHIP in living cells. We demonstrate that BAG5 mitigates the ability of CHIP to reduce α-syn oligomerization and that non-ubiquitinylated α-syn has an increased propensity for oligomerization. Thus, our results identify CHIP as an E3 ubiquitin ligase of α-syn and suggest a novel function for BAG5 as a modulator of CHIP E3

  11. Ubiquitinylation of α-synuclein by carboxyl terminus Hsp70-interacting protein (CHIP is regulated by Bcl-2-associated athanogene 5 (BAG5.

    Directory of Open Access Journals (Sweden)

    Lorraine V Kalia

    2011-02-01

    Full Text Available Parkinson's disease (PD is a common neurodegenerative condition in which abnormalities in protein homeostasis, or proteostasis, may lead to accumulation of the protein α-synuclein (α-syn. Mutations within or multiplications of the gene encoding α-syn are known to cause genetic forms of PD and polymorphisms in the gene are recently established risk factors for idiopathic PD. α-syn is a major component of Lewy bodies, the intracellular proteinaceous inclusions which are pathological hallmarks of most forms of PD. Recent evidence demonstrates that α-syn can self associate into soluble oligomeric species and implicates these α-syn oligomers in cell death. We have previously shown that carboxyl terminus of Hsp70-interacting protein (CHIP, a co-chaperone molecule with E3 ubiquitin ligase activity, may reduce the levels of toxic α-syn oligomers. Here we demonstrate that α-syn is ubiquitinylated by CHIP both in vitro and in cells. We find that the products from ubiquitinylation by CHIP include both monoubiquitinylated and polyubiquitinylated forms of α-syn. We also demonstrate that CHIP and α-syn exist within a protein complex with the co-chaperone bcl-2-associated athanogene 5 (BAG5 in brain. The interaction of CHIP with BAG5 is mediated by Hsp70 which binds to the tetratricopeptide repeat domain of CHIP and the BAG domains of BAG5. The Hsp70-mediated association of BAG5 with CHIP results in inhibition of CHIP E3 ubiquitin ligase activity and subsequently reduces α-syn ubiquitinylation. Furthermore, we use a luciferase-based protein-fragment complementation assay of α-syn oligomerization to investigate regulation of α-syn oligomers by CHIP in living cells. We demonstrate that BAG5 mitigates the ability of CHIP to reduce α-syn oligomerization and that non-ubiquitinylated α-syn has an increased propensity for oligomerization. Thus, our results identify CHIP as an E3 ubiquitin ligase of α-syn and suggest a novel function for BAG5 as a

  12. Differential Targeting of Hsp70 Heat Shock Proteins HSPA6 and HSPA1A with Components of a Protein Disaggregation/Refolding Machine in Differentiated Human Neuronal Cells following Thermal Stress

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    Ian R. Brown

    2017-04-01

    Full Text Available Heat shock proteins (Hsps co-operate in multi-protein machines that counter protein misfolding and aggregation and involve DNAJ (Hsp40, HSPA (Hsp70, and HSPH (Hsp105α. The HSPA family is a multigene family composed of inducible and constitutively expressed members. Inducible HSPA6 (Hsp70B' is found in the human genome but not in the genomes of mouse and rat. To advance knowledge of this little studied HSPA member, the targeting of HSPA6 to stress-sensitive neuronal sites with components of a disaggregation/refolding machine was investigated following thermal stress. HSPA6 targeted the periphery of nuclear speckles (perispeckles that have been characterized as sites of transcription. However, HSPA6 did not co-localize at perispeckles with DNAJB1 (Hsp40-1 or HSPH1 (Hsp105α. At 3 h after heat shock, HSPA6 co-localized with these members of the disaggregation/refolding machine at the granular component (GC of the nucleolus. Inducible HSPA1A (Hsp70-1 and constitutively expressed HSPA8 (Hsc70 co-localized at nuclear speckles with components of the machine immediately after heat shock, and at the GC layer of the nucleolus at 1 h with DNAJA1 and BAG-1. These results suggest that HSPA6 exhibits targeting features that are not apparent for HSPA1A and HSPA8.

  13. Identification of the divergent calmodulin binding motif in yeast Ssb1/Hsp75 protein and in other HSP70 family members

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    R.C. Heinen

    2006-11-01

    Full Text Available Yeast soluble proteins were fractionated by calmodulin-agarose affinity chromatography and the Ca2+/calmodulin-binding proteins were analyzed by SDS-PAGE. One prominent protein of 66 kDa was excised from the gel, digested with trypsin and the masses of the resultant fragments were determined by MALDI/MS. Twenty-one of 38 monoisotopic peptide masses obtained after tryptic digestion were matched to the heat shock protein Ssb1/Hsp75, covering 37% of its sequence. Computational analysis of the primary structure of Ssb1/Hsp75 identified a unique potential amphipathic alpha-helix in its N-terminal ATPase domain with features of target regions for Ca2+/calmodulin binding. This region, which shares 89% similarity to the experimentally determined calmodulin-binding domain from mouse, Hsc70, is conserved in near half of the 113 members of the HSP70 family investigated, from yeast to plant and animals. Based on the sequence of this region, phylogenetic analysis grouped the HSP70s in three distinct branches. Two of them comprise the non-calmodulin binding Hsp70s BIP/GR78, a subfamily of eukaryotic HSP70 localized in the endoplasmic reticulum, and DnaK, a subfamily of prokaryotic HSP70. A third heterogeneous group is formed by eukaryotic cytosolic HSP70s containing the new calmodulin-binding motif and other cytosolic HSP70s whose sequences do not conform to those conserved motif, indicating that not all eukaryotic cytosolic Hsp70s are target for calmodulin regulation. Furthermore, the calmodulin-binding domain found in eukaryotic HSP70s is also the target for binding of Bag-1 - an enhancer of ADP/ATP exchange activity of Hsp70s. A model in which calmodulin displaces Bag-1 and modulates Ssb1/Hsp75 chaperone activity is discussed.

  14. The small-molecule kinase inhibitor D11 counteracts 17-AAG-mediated up-regulation of HSP70 in brain cancer cells.

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    Susanne Schaefer

    Full Text Available Many types of cancer express high levels of heat shock proteins (HSPs that are molecular chaperones regulating protein folding and stability ensuring protection of cells from potentially lethal stress. HSPs in cancer cells promote survival, growth and spreading even in situations of growth factors deprivation by associating with oncogenic proteins responsible for cell transformation. Hence, it is not surprising that the identification of potent inhibitors of HSPs, notably HSP90, has been the primary research focus, in recent years. Exposure of cancer cells to HSP90 inhibitors, including 17-AAG, has been shown to cause resistance to chemotherapeutic treatment mostly attributable to induction of the heat shock response and increased cellular levels of pro-survival chaperones. In this study, we show that treatment of glioblastoma cells with 17-AAG leads to HSP90 inhibition indicated by loss of stability of the EGFR client protein, and significant increase in HSP70 expression. Conversely, co-treatment with the small-molecule kinase inhibitor D11 leads to suppression of the heat shock response and inhibition of HSF1 transcriptional activity. Beside HSP70, Western blot and differential mRNA expression analysis reveal that combination treatment causes strong down-regulation of the small chaperone protein HSP27. Finally, we demonstrate that incubation of cells with both agents leads to enhanced cytotoxicity and significantly high levels of LC3-II suggesting autophagy induction. Taken together, results reported here support the notion that including D11 in future treatment regimens based on HSP90 inhibition can potentially overcome acquired resistance induced by the heat shock response in brain cancer cells.

  15. The p53/HSP70 inhibitor, 2-phenylethynesulfonamide, causes oxidative stress, unfolded protein response and apoptosis in rainbow trout cells

    Energy Technology Data Exchange (ETDEWEB)

    Zeng, Fanxing; Tee, Catherine; Liu, Michelle [Department of Biology, University of Waterloo, Waterloo, Ontario N2L 3G1 (Canada); Sherry, James P. [Aquatic Contaminants Research Division, Environment Canada, Burlington, Ontario L7R 4A6 (Canada); Dixon, Brian; Duncker, Bernard P. [Department of Biology, University of Waterloo, Waterloo, Ontario N2L 3G1 (Canada); Bols, Niels C., E-mail: ncbols@uwaterloo.ca [Department of Biology, University of Waterloo, Waterloo, Ontario N2L 3G1 (Canada)

    2014-01-15

    Highlights: •2-Phenylethynesulfonamide (PES) is an inhibitor of p53 and HSP 70 in mammals. •In the fish epithelial cell line, RTgill-W1, PES enhanced ROS generation and was cytotoxic. •RTgill-W1 death was by apoptosis and blocked by the anti-oxidant N-acetylcysteine. •This is the first report linking PES-induced cell death to ROS. •With this background PES should be useful for studying fish cell survival pathways. -- Abstract: The effect of 2-phenylethynesulfonamide (PES), which is a p53 and HSP70 inhibitor in mammalian cells, was studied on the rainbow trout (Oncorhynchus mykiss) gill epithelial cell line, RTgill-W1, in order to evaluate PES as a tool for understanding the cellular survival pathways operating in fish. As judged by three viability assays, fish cells were killed by 24 h exposures to PES, but cell death was blocked by the anti-oxidant N-acetylcysteine (NAC). Cell death had several hallmarks of apoptosis: DNA laddering, nuclear fragmentation, Annexin V staining, mitochondrial membrane potential decline, and caspases activation. Reactive oxygen species (ROS) production peaked in several hours after the addition of PES and before cell death. HSP70 and BiP levels were higher in cultures treated with PES for 24 h, but this was blocked by NAC. As well, PES treatment caused HSP70, BiP and p53 to accumulate in the detergent-insoluble fraction, and this too was prevented by NAC. Of several possible scenarios to explain the results, the following one is the simplest. PES enhances the generation of ROS, possibly by inhibiting the anti-oxidant actions of p53 and HSP70. ER stress arises from the ROS and from PES inhibiting the chaperone activities of HSP70. The ER stress in turn initiates the unfolded protein response (UPR), but this fails to restore ER homeostasis so proteins aggregate and cells die. Despite these multiple actions, PES should be useful for studying fish cellular survival pathways.

  16. Annona muricata leaves accelerate wound healing in rats via involvement of Hsp70 and antioxidant defence.

    Science.gov (United States)

    Moghadamtousi, Soheil Zorofchian; Rouhollahi, Elham; Hajrezaie, Maryam; Karimian, Hamed; Abdulla, Mahmood Ameen; Kadir, Habsah Abdul

    2015-06-01

    Annona muricata, a member of the Annonaceae family, is commonly known as soursop and graviola. The leaves of this tropical fruit tree are widely used in folk medicine against skin diseases and abscesses, however there is no scientific evidence justifying the use of A. muricata leaves. The aim of the present study is to evaluate the wound healing potential of ethyl acetate extract of A. muricata leaves (EEAM) towards excisional wound models in rats. Sprague Dawley rats (24) were randomly divided into four groups, viz. (A) vehicle control, (B) low dose of EEAM (5% w/w), (C) high dose of EEAM (10% w/w) and (D) positive control with excisional wound created on the neck area. Wounds were topically dressed twice a day for 15 days. On the 15th day, animals were sacrificed and then processed for immunohistochemical and histological evaluations, including Hematoxylin & Eosin and Masson Trichrome stainings. The activity of antioxidants, namely catalase, glutathione peroxidase and superoxide dismutase, and malondialdehyde (MDA) was measured in wound tissue homogenate. Macroscopic and microscopic analysis of wounds demonstrated a significant wound healing activity shown by EEAM at two doses. Treatment of wounds with ointment containing EEAM caused significant surge in antioxidants activities and decrease in the MDA level of wound tissues compared with vehicle control. The immunohistochemical evaluation revealed conspicuous up-regulation of Hsp70 in treated wounds with EEAM, suggesting the anti-inflammatory effect of EEAM. EEAM exhibited a promising wound healing potential towards excisional wound models in rats. Copyright © 2015. Published by Elsevier Ltd.

  17. Effects of heavy metals on ultrastructure and HSP70s induction in the aquatic moss Leptodictyum riparium Hedw.

    Science.gov (United States)

    Esposito, S; Sorbo, S; Conte, B; Basile, A

    2012-04-01

    The effects of heavy metals, both toxic (Pb, Cd) and essential (Cu, Zn) on the ultrastructure and the induction of Heat Shock Protein 70 (HSP70) have been studied in the aquatic moss Leptodictyum riparium Hedw. In vitro cultured L. riparium was treated with different heavy metals, both toxic, as cadmium or lead; and essential microelements such as Copper or Zinc concentrations ranging from 10(-3) to 10(-6) M to investigate both ultrastructural damage and HSP induction. TEM observations showed that sub-lethal concentrations of heavy metals caused only slight changes, largely localized in the chloroplasts. Among all the heavy metals tested, cadmium caused the most severe modifications. Heavy metals caused the decrease of the soluble protein content and the enhancement of proteins reacting versus HSP70 antibodies, suggesting that molecular chaperons might be involved in the resistance to toxic effects of lead, cadmium, copper and zinc. Therefore, the induction of HSP70 in L. riparium would confer a higher resistance to pollutants under stressful conditions lethal for other mosses and higher plant species. These results suggest that the moss L. riparium can tolerate heavy metals stress without incurring severe cellular/subcellular damage. Therefore it can be used as a useful indicator of heavy metals accumulation.

  18. Effects of heated hydrotherapy on muscle HSP70 and glucose metabolism in old and young vervet monkeys.

    Science.gov (United States)

    Kavanagh, Kylie; Davis, Ashely T; Jenkins, Kurt A; Flynn, D Mickey

    2016-07-01

    Increasing heat shock protein 70 (HSP70) in aged and/or insulin-resistant animal models confers benefits to healthspan and lifespan. Heat application to increase core temperature induces HSPs in metabolically important tissues, and preliminary human and animal data suggest that heated hydrotherapy is an effective method to achieve increased HSPs. However, safety concerns exist, particularly in geriatric medicine where organ and cardiovascular disease commonly will preexist. We evaluated young vervet monkeys compared to old, insulin-resistant vervet monkeys (Chlorocebus aethiops sabaeus) in their core temperatures, glucose tolerance, muscle HSP70 level, and selected safety biomarkers after 10 sessions of hot water immersions administered twice weekly. Hot water immersion robustly induced the heat shock response in muscles. We observed that heat-treated old and young monkeys have significantly higher muscle HSP70 than control monkeys and treatment was without significant adverse effects on organ or cardiovascular health. Heat therapy improved pancreatic responses to glucose challenge and tended to normalize glucose excursions. A trend for worsened blood pressure and glucose values in the control monkeys and improved values in heat-treated monkeys were seen to support further investigation into the safety and efficacy of this intervention for metabolic syndrome or diabetes in young or old persons unable to exercise.

  19. Hsp70 oligomerization is mediated by an interaction between the interdomain linker and the substrate-binding domain.

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    Francesco A Aprile

    Full Text Available Oligomerization in the heat shock protein (Hsp 70 family has been extensively documented both in vitro and in vivo, although the mechanism, the identity of the specific protein regions involved and the physiological relevance of this process are still unclear. We have studied the oligomeric properties of a series of human Hsp70 variants by means of nanoelectrospray ionization mass spectrometry, optical spectroscopy and quantitative size exclusion chromatography. Our results show that Hsp70 oligomerization takes place through a specific interaction between the interdomain linker of one molecule and the substrate-binding domain of a different molecule, generating dimers and higher-order oligomers. We have found that substrate binding shifts the oligomerization equilibrium towards the accumulation of functional monomeric protein, probably by sequestering the helical lid sub-domain needed to stabilize the chaperone: substrate complex. Taken together, these findings suggest a possible role of chaperone oligomerization as a mechanism for regulating the availability of the active monomeric form of the chaperone and for the control of substrate binding and release.

  20. Selection of an Anticalin® against the membrane form of Hsp70 via bacterial surface display and its theranostic application in tumour models.

    Science.gov (United States)

    Friedrich, Lars; Kornberger, Petra; Mendler, Claudia T; Multhoff, Gabriele; Schwaiger, Markus; Skerra, Arne

    2017-11-27

    We describe the selection of Anticalins against a common tumour surface antigen, human Hsp70, using functional display on live E. coli cells as fusion with a truncated EspP autotransporter. While found intracellularly in normal cells, Hsp70 is frequently exposed in a membrane-bound state on the surface of tumour cells and, even more pronounced, in metastases or after radiochemotherapy. Employing a recombinant Hsp70 fragment comprising residues 383-548 as target, Anticalins were selected from a naive bacterial library. The Anticalin with the highest affinity (KD = 13 nM), as determined towards recombinant full-length Hsp70 by real-time surface plasmon resonance analysis, was improved to KD = 510 pM by doped random mutagenesis and another cycle of E. coli surface display, followed by rational combination of mutations. This Anticalin, which recognizes a linear peptide epitope located in the interdomain linker of Hsp70, was demonstrated to specifically bind Hsp70 in its membrane-associated form in immunofluorescence microscopy and via flow cytometry using the FaDu cell line, which is positive for surface Hsp70. The radiolabeled and PASylated Anticalin revealed specific tumour accumulation in xenograft mice using PET imaging. Furthermore, after enzymatic coupling to the protein toxin gelonin, the Anticalin showed potent cytotoxicity on FaDu cells in vitro.

  1. Involvement of p27CIP/KIP in HSP25 or inducible HSP70 mediated adaptive response by low dose radiation

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    Seo, Hang Rhan; Lee, Yoon Jin; Bae, Sangwoo; Lee, Yun Sil; Lee, Su Jae [Korea Institute of Radiological and Medical Science, Seoul (Korea, Republic of); Chung, Hee Yong [Hanyang University, Seoul (Korea, Republic of); Baek, Min [Ministry of Science and Technology, Gwacheon (Korea, Republic of)

    2006-04-15

    ThermoResistant (TR) clones of Radiation-Induced Fibrosarcoma (RIF) cells have been reported to show an adaptive response to 1cGy of low dose radiation, and HSP25 and inducible HSP70 are involved in this process. In this study, to further elucidate the mechanism by which HSP25 and inducible HSP70 regulate the adaptive response, HSP25 or inducible HSP70 overexpressed RIF cells were irradiated with 1cGy and the cell cycle was analyzed. HSP25 or inducible HSP70 overexpressed cells together with TR cells showed increased G1 phase after 1cGy irradiation, while RIF cells did not. [{sup 3}H]-Thymidine and BrdU incorporation also indicated that both HSP25 and inducible HSP70 are involved in G1 arrest after 1cGy irradiation. Molecular analysis revealed upregulation of p27Cip/Kip protein in HSP25 and inducible HSP70 overexpressed cells, and cotransfection of p27Cip/Kip antisense abolished the induction of the adaptive response and 1cGy-mediated G1 arrest. The above results indicate that induction of an adaptive response by HSP25 and inducible HSP70 is mediated by upregulation of p27Cip/Kip protein, resulting in low dose radiation-induced G1 arrest.

  2. Evaluation of the Toxic Potential of Graphene Copper Nanocomposite (GCNC) in the Third Instar Larvae of Transgenic Drosophila melanogaster (hsp70-lacZ)Bg9

    Science.gov (United States)

    Siddique, Yasir Hasan; Fatima, Ambreen; Jyoti, Smita; Naz, Falaq; Rahul; Khan, Wasi; Singh, Braj Raj; Naqvi, Alim Hussain

    2013-01-01

    Graphene, a two-dimensional carbon sheet with single-atom thickness, have attracted the scientific world for its potential applications in various field including the biomedical areas. In the present study the graphene copper nanocomposite (GCNC) was synthesized, characterized and evaluated for its toxic potential on third instar larvae of transgenic Drosophila melanogaster (hsp70-lacZ)Bg9. The synthesized GCNC was analyzed by X-ray diffraction (XRD), scanning/transmission electron microscopy (SEM/TEM), atomic force microscopy (AFM), and fourier transform infrared spectroscopy (FTIR). The GCNC in 0.1% DMSO was sonicated for 10 min and the final concentration of 0.033, 0.099, 0.199 and 3.996 µg/µl of diet were established. The third instar larvae were allowed to feed on it separately for 24 and 48 hrs. The hsp70 expression was measured by O-nitrophenyl-β-D-galactopyranoside assay, tissue damage by trypan blue exclusion test and β-galactosidase activity was monitored by in situ histochemical β-galactosidase staining. Oxidative stress was monitored by performing lipid peroxidation assay and total protein estimation. Ethidium bromide/acridine orange staining was performed on midgut cells for apoptotic index and the comet assay was performed for the DNA damage. The results of the present study showed that the exposure of 0.199 and 3.996 µg/µl of GCNC were toxic for 24 hr of exposure and for 48 hr of exposure: 0.099, 0.199 and 3.996 µg/µl of GCNC was toxic. The dose of 0.033 µg/µl of GCNC showed no toxic effects on its exposure to the third instar larvae for 24 hr as well as 48 hrs. This dose can be considered as No Observed Adverse Effect Level (NOAEL). PMID:24339891

  3. Evaluation of the toxic potential of graphene copper nanocomposite (GCNC in the third instar larvae of transgenic Drosophila melanogaster (hsp70-lacZBg(9..

    Directory of Open Access Journals (Sweden)

    Yasir Hasan Siddique

    Full Text Available Graphene, a two-dimensional carbon sheet with single-atom thickness, have attracted the scientific world for its potential applications in various field including the biomedical areas. In the present study the graphene copper nanocomposite (GCNC was synthesized, characterized and evaluated for its toxic potential on third instar larvae of transgenic Drosophila melanogaster (hsp70-lacZBg(9 . The synthesized GCNC was analyzed by X-ray diffraction (XRD, scanning/transmission electron microscopy (SEM/TEM, atomic force microscopy (AFM, and fourier transform infrared spectroscopy (FTIR. The GCNC in 0.1% DMSO was sonicated for 10 min and the final concentration of 0.033, 0.099, 0.199 and 3.996 µg/µl of diet were established. The third instar larvae were allowed to feed on it separately for 24 and 48 hrs. The hsp70 expression was measured by O-nitrophenyl-β-D-galactopyranoside assay, tissue damage by trypan blue exclusion test and β-galactosidase activity was monitored by in situ histochemical β-galactosidase staining. Oxidative stress was monitored by performing lipid peroxidation assay and total protein estimation. Ethidium bromide/acridine orange staining was performed on midgut cells for apoptotic index and the comet assay was performed for the DNA damage. The results of the present study showed that the exposure of 0.199 and 3.996 µg/µl of GCNC were toxic for 24 hr of exposure and for 48 hr of exposure: 0.099, 0.199 and 3.996 µg/µl of GCNC was toxic. The dose of 0.033 µg/µl of GCNC showed no toxic effects on its exposure to the third instar larvae for 24 hr as well as 48 hrs. This dose can be considered as No Observed Adverse Effect Level (NOAEL.

  4. Evaluation of the toxic potential of graphene copper nanocomposite (GCNC) in the third instar larvae of transgenic Drosophila melanogaster (hsp70-lacZ)Bg(9.).

    Science.gov (United States)

    Siddique, Yasir Hasan; Fatima, Ambreen; Jyoti, Smita; Naz, Falaq; Rahul; Khan, Wasi; Singh, Braj Raj; Naqvi, Alim Hussain

    2013-01-01

    Graphene, a two-dimensional carbon sheet with single-atom thickness, have attracted the scientific world for its potential applications in various field including the biomedical areas. In the present study the graphene copper nanocomposite (GCNC) was synthesized, characterized and evaluated for its toxic potential on third instar larvae of transgenic Drosophila melanogaster (hsp70-lacZ)Bg(9) . The synthesized GCNC was analyzed by X-ray diffraction (XRD), scanning/transmission electron microscopy (SEM/TEM), atomic force microscopy (AFM), and fourier transform infrared spectroscopy (FTIR). The GCNC in 0.1% DMSO was sonicated for 10 min and the final concentration of 0.033, 0.099, 0.199 and 3.996 µg/µl of diet were established. The third instar larvae were allowed to feed on it separately for 24 and 48 hrs. The hsp70 expression was measured by O-nitrophenyl-β-D-galactopyranoside assay, tissue damage by trypan blue exclusion test and β-galactosidase activity was monitored by in situ histochemical β-galactosidase staining. Oxidative stress was monitored by performing lipid peroxidation assay and total protein estimation. Ethidium bromide/acridine orange staining was performed on midgut cells for apoptotic index and the comet assay was performed for the DNA damage. The results of the present study showed that the exposure of 0.199 and 3.996 µg/µl of GCNC were toxic for 24 hr of exposure and for 48 hr of exposure: 0.099, 0.199 and 3.996 µg/µl of GCNC was toxic. The dose of 0.033 µg/µl of GCNC showed no toxic effects on its exposure to the third instar larvae for 24 hr as well as 48 hrs. This dose can be considered as No Observed Adverse Effect Level (NOAEL).

  5. Toxic Potential of Synthesized Graphene Zinc Oxide Nanocomposite in the Third Instar Larvae of Transgenic Drosophila melanogaster (hsp70-lacZBg9

    Directory of Open Access Journals (Sweden)

    Yasir Hasan Siddique

    2014-01-01

    Full Text Available In the present study the graphene zinc oxide nanocomposite (GZNC was synthesized, characterized, and evaluated for its toxic potential on third instar larvae of transgenic Drosophila melanogaster (hsp70-lacZBg9. The synthesized GZNC was characterized by X-ray diffraction (XRD, Fourier transform infrared spectroscopy (FTIR, thermogravimetric analysis (TGA, scanning electron microscopy (SEM, and transmission electron microscopy (TEM. The GZNC in 0.1% dimethyl sulphoxide (DMSO was sonicated for 10 minutes and the final concentrations 0.033, 0.099, 0.199, and 3.996 μg/μL of diet were established. The third instar larvae were allowed to feed on it separately for 24 and 48 hr. The hsp70 expression was measured by o-nitrophenyl-β-D-galactopyranoside assay, tissue damage was measured by trypan blue exclusion test, and β-galactosidase activity was monitored by in situ histochemical β-galactosidase staining. Oxidative stress was monitored by performing lipid peroxidation assay and total protein estimation. Ethidium bromide/acridine orange staining was performed on midgut cells for apoptotic index and the comet assay was performed for the DNA damage. The results of the present study showed that the exposure of 0.199 and 3.996 μg/μL of GZNC was toxic for both 24 hr and 48 hr of exposure. The doses of 0.033 μg/μL and 0.099 of GZNC showed no toxic effects on its exposure to the third instar larvae for 24 hr as well as 48 hr of duration.

  6. Enhancing insecticidal efficacy of baculovirus by early expressing an insect neurotoxin, LqhIT2, in infected Trichoplusia ni larvae.

    Science.gov (United States)

    Jinn, Tzyy-Rong; Tu, Wu-Chun; Lu, Cha-I; Tzen, Jason T C

    2006-10-01

    LqhIT(2), an insect specific neurotoxin from the venom of Leiurus quinquestriatus hebraeus, has been demonstrated to improve insecticidal efficacy of Autographa californica nuclear polyhedrosis virus (AcMNPV). A polyhedrin-positive recombinant AcMNPVvAcP(hsp70)EGFP/P(pag90)IT(2) was engineered for larvae to express the enhanced green fluorescence protein (EGFP) and LqhIT(2) under the control of P(hsp70) and P(pag90) promoters, respectively. This would allow a visual observation of the viral infection and an improvement of the insecticidal efficacy. The insecticidal activity of this recombinant baculovirus, a wild type AcMNPV and four other recombinant baculoviruses, was evaluated and compared in terms of mortality, body weight, median lethal time (LT(50)), and median lethal concentration (LC(50)). Insecticidal efficacy was unaltered when treated with vAcP(hsp70)EGFP, moderately improved when infected by vAcP(10)IT(2) (a P(10)-promoted LqhIT ( 2 ) gene), and significantly elevated when treated with vAcP(pag90)IT(2) or vAcP(hsp70)EGFP/P(pag90)IT(2). No apparent difference was observed in insecticidal efficacy when additional EGFP was expressed as a visible marker. These results suggest that recombinant AcMNPV vAcP(hsp70)EGFP/P(pag90)IT(2) may be used as an effective insecticide against Trichoplusia ni and other lepidopterous insect pests.

  7. Molecular cloning of the heat-shock cognate 70 (Hsc70) gene from the two-spotted spider mite, Tetranychus urticae, and its expression in response to heat shock and starvation.

    Science.gov (United States)

    Shim, Jae-Kyoung; Jung, Duck-Oung; Park, Jae-Woo; Kim, Dong-Woo; Ha, Dae-Myung; Lee, Kyeong-Yeoll

    2006-01-01

    We isolated a heat shock cognate 70 (hsc70) gene from the two-spotted spider mite, Tetranychus urticae, a serious agricultural pest. The hsc70 cDNA is 2275 bp and contains a 1962 bp open reading frame. The translated amino acid sequence consists of 654 residues with a calculated molecular mass of 71,275 Da and an isoelectronic point (pI) of 5.52. It also contains the highly conserved functional motifs of the Hsp70 family. A comparison of the deduced amino acid sequence shows a high identity (81-84%) with Hsp70s/Hsc70s of insects but the highest identity is with mussel Hsc71 (86%). Northern blot hybridization indicates that the hsc70 transcript level of female adults is higher than that of male adults. We evaluated the response of hsc70 gene to stresses from temperature and starvation. The level of hsc70 mRNA was not significantly changed by heat and cold shocks nor by recovery after the shocks. However, the hsc70 mRNA level was decreased by food restriction of female mites. Analysis of nucleotide and deduced amino acid sequences of hsc70 gene from T. urticae suggests that it is a member of heat shock cognate 70 gene in the highly conserved Hsp70 family but that its expression is influenced by food restriction rather than thermal stress. This is the first molecular analysis of a heat shock protein gene in an acarid.

  8. Vitrification affects nuclear maturation and gene expression of immature human oocytes

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    Abbas Shahedi

    2017-02-01

    Full Text Available Background: Vitrification of oocytes is a fast-freezing technique, which may affect the quality of the human oocyte, and consequently affects the embryo development, pregnancy and birth. The aim of the current study was to investigate the consequence of in-vitro vitrification on maturation status of immature human oocytes, additionally, expression levels of stress, and apoptosis related genes. Materials and Methods: The total of 213 human immature oocytes which routinely discarded from assisted reproduction clinics were collected and divided into two groups including: (I fresh germinal vesicle (GV oocytes (n=106 (matured in-vitro  (fIVM , and  (II GV oocytes (n=107 that initially vitrified, then matured in  in-vitro (vIVM. After 36 hours of incubation, the oocytes were evaluated for nuclear maturation and expression level of DNA methyltransferase (DNMT1, stress related genes (Sod1 and Hsp70, and apoptotic related genes (Bax and Bcl-2 by quantitative Real-Time PCR. Results: Oocyte maturation rates were reduced in vIVM compared to fIVM oocytes (P=0.001. The expression of stress (Sod1 and Hsp70, and apoptotic-related genes (Bax and Bcl-2 in vIVM were significantly higher compared to the fIVM group. Additionally, pro-apoptotic gene up-regulated 4.3 times more than anti-apoptotic gene in vIVM oocyte. However, DNMT1 gene expression was reduced in vIVM oocyte (P = 0.047. Conclusions: The low survival rate of vitrified In-vitro matured GV oocytes could definitely be explained by the alterations of their gene expression profile. 

  9. ANTI-HSP60 and ANTI-HSP70 antibody levels and micro/ macrovascular complications in type 1 diabetes: the EURODIAB Study

    DEFF Research Database (Denmark)

    Gruden, G.; Bruno, G.; Chaturvedi, N.

    2009-01-01

    diabetes is associated with a greatly increased risk of micro- and macrovascular complications. Therefore, we investigated whether anti-HSP60 and anti-HSP70 antibody levels were associated with micro- and macrovascular complications in type 1 diabetic patients. DESIGN: A cross-sectional nested case......-control study from the EURODIAB Study of 531 type 1 diabetic patients was performed. SUBJECTS: Cases (n = 363) were defined as those with one or more complications of diabetes; control subjects (n = 168) were all those with no evidence of any complication. We measured anti-HSP60 and anti-HSP70 antibody levels...... and investigated their cross-sectional associations with diabetic complications. RESULTS: Anti-HSP70 antibody levels were significantly greater in control than in case subjects, whereas anti-HSP60 antibody levels were similar in the two groups. In logistic regression analysis, anti-HSP70 levels in the upper...

  10. A fragment-based approach applied to a highly flexible target: Insights and challenges towards the inhibition of HSP70 isoforms

    Science.gov (United States)

    Jones, Alan M.; Westwood, Isaac M.; Osborne, James D.; Matthews, Thomas P.; Cheeseman, Matthew D.; Rowlands, Martin G.; Jeganathan, Fiona; Burke, Rosemary; Lee, Diane; Kadi, Nadia; Liu, Manjuan; Richards, Meirion; McAndrew, Craig; Yahya, Norhakim; Dobson, Sarah E.; Jones, Keith; Workman, Paul; Collins, Ian; van Montfort, Rob L. M.

    2016-10-01

    The heat shock protein 70s (HSP70s) are molecular chaperones implicated in many cancers and of significant interest as targets for novel cancer therapies. Several HSP70 inhibitors have been reported, but because the majority have poor physicochemical properties and for many the exact mode of action is poorly understood, more detailed mechanistic and structural insight into ligand-binding to HSP70s is urgently needed. Here we describe the first comprehensive fragment-based inhibitor exploration of an HSP70 enzyme, which yielded an amino-quinazoline fragment that was elaborated to a novel ATP binding site ligand with different physicochemical properties to known adenosine-based HSP70 inhibitors. Crystal structures of amino-quinazoline ligands bound to the different conformational states of the HSP70 nucleotide binding domain highlighted the challenges of a fragment-based approach when applied to this particular flexible enzyme class with an ATP-binding site that changes shape and size during its catalytic cycle. In these studies we showed that Ser275 is a key residue in the selective binding of ATP. Additionally, the structural data revealed a potential functional role for the ATP ribose moiety in priming the protein for the formation of the ATP-bound pre-hydrolysis complex by influencing the conformation of one of the phosphate binding loops.

  11. Non-lethal heat shock increased Hsp70 and immune protein transcripts but not Vibrio tolerance in the white-leg shrimp.

    Directory of Open Access Journals (Sweden)

    Nguyen Hong Loc

    Full Text Available Non-lethal heat shock boosts bacterial and viral disease tolerance in shrimp, possibly due to increases in endogenous heat shock protein 70 (Hsp70 and/or immune proteins. To further understand the mechanisms protecting shrimp against infection, Hsp70 and the mRNAs encoding the immune-related proteins prophenoloxidase (proPO, peroxinectin, penaeidin, crustin and hemocyanin were studied in post-larvae of the white-leg shrimp Litopenaeus vannamei, following a non-lethal heat shock. As indicated by RT-qPCR, a 30 min abrupt heat shock increased Hsp70 mRNA in comparison to non-heated animals. Immunoprobing of western blots and quantification by ELISA revealed that Hsp70 production after heat shock was correlated with enhanced Hsp70 mRNA. proPO and hemocyanin mRNA levels were augmented, whereas peroxinectin and crustin mRNA levels were unchanged following non-lethal heat shock. Penaeidin mRNA was decreased by all heat shock treatments. Thirty min abrupt heat shock failed to improve survival of post-larvae in a standardized challenge test with Vibrio harveyi, indicating that under the conditions of this study, L. vannamei tolerance to Vibrio infection was influenced neither by Hsp70 accumulation nor the changes in the immune-related proteins, observations dissimilar to other shrimp species examined.

  12. HSP70 immune reactivity and TUNEL positivity in the liver of toluene-inhaled and melatonin-treated rats.

    Science.gov (United States)

    Tas, Ufuk; Ogeturk, Murat; Kuloglu, Tuncay; Sapmaz, Hilal Irmak; Kocaman, Nevin; Zararsiz, Ismail; Sarsilmaz, Mustafa

    2013-07-01

    Toluene is a clear, colorless and volatile hydrocarbon that is metabolized in liver, produced free oxygen radicals and can mediate cellular damage. Melatonin which is a pineal gland hormone is a very potent antioxidant. It can make the cellular membrane more durable against oxidative attacks and protect nuclear DNA from oxidative damage. This study aimed to investigate heat shock protein (HSP)70 immune reactivity and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) positivity (apoptotic activity) in the liver of toluene-inhaled and melatonin-treated rats. A total of 21 adult male Wistar albino rats were divided at random into 3 equal groups. Animals in group I were designated as control. The rats in group II were exposed to toluene (3000 ppm/1 h/day) for 30 days, while the rats in group III were treated with melatonin (10 mg/kg/day, intraperitoneally) plus toluene inhalation. At the end of the 30-day experimental period, all rats were killed by decapitation. Then the liver tissues of rats were removed and tissue specimens were embedded in paraffin blocks. The specimens were stained with periodic acid-schiff (PAS) following routine histological procedures. Sections obtained from paraffin blocks were used for immune detection of TUNEL and HSP70. In light microscopic observations of tissues from toluene-inhaled rats, massive hepatocyte degeneration, ballooning degeneration and decreased PAS positivity were observed. Increased TUNEL positivity and HSP70 immune reactivity were determined in toluene-inhaled group and melatonin treatment decreased all these adverse effects.

  13. A mitochondrial homolog of bacterial GrpE interacts with mitochondrial hsp70 and is essential for viability.

    OpenAIRE

    Bolliger, L; Deloche, O; Glick, B S; Georgopoulos, C; Jenö, P; Kronidou, N; Horst, M; Morishima, N; Schatz, G.

    1994-01-01

    Mitochondrial hsp70 (mhsp70) is located in the matrix and an essential component of the mitochondrial protein import system. To study the function of mhsp70 and to identify possible partner proteins we constructed a yeast strain in which all mhsp70 molecules carry a C-terminal hexa-histidine tag. The tagged mhsp70 appears to be functional in vivo. When an ATP depleted mitochondrial extract was incubated with a nickel-derivatized affinity resin, the resin bound not only mhsp70, but also a 23 k...

  14. Developmental competence and expression profile of genes in buffalo (Bubalus bubalis) oocytes and embryos collected under different environmental stress.

    Science.gov (United States)

    Sadeesh, E M; Sikka, P; Balhara, A K; Balhara, S

    2016-12-01

    The study examined the effects of different environmental stress on developmental competence and the relative abundance (RA) of various gene transcripts in oocytes and embryos of buffalo. Oocytes collected during cold period (CP) and hot period (HP) were matured, fertilized and cultured in vitro to blastocyst hatching stage. The mRNA expression patterns of genes implicated in developmental competence (OCT-4, IGF-2R and GDF-9), heat shock (HSP-70.1), oxidative stress (MnSOD), metabolism (GLUT-1), pro-apoptosis (BAX) and anti-apoptosis (BCL-2) were evaluated in immature and matured oocytes as well as in pre-implantation stage embryos. Oocytes reaching MII stage, cleavage rates, blastocyst yield and hatching rates increased (P < 0.05) during the CP. In MII oocytes and 2-cell embryos, the RA of OCT-4, IGF-2R, GDF-9, MnSOD and GLUT-1 decreased (P < 0.05) during the HP. In 4-cell embryos, the RA of OCT-4, IGF-2R and BCL-2 decreased (P < 0.05) in the HP, whereas GDF-9 increased (P < 0.05). In 8-to 16-cell embryos, the RA of OCT-4 and BCL-2 decreased (P < 0. 05) in the HP, whereas HSP-70.1 and BAX expression increased (P < 0.05). In morula and blastocyst, the RA of OCT-4, IGF-2R and MnSOD decreased (P < 0.05) during the HP, whereas HSP-70.1 was increased (P < 0.05). In conclusion, deleterious seasonal effects induced at the GV-stage carry-over to subsequent embryonic developmental stages and compromise oocyte developmental competence and quality of developed blastocysts.

  15. Brain death induces renal expression of heme oxygenase-1 and heat shock protein 70

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    van Dullemen Leon FA

    2013-01-01

    Full Text Available Abstract Background Kidneys derived from brain dead donors have lower graft survival and higher graft-function loss compared to their living donor counterpart. Heat Shock Proteins (HSP are a large family of stress proteins involved in maintaining cell homeostasis. We studied the role of stress-inducible genes Heme Oxygenase-1 (HO-1, HSP27, HSP40, and HSP70 in the kidney following a 4 hour period of brain death. Methods Brain death was induced in rats (n=6 by inflating a balloon catheter in the epidural space. Kidneys were analysed for HSPs using RT-PCR, Western blotting, and immunohistochemistry. Results RT-PCR data showed a significant increase in gene expression for HO-1 and HSP70 in kidneys of brain dead rats. Western blotting revealed a massive increase in HO-1 protein in brain dead rat kidneys. Immunohistochemistry confirmed these findings, showing extensive HO-1 protein expression in the renal cortical tubules of brain dead rats. HSP70 protein was predominantly increased in renal distal tubules of brain dead rats treated for hypotension. Conclusion Renal stress caused by brain death induces expression of the cytoprotective genes HO-1 and HSP70, but not of HSP27 and HSP40. The upregulation of these cytoprotective genes indicate that renal damage occurs during brain death, and could be part of a protective or recuperative mechanism induced by brain death-associated stress.

  16. Cryo-thermal therapy elicits potent anti-tumor immunity by inducing extracellular Hsp70-dependent MDSC differentiation

    Science.gov (United States)

    Zhu, Jun; Zhang, Yan; Zhang, Aili; He, Kun; Liu, Ping; Xu, Lisa X.

    2016-01-01

    Achieving control of metastatic disease is a long-sought goal in cancer therapy. Treatments that encourage a patient’s own immune system are bringing new hopes in reaching such a goal. In clinic, local hyperthermia and cryoablation have been explored to induce anti-tumor immune responses against tumors. We have also developed a novel therapeutic modality of cryo-thermal treatment by alternating liquid nitrogen (LN2) cooling and radio frequency (RF) heating, and better therapeutic effect was achieved in treating metastatic cancer in animal model. In this study, we investigated the mechanism of systemic immune response elicited by cryo-thermal therapy. In the 4T1 murine mammary carcinoma model, we found that local cryo-thermal therapy resulted in a considerable reduction of distant lung metastases, and improved long-term survival. Moreover, results of tumor re-challenge experiments indicated generation of a strong tumor-specific immune memory after the local treatment of primary tumors. Our further study indicated that cryo-thermal therapy caused an elevated extracellular release of Hsp70. Subsequently, Hsp70 induced differentiation of MDSCs into mature DCs, contributing to the relief of MDSCs-mediated immunosuppression and ultimately the activation of strong anti-tumor immune response. Our findings reveal new insight into the mechanism of robust therapeutic effects of cryo-thermal therapy against metastatic cancers. PMID:27256519

  17. Polydopamine-Encapsulated Fe3O4 with an Adsorbed HSP70 Inhibitor for Improved Photothermal Inactivation of Bacteria.

    Science.gov (United States)

    Liu, Dongdong; Ma, Liyi; Liu, Lidong; Wang, Lu; Liu, Yuxin; Jia, Qi; Guo, Quanwei; Zhang, Ge; Zhou, Jing

    2016-09-21

    Photothermal treatment, a new approach for inactivation of bacteria and pathogens that does not depend on traditional therapeutic approaches, has recently received much attention. In this study, a new type of nanoplatform (PDA@Fe3O4 + PES) was fabricated by using polydopamine (PDA, a photothermal conversion agent) to encapsulate Fe3O4 (a magnetic nanoparticle) and support 2-phenylethynesulfonamide (PES, an inhibitor of heat shock protein 70 (HSP70)). Upon near-infrared light irradiation, the increased temperature weakens π-π and hydrogen bonding interactions, and PES is released from the PDA@Fe3O4 + PES. The released PES inhibits the function of HSP70, reducing bacterial tolerance to photothermal therapy and improving the therapeutic effect against infectious bacterial pathogens. After treatment, PDA@Fe3O4 + PES can be recovered using the magnetic property of the Fe3O4 cores. Consequently, PDA@Fe3O4 + PES possesses the potential to be a recyclable photothermal agent for enhanced photothermal bacterial inactivation without causing secondary pollution.

  18. Cryo-thermal therapy elicits potent anti-tumor immunity by inducing extracellular Hsp70-dependent MDSC differentiation

    Science.gov (United States)

    Zhu, Jun; Zhang, Yan; Zhang, Aili; He, Kun; Liu, Ping; Xu, Lisa X.

    2016-06-01

    Achieving control of metastatic disease is a long-sought goal in cancer therapy. Treatments that encourage a patient’s own immune system are bringing new hopes in reaching such a goal. In clinic, local hyperthermia and cryoablation have been explored to induce anti-tumor immune responses against tumors. We have also developed a novel therapeutic modality of cryo-thermal treatment by alternating liquid nitrogen (LN2) cooling and radio frequency (RF) heating, and better therapeutic effect was achieved in treating metastatic cancer in animal model. In this study, we investigated the mechanism of systemic immune response elicited by cryo-thermal therapy. In the 4T1 murine mammary carcinoma model, we found that local cryo-thermal therapy resulted in a considerable reduction of distant lung metastases, and improved long-term survival. Moreover, results of tumor re-challenge experiments indicated generation of a strong tumor-specific immune memory after the local treatment of primary tumors. Our further study indicated that cryo-thermal therapy caused an elevated extracellular release of Hsp70. Subsequently, Hsp70 induced differentiation of MDSCs into mature DCs, contributing to the relief of MDSCs-mediated immunosuppression and ultimately the activation of strong anti-tumor immune response. Our findings reveal new insight into the mechanism of robust therapeutic effects of cryo-thermal therapy against metastatic cancers.

  19. Polyglutamine-rich suppressors of huntingtin toxicity act upstream of Hsp70 and Sti1 in spatial quality control of amyloid-like proteins.

    Directory of Open Access Journals (Sweden)

    Katie J Wolfe

    Full Text Available Protein conformational maladies such as Huntington Disease are characterized by accumulation of intracellular and extracellular protein inclusions containing amyloid-like proteins. There is an inverse correlation between proteotoxicity and aggregation, so facilitated protein aggregation appears cytoprotective. To define mechanisms for protective protein aggregation, a screen for suppressors of nuclear huntingtin (Htt103Q toxicity was conducted. Nuclear Htt103Q is highly toxic and less aggregation prone than its cytosolic form, so we identified suppressors of cytotoxicity caused by Htt103Q tagged with a nuclear localization signal (NLS. High copy suppressors of Htt103Q-NLS toxicity include the polyQ-domain containing proteins Nab3, Pop2, and Cbk1, and each suppresses Htt toxicity via a different mechanism. Htt103Q-NLS appears to inactivate the essential functions of Nab3 in RNA processing in the nucleus. Function of Pop2 and Cbk1 is not impaired by nuclear Htt103Q, as their respective polyQ-rich domains are sufficient to suppress Htt103Q toxicity. Pop2 is a subunit of an RNA processing complex and is localized throughout the cytoplasm. Expression of just the Pop2 polyQ domain and an adjacent proline-rich stretch is sufficient to suppress Htt103Q toxicity. The proline-rich domain in Pop2 resembles an aggresome targeting signal, so Pop2 may act in trans to positively impact spatial quality control of Htt103Q. Cbk1 accumulates in discrete perinuclear foci and overexpression of the Cbk1 polyQ domain concentrates diffuse Htt103Q into these foci, which correlates with suppression of Htt toxicity. Protective action of Pop2 and Cbk1 in spatial quality control is dependent upon the Hsp70 co-chaperone Sti1, which packages amyloid-like proteins into benign foci. Protein:protein interactions between Htt103Q and its intracellular neighbors lead to toxic and protective outcomes. A subset of polyQ-rich proteins buffer amyloid toxicity by funneling toxic

  20. Hsp70 and lipid peroxide levels following heat stress in Xeropicta derbentina (Krynicki 1836) (Gastropoda, Pulmonata) with regard to different colour morphs.

    Science.gov (United States)

    Dieterich, A; Troschinski, S; Schwarz, S; Di Lellis, M A; Henneberg, A; Fischbach, U; Ludwig, M; Gärtner, U; Triebskorn, R; Köhler, H-R

    2015-01-01

    Terrestrial snails which live under dry and hot conditions need efficient mechanisms of adaptation to counteract the problems of desiccation and over-heating. A profoundly heat tolerant snail species is the Mediterranean Xeropicta derbentina, exhibiting different shell colour morphs ranging from pale white to darkly banded. Considering that dark-pigmented snails are believed to have a disadvantage due to faster heating, we investigated possible differences in the stress markers Hsp70 and lipid peroxideation between four pre-defined colour morphs which were exposed to different temperatures for eight hours. The highest Hsp70 levels were observed in response to 38-40 °C. Levels decreased when this temperature was exceeded. Snails of a pre-defined colour category 3 (with a large black band at the umbilicus side of the shell) showed the most prominent Hsp70 response. Lipid peroxideation levels also showed a maximum at 38 °C but displayed a second peak at rather high temperatures at which the Hsp70 level already had decreased (45-48 °C). Particularly pure white snails (category 1) and the most pigmented ones (category 4) were found to have different levels of lipid peroxidation at 38 °C and 45 °C compared to the other morphs. A hypothesis involving a combined two-phase defence mechanism, to which both, the Hsp70 protection system and the antioxidant defence system, may contribute, is discussed.

  1. Bag1 Co-chaperone Promotes TRC8 E3 Ligase-dependent Degradation of Misfolded Human Ether a Go-Go-related Gene (hERG) Potassium Channels.

    Science.gov (United States)

    Hantouche, Christine; Williamson, Brittany; Valinsky, William C; Solomon, Joshua; Shrier, Alvin; Young, Jason C

    2017-02-10

    Cardiac long QT syndrome type 2 is caused by mutations in the human ether a go-go-related gene (hERG) potassium channel, many of which cause misfolding and degradation at the endoplasmic reticulum instead of normal trafficking to the cell surface. The Hsc70/Hsp70 chaperones assist the folding of the hERG cytosolic domains. Here, we demonstrate that the Hsp70 nucleotide exchange factor Bag1 promotes hERG degradation by the ubiquitin-proteasome system at the endoplasmic reticulum to regulate hERG levels and channel activity. Dissociation of hERG complexes containing Hsp70 and the E3 ubiquitin ligase CHIP requires the interaction of Bag1 with Hsp70, but this does not involve the Bag1 ubiquitin-like domain. The interaction with Bag1 then shifts hERG degradation to the membrane-anchored E3 ligase TRC8 and its E2-conjugating enzyme Ube2g2, as determined by siRNA screening. TRC8 interacts through the transmembrane region with hERG and decreases hERG functional expression. TRC8 also mediates degradation of the misfolded hERG-G601S disease mutant, but pharmacological stabilization of the mutant structure prevents degradation. Our results identify TRC8 as a previously unknown Hsp70-independent quality control E3 ligase for hERG. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  2. Expression of Mytilus immune genes in response to experimental challenges varied according to the site of collection.

    Science.gov (United States)

    Li, Hui; Venier, Paola; Prado-Alvárez, María; Gestal, Camino; Toubiana, Mylène; Quartesan, Rosita; Borghesan, Fabio; Novoa, Beatriz; Figueras, Antonio; Roch, Philippe

    2010-04-01

    Mussels live in diverse coastal environments experience various physical, chemical and biological conditions, which they counteract with functional adjustments and heritable adaptive changes. In order to investigate possible differences in immune system capabilities, we analyzed by qPCR the expression levels of 4 immune genes (defensin, mytilin B, myticin B, lysozyme) and HSP70 in the Mediterranean mussel, Mytilus galloprovincialis collected in 3 European farming areas {Atlantic Ocean-Ría de Vigo-Spain (RV), French Mediterranean Gulf of Lion-Palavas-Prévost lagoon (PP) and Northern Adriatic Sea-Venice-Italy (VI)} in response to one injection of one of the 3 bacterial species (Vibrio splendidus LGP32, Vibrio anguillarum, Micrococcus lysodeikticus), and to heat shock or cold stress. We confirmed that the 5 genes are constitutively expressed in hemocytes, defensin being the less expressed, myticin B the highest. As suspected, the same gene resulted differently expressed according to mussel group, with the biggest difference being for HSP70 and lysozyme and lowest expression of all the 5 genes in mussels from RV. In addition, gene expression levels varied according to the challenge. Most frequent effect of bacterial injections was down-regulation, especially for mytilin B and myticin B. Heat shock enhanced transcript levels, particularly in mussels from RV, whereas cold stress had no effect. In situ hybridization of labelled probes on mussel hemocytes indicated that bacterial injections did not change the mRNA patterns of defensin and myticin B whereas mytilin B mRNA almost disappeared. In conclusion, these results demonstrated that constitutive level, nature and intensity of immune gene expression regulations strongly depended from mussel group, and support the concept of gene-environment interactions. Copyright 2009 Elsevier Ltd. All rights reserved.

  3. Cathepsin B is involved in the heat shock induced cardiomyocytes apoptosis as well as the anti-apoptosis effect of HSP-70.

    Science.gov (United States)

    Hsu, Shu-Fen; Hsu, Chuan-Chih; Cheng, Bor-Chih; Lin, Cheng-Hsien

    2014-11-01

    Cathepsin B is one of the major lysosomal cysteine proteases that plays an important role in apoptosis. Herein, we investigated whether Cathepsin B is involved in cardiomyocyte apoptosis caused by hyperthermic injury (HI) and heat shock protein (HSP)-70 protects these cells from HI-induced apoptosis mediated by Cathepsin. HI was produced in H9C2 cells by putting them in a circulating 43 °C water bath for 120 min, whereas preinduction of HSP-70 was produced in H9C2 cells by mild heat preconditioning (or putting them in 42 °C water bath for 30 min) 8 h before the start of HI. It was found that HI caused both cardiomyocyte apoptosis and increased Cathepsin B activity in H9C2 cells. E-64-c, in addition to reducing Cathepsin B activity, significantly attenuated HI-induced cardiomyocyte apoptosis (evidenced by increased apoptotic cell numbers, increased tuncated Bid (t-Bid), increased cytochrome C, increased caspase-9/-3, and decreased Bcl-2/Bax) in H9C2 cells. In addition, preinduction of HSP-70 by mild heat preconditioning or inhibition of HSP-70 by Tripolide significantly attenuated or exacerbated respectively both the cardiomyocyte apoptosis and increased Cathepsin B activity in H9C2 cells. Furthermore, the beneficial effects of pre-induction of HSP-70 by mild heat production in reducing both cardiomyocyte apoptosis and increased Cathepsin B activity caused by HI can be significantly reduced by Triptolide preconditioning. These results indicate that Cathepsin B is involved in HI-induced cardiomyocyte apoptosis in H9C2 cells and HSP-70 protects these cells from HI-induced cardiomyocyte apoptosis through Cathepsin B pathways.

  4. DnaK as Antibiotic Target: Hot Spot Residues Analysis for Differential Inhibition of the Bacterial Protein in Comparison with the Human HSP70.

    Directory of Open Access Journals (Sweden)

    Federica Chiappori

    Full Text Available DnaK, the bacterial homolog of human Hsp70, plays an important role in pathogens survival under stress conditions, like antibiotic therapies. This chaperone sequesters protein aggregates accumulated in bacteria during antibiotic treatment reducing the effect of the cure. Although different classes of DnaK inhibitors have been already designed, they present low specificity. DnaK is highly conserved in prokaryotes (identity 50-70%, which encourages the development of a unique inhibitor for many different bacterial strains. We used the DnaK of Acinetobacter baumannii as representative for our analysis, since it is one of the most important opportunistic human pathogens, exhibits a significant drug resistance and it has the ability to survive in hospital environments. The E.coli DnaK was also included in the analysis as reference structure due to its wide diffusion. Unfortunately, bacterial DnaK and human Hsp70 have an elevated sequence similarity. Therefore, we performed a differential analysis of DnaK and Hsp70 residues to identify hot spots in bacterial proteins that are not present in the human homolog, with the aim of characterizing the key pharmacological features necessary to design selective inhibitors for DnaK. Different conformations of DnaK and Hsp70 bound to known inhibitor-peptides for DnaK, and ineffective for Hsp70, have been analysed by molecular dynamics simulations to identify residues displaying stable and selective interactions with these peptides. Results achieved in this work show that there are some residues that can be used to build selective inhibitors for DnaK, which should be ineffective for the human Hsp70.

  5. Heat Shock Protein 70 and 90 Genes in the Harmful Dinoflagellate Cochlodinium polykrikoides: Genomic Structures and Transcriptional Responses to Environmental Stresses

    Directory of Open Access Journals (Sweden)

    Ruoyu Guo

    2015-01-01

    Full Text Available The marine dinoflagellate Cochlodinium polykrikoides is responsible for harmful algal blooms in aquatic environments and has spread into the world’s oceans. As a microeukaryote, it seems to have distinct genomic characteristics, like gene structure and regulation. In the present study, we characterized heat shock protein (HSP 70/90 of C. polykrikoides and evaluated their transcriptional responses to environmental stresses. Both HSPs contained the conserved motif patterns, showing the highest homology with those of other dinoflagellates. Genomic analysis showed that the CpHSP70 had no intron but was encoded by tandem arrangement manner with separation of intergenic spacers. However, CpHSP90 had one intron in the coding genomic regions, and no intergenic region was found. Phylogenetic analyses of separate HSPs showed that CpHSP70 was closely related with the dinoflagellate Crypthecodinium cohnii and CpHSP90 with other Gymnodiniales in dinoflagellates. Gene expression analyses showed that both HSP genes were upregulated by the treatments of separate algicides CuSO4 and NaOCl; however, they displayed downregulation pattern with PCB treatment. The transcription of CpHSP90 and CpHSP70 showed similar expression patterns under the same toxicant treatment, suggesting that both genes might have cooperative functions for the toxicant induced gene regulation in the dinoflagellate.

  6. Studies on the protective efficacy and immunogenicity of Hsp70 and Hsp83 based vaccine formulations in Leishmania donovani infected BALB/c mice.

    Science.gov (United States)

    Kaur, Jaspreet; Kaur, Tejinder; Kaur, Sukhbir

    2011-07-01

    Visceral leishmaniasis, a chronic systemic infection, is the major cause of morbidity and mortality in many parts of world. The current drugs for the treatment of leishmaniasis are toxic, expensive, difficult to administer and becoming ineffective due to the emergence of drug resistance. In the absence of effective treatment, vaccination remains the only hope for control of the disease. We have evaluated the protective efficacy of two heat shock proteins (Hsp70 and Hsp83) in combination with two different adjuvants (MPLA and ALD) in Leishmania donovani infected inbred BALB/c mice. The proteins were isolated by SDS-PAGE and the mice were immunized subcutaneously with Hsp70+Hsp83, Hsp70+Hsp83+ALD and Hsp70+Hsp83+MPLA. These were challenged with 10(7) promastigotes of L. donovani. The animals were sacrificed on 30, 60 and 90 days post challenge for the assessment of parasite load and generation of cellular and humoral immune responses. The vaccines induced a strong protective response against experimental visceral leishmaniasis as shown by reduced parasite load in liver of all immunized groups as compared to the infected controls. The vaccines also led to the augmentation of DTH responses, increased levels of IgG2a, IFN-γ and IL-2. Both the adjuvants raised significantly the level of protection imparted by the proteins but MPLA was more effective in comparison to ALD. Copyright © 2011 Elsevier B.V. All rights reserved.

  7. Stress Proteins (hsp70, hsp60) Induced in Isopods and Nematodes by Field Exposure to Metals in a Gradient near Avonmouth, UK

    NARCIS (Netherlands)

    Arts, M.S.J.; Schill, R.O.; Knigge, T.; Eckwert, H.; Kammenga, J.E.; Köhler, H.R.

    2004-01-01

    Heat shock proteins (hsps) are potential biomarkers for monitoring environmental pollution. In this study, the use of hsps as biomarkers in field bioassays was evaluated in terrestrial invertebrates exposed to a metal gradient near Avonmouth, UK. We investigated the hsp70 response in resident and

  8. Daily and seasonal changes in heat exposure and the Hsp70 level of individuals from a field population of Xeropicta derbentina (Krynicki 1836) (Pulmonata, Hygromiidae) in Southern France.

    Science.gov (United States)

    Dieterich, A; Fischbach, U; Ludwig, M; Di Lellis, M A; Troschinski, S; Gärtner, U; Triebskorn, R; Köhler, H-R

    2013-07-01

    The Mediterranean land snail Xeropicta derbentina forms huge populations in Southern France. In order to characterize heat exposure and the induction of the 70-kD heat shock protein (Hsp70) response system during the life cycle of this snail, a selected population from the Vaucluse area, Provence, was investigated encompassing the issues of morphological life cycle parameters (shell size and colouration), the daily courses of heat exposure at different heights above the ground, of shell temperature, and that of the individual Hsp70 levels. The study covered all four seasons of the year 2011. Snails were found to be annual, reaching their final size in August. The shell colouration pattern showed high variation in juveniles (spring) with a strong tendency towards becoming uniformly white at old age in autumn. In all seasons, ambient air temperature decreased with increasing distance from the ground surface during daytime while remaining constantly low in the night. Overall, the Hsp70 level of individuals followed the ambient temperature during diurnal and seasonal variations. Correlation analysis revealed a positive association of individual shell temperature and Hsp70 level for the most part of the life cycle of the snails until late summer, whereas a negative correlation was found for aged animals indicating senescence effects on the capacity of the stress response system.

  9. Efeito do estresse térmico agudo sobre os níveis da proteína e RNA mensageiro da Hsp70, em fígado e cérebro de pintos de corte de diferentes linhagens

    Directory of Open Access Journals (Sweden)

    Dionello Nelson José Laurino

    2001-01-01

    Full Text Available Oitenta pintos de corte de duas linhagens, pescoço pelado (Na/na e Hubbard-Pettersen, entre 2 e 5 dias de idade, foram expostos a estresse térmico agudo (36-37°C, durante cinco horas. Foram avaliados temperatura cloacal e peso corporal individuais ao início e final do período de estresse térmico agudo. Os pintos foram sacrificados ao final e amostras de fígado e cérebro foram coletadas e analisadas por Western Blotting e Northern Blotting, para quantificação da proteína e RNA mensageiro da Hsp70, respectivamente. Os resultados apresentaram maiores níveis protéicos de Hsp70 no tecido cerebral do que no hepático. Antes do estresse térmico, não houve diferenças de linhagens para expressão e síntese de Hsp70, em ambos os tecidos. Após o estresse térmico, as aves estressadas apresentaram maiores níveis protéicos de Hsp70, quando o tecido analisado foi o fígado (para as pescoço pelado nas idades de 4 e 5 dias, e menores níveis, em relação às controles, quando o tecido analisado foi o cérebro (para as Pescoço pelado na idade de 2 dias. O tamanho do transcrito de RNA mensageiro de Hsp70 foi de 2,7 kb. Os resultados do presente experimento sugerem que, para o tecido cerebral, a indução de Hsp70 ocorreu em níveis transcripcional e traducional e para o tecido hepático foi detectada apenas alteração em nível traducional, para ambas as linhagens.

  10. [Potential protective role of nitric oxide and Hsp70 linked to functional foods in the atherosclerosis].

    Science.gov (United States)

    Camargo, Alejandra B; Manucha, Walter

    Atherosclerosis, one of the main pathologic entities considered epidemic and a worldwide public health problem, is currently under constant review as regards its basic determining mechanisms and therapeutic possibilities. In this regard, all patients afflicted with the disease exhibit mitochondrial dysfunction, oxidative stress and inflammation. Interestingly, nitric oxide - a known vasoactive messenger gas - has been closely related to the inflammatory, oxidative and mitochondrial dysfunctional process that characterizes atherosclerosis. In addition, it has recently been demonstrated that alterations in the bioavailability of nitric oxide would induce the expression of heat shock proteins. This agrees with the use of functional foods as a strategy to prevent both vascular aging and the development of atherosclerosis. Finally, a greater knowledge regarding the mechanisms implied in the development of atherosclerosis will enable proposing new and possible hygiene, health and therapeutic interventions. Copyright © 2016 Sociedad Española de Arteriosclerosis. Publicado por Elsevier España, S.L.U. All rights reserved.

  11. Dancing through Life: Molecular Dynamics Simulations and Network-Centric Modeling of Allosteric Mechanisms in Hsp70 and Hsp110 Chaperone Proteins.

    Science.gov (United States)

    Stetz, Gabrielle; Verkhivker, Gennady M

    2015-01-01

    Hsp70 and Hsp110 chaperones play an important role in regulating cellular processes that involve protein folding and stabilization, which are essential for the integrity of signaling networks. Although many aspects of allosteric regulatory mechanisms in Hsp70 and Hsp110 chaperones have been extensively studied and significantly advanced in recent experimental studies, the atomistic picture of signal propagation and energetics of dynamics-based communication still remain unresolved. In this work, we have combined molecular dynamics simulations and protein stability analysis of the chaperone structures with the network modeling of residue interaction networks to characterize molecular determinants of allosteric mechanisms. We have shown that allosteric mechanisms of Hsp70 and Hsp110 chaperones may be primarily determined by nucleotide-induced redistribution of local conformational ensembles in the inter-domain regions and the substrate binding domain. Conformational dynamics and energetics of the peptide substrate binding with the Hsp70 structures has been analyzed using free energy calculations, revealing allosteric hotspots that control negative cooperativity between regulatory sites. The results have indicated that cooperative interactions may promote a population-shift mechanism in Hsp70, in which functional residues are organized in a broad and robust allosteric network that can link the nucleotide-binding site and the substrate-binding regions. A smaller allosteric network in Hsp110 structures may elicit an entropy-driven allostery that occurs in the absence of global structural changes. We have found that global mediating residues with high network centrality may be organized in stable local communities that are indispensable for structural stability and efficient allosteric communications. The network-centric analysis of allosteric interactions has also established that centrality of functional residues could correlate with their sensitivity to mutations

  12. The Hsp70-like StkA functions between T4P and Dif signaling proteins as a negative regulator of exopolysaccharide in Myxococcus xanthus

    Directory of Open Access Journals (Sweden)

    Pamela L. Moak

    2015-02-01

    Full Text Available Myxococcus xanthus displays a form of surface motility known as social (S gliding. It is mediated by the type IV pilus (T4P and requires the exopolysaccharide (EPS to function. It is clear that T4P retraction powers S motility. EPS on a neighboring cell or deposited on a gliding surface is proposed to anchor the distal end of a pilus and trigger T4P retraction at its proximal end. Inversely, T4P has been shown to regulate EPS production upstream of the Dif signaling pathway. Here we describe the isolation of two Tn insertions at the stk locus which had been known to play roles in cellular cohesion and formation of cell groups. An insertion in stkA (MXAN_3474 was identified based on its ability to restore EPS to a pilA deletion mutant. The stkA encodes a DnaK or Hsp70 homolog and it is upstream of stkB (MXAN_3475 and stkC (MXAN_3476. A stkB insertion was identified in a separate genetic screen because it eliminated EPS production of an EPS+ parental strain. Our results with in-frame deletions of these three stk genes indicated that the stkA mutant produced increased level of EPS while stkB and stkC mutants produced less EPS relative to the wild type. S motility and developmental aggregation were affected by deletions of stkA and stkB but only minimally by the deletion of stkC. Genetic epistasis indicated that StkA functions downstream of T4P but upstream of the Dif proteins as a negative regulator of EPS production in M. xanthus.

  13. The Hsp70-like StkA functions between T4P and Dif signaling proteins as a negative regulator of exopolysaccharide in Myxococcus xanthus.

    Science.gov (United States)

    Moak, Pamela L; Black, Wesley P; Wallace, Regina A; Li, Zhuo; Yang, Zhaomin

    2015-01-01

    Myxococcus xanthus displays a form of surface motility known as social (S) gliding. It is mediated by the type IV pilus (T4P) and requires the exopolysaccharide (EPS) to function. It is clear that T4P retraction powers S motility. EPS on a neighboring cell or deposited on a gliding surface is proposed to anchor the distal end of a pilus and trigger T4P retraction at its proximal end. Inversely, T4P has been shown to regulate EPS production upstream of the Dif signaling pathway. Here we describe the isolation of two Tn insertions at the stk locus which had been known to play roles in cellular cohesion and formation of cell groups. An insertion in stkA (MXAN_3474) was identified based on its ability to restore EPS to a pilA deletion mutant. The stkA encodes a DnaK or Hsp70 homolog and it is upstream of stkB (MXAN_3475) and stkC (MXAN_3476). A stkB insertion was identified in a separate genetic screen because it eliminated EPS production of an EPS(+) parental strain. Our results with in-frame deletions of these three stk genes indicated that the stkA mutant produced increased level of EPS while stkB and stkC mutants produced less EPS relative to the wild type. S motility and developmental aggregation were affected by deletions of stkA and stkB but only minimally by the deletion of stkC. Genetic epistasis indicated that StkA functions downstream of T4P but upstream of the Dif proteins as a negative regulator of EPS production in M. xanthus.

  14. Insulin Promotes Survival of Amyloid-Beta Oligomers Neuroblastoma Damaged Cells via Caspase 9 Inhibition and Hsp70 Upregulation

    Directory of Open Access Journals (Sweden)

    M. Di Carlo

    2010-01-01

    Full Text Available Alzheimer's disease (AD and type 2 diabetes are connected in a way that is still not completely understood, but insulin resistance has been implicated as a risk factor for developing AD. Here we show an evidence that insulin is capable of reducing cytotoxicity induced by Amyloid-beta peptides (A-beta in its oligomeric form in a dose-dependent manner. By TUNEL and biochemical assays we demonstrate that the recovery of the cell viability is obtained by inhibition of intrinsic apoptotic program, triggered by A-beta and involving caspase 9 and 3 activation. A protective role of insulin on mitochondrial damage is also shown by using Mito-red vital dye. Furthermore, A-beta activates the stress inducible Hsp70 protein in LAN5 cells and an overexpression is detectable after the addition of insulin, suggesting that this major induction is the necessary condition to activate a cell survival program. Together, these results may provide opportunities for the design of preventive and therapeutic strategies against AD.

  15. An androgen receptor NH2-terminal conserved motif interacts with the COOH terminus of the Hsp70-interacting protein (CHIP).

    Science.gov (United States)

    He, Bin; Bai, Suxia; Hnat, Andrew T; Kalman, Rebecca I; Minges, John T; Patterson, Cam; Wilson, Elizabeth M

    2004-07-16

    The NH2-terminal sequence of steroid receptors is highly variable between different receptors and in the same receptor from different species. In this study, a primary sequence homology comparison identified a 14-amino acid NH2-terminal motif of the human androgen receptor (AR) that is common to AR from all species reported, including the lower vertebrates. The evolutionarily conserved motif is unique to AR, with the exception of a partial sequence in the glucocorticoid receptor of higher species. The presence of the conserved motif in AR and the glucocorticoid receptor and its absence in other steroid receptors suggests convergent evolution. The function of the AR NH2-terminal conserved motif was suggested from a yeast two-hybrid screen that identified the COOH terminus of the Hsp70-interacting protein (CHIP) as a binding partner. We found that CHIP functions as a negative regulator of AR transcriptional activity by promoting AR degradation. In support of this, two mutations in the AR NH2-terminal conserved motif previously identified in the transgenic adenocarcinoma of mouse prostate model reduced the interaction between CHIP and AR. Our results suggest that the AR NH2-terminal domain contains an evolutionarily conserved motif that functions to limit AR transcriptional activity. Moreover, we demonstrate that the combination of comparative sequence alignment and yeast two-hybrid screening using short conserved peptides as bait provides an effective strategy to probe the structure-function relationships of steroid receptor NH2-terminal domains and other intrinsically unstructured transcriptional regulatory proteins.

  16. gene structure, gene expression

    Indian Academy of Sciences (India)

    Primer 5.0 software. To adjust for RNA quality and diffe- rences in cDNA concentration, we amplified actin as an internal control with the following primers: PtActin-F (5′-TG. AAGGAGAAACTTGCGTAT-3′) and PtActin-R (5′-GCA. CAATGTTACCGTACAGAT-3′). These genes were ampli- fied from first-strand cDNA using ...

  17. 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.

  18. Cloning and differential expression of five heat shock protein genes associated with thermal stress and development in the polyphagous predatory mite Neoseiulus cucumeris (Acari: Phytoseiidae).

    Science.gov (United States)

    Chen, Wei; Li, Dunsong; Zhang, Min; Zhao, Yunlong; Wu, Wenjing; Zhang, Guren

    2015-09-01

    In order to explore the role of heat shock proteins (Hsps) during thermal stress and development in the predatory mite Neoseiulus cucumeris (Oudemans), we cloned and characterized five full-length Hsp genes. We investigated the expression levels of these genes by quantitative real-time PCR. The five genes characterized here were NcHsp90, NcHsp75, NcHsp70, NcHsp60, and NcHsp40. These Hsps showed high sequence conservation and had greatest identity with heat shock proteins of Metaseiulus occidentalis and other mite and insect species. All five NcHsp genes showed changes in their levels of expression during development. Higher levels of expression were observed in adult females than in adult males, but there were no significant changes between pre-oviposition and post-oviposition stages in the females. NcHsp90, NcHsp75, and NcHsp70 expression levels were up-regulated after a heat shock, and the increases in NcHsp75 and NcHsp70 expression levels were maintained for at least 3 h. Up-regulation of NcHsp60 and NcHsp40 was not detected after 1 h at a high temperature (35-45 °C); however, a significant down-regulation was observed after 3 h heat exposure at 35 °C and 3 h recovery at 25 °C. Cold shock treatment (-5 to 15 °C) for 1 h did not acute elicit changes in the expression levels of any of the genes. At 5 °C, the expression levels of NcHsp90 significantly increased after 6 or 24 h exposure compared to the levels after 1 h exposure. Thus, expression of Hsp genes in N. cucumeris reflected developmental changes, sexual difference, and variable induced response to thermal stress. Increased expression of Hsps might protect N. cucumeris individuals under extreme temperature conditions. Therefore, it may be possible to enhance the thermal tolerance of commercially available N. cucumeris using temperature acclimation. Treatment at 35 °C should be suitable for such acclimation.

  19. Multiple controls affect arsenite oxidase gene expression in Herminiimonas arsenicoxydans

    Directory of Open Access Journals (Sweden)

    Coppée Jean-Yves

    2010-02-01

    Full Text Available Abstract Background Both the speciation and toxicity of arsenic are affected by bacterial transformations, i.e. oxidation, reduction or methylation. These transformations have a major impact on environmental contamination and more particularly on arsenic contamination of drinking water. Herminiimonas arsenicoxydans has been isolated from an arsenic- contaminated environment and has developed various mechanisms for coping with arsenic, including the oxidation of As(III to As(V as a detoxification mechanism. Results In the present study, a differential transcriptome analysis was used to identify genes, including arsenite oxidase encoding genes, involved in the response of H. arsenicoxydans to As(III. To get insight into the molecular mechanisms of this enzyme activity, a Tn5 transposon mutagenesis was performed. Transposon insertions resulting in a lack of arsenite oxidase activity disrupted aoxR and aoxS genes, showing that the aox operon transcription is regulated by the AoxRS two-component system. Remarkably, transposon insertions were also identified in rpoN coding for the alternative N sigma factor (σ54 of RNA polymerase and in dnaJ coding for the Hsp70 co-chaperone. Western blotting with anti-AoxB antibodies and quantitative RT-PCR experiments allowed us to demonstrate that the rpoN and dnaJ gene products are involved in the control of arsenite oxidase gene expression. Finally, the transcriptional start site of the aoxAB operon was determined using rapid amplification of cDNA ends (RACE and a putative -12/-24 σ54-dependent promoter motif was identified upstream of aoxAB coding sequences. Conclusion These results reveal the existence of novel molecular regulatory processes governing arsenite oxidase expression in H. arsenicoxydans. These data are summarized in a model that functionally integrates arsenite oxidation in the adaptive response to As(III in this microorganism.

  20. Hepatobiliary transporter expression and post-operative jaundice in patients undergoing partial hepatectomy.

    Science.gov (United States)

    Bernhardt, Gerwin A; Zollner, Gernot; Cerwenka, Herwig; Kornprat, Peter; Fickert, Peter; Bacher, Heinz; Werkgartner, Georg; Müller, Gabriele; Zatloukal, Kurt; Mischinger, Hans-Jörg; Trauner, Michael

    2012-01-01

    Post-operative hyperbilirubinaemia in patients undergoing liver resections is associated with high morbidity and mortality. Apart from different known factors responsible for the development of post-operative jaundice, little is known about the role of hepatobiliary transport systems in the pathogenesis of post-operative jaundice in humans after liver resection. Two liver tissue samples were taken from 14 patients undergoing liver resection before and after Pringle manoeuvre. Patients were retrospectively divided into two groups according to post-operative bilirubin serum levels. The two groups were analysed comparing the results of hepatobiliary transporter [Na-taurocholate cotransporter (NTCP); multidrug resistance gene/phospholipid export pump(MDR3); bile salt export pump (BSEP); canalicular bile salt export pump (MRP2)], heat shock protein 70 (HSP70) expression as well as the results of routinely taken post-operative liver chemistry tests. Patients with low post-operative bilirubin had lower levels of NTCP, MDR3 and BSEP mRNA compared to those with high bilirubin after Pringle manoeuvre. HSP70 levels were significantly higher after ischaemia-reperfusion (IR) injury in both groups resulting in 4.5-fold median increase. Baseline median mRNA expression of all four transporters prior to Pringle manoeuvre tended to be lower in the low bilirubin group whereas expression of HSP70 was higher in the low bilirubin group compared to the high bilirubin group. Higher mRNA levels of HSP70 in the low bilirubin group could indicate a possible protective effect of high HSP70 levels against IR injury. Although the exact role of hepatobiliary transport systems in the development of post-operative hyper bilirubinemia is not yet completely understood, this study provides new insights into the molecular aspects of post-operative jaundice after liver surgery. © 2011 John Wiley & Sons A/S.

  1. Gastroprotective activity of Annona muricata leaves against ethanol-induced gastric injury in rats via Hsp70/Bax involvement.

    Science.gov (United States)

    Moghadamtousi, Soheil Zorofchian; Rouhollahi, Elham; Karimian, Hamed; Fadaeinasab, Mehran; Abdulla, Mahmood Ameen; Kadir, Habsah Abdul

    2014-01-01

    The popular fruit tree of Annona muricata L. (Annonaceae), known as soursop and graviola, is a widely distributed plant in Central and South America and tropical countries. Leaves of A. muricata have been reported to possess antioxidant and anti-inflammatory activities. In this study, the gastroprotective effects of ethyl acetate extract of A. muricata leaves (EEAM) were investigated against ethanol-induced gastric injury models in rats. The acute toxicity test of EEAM in rats, carried out in two doses of 1 g/kg and 2 g/kg, showed the safety of this plant, even at the highest dose of 2 g/kg. The antiulcer study in rats (five groups, n=6) was performed with two doses of EEAM (200 mg/kg and 400 mg/kg) and with omeprazole (20 mg/kg), as a standard antiulcer drug. Gross and histological features showed the antiulcerogenic characterizations of EEAM. There was significant suppression on the ulcer lesion index of rats pretreated with EEAM, which was comparable to the omeprazole effect in the omeprazole control group. Oral administration of EEAM to rats caused a significant increase in the level of nitric oxide and antioxidant activities, including catalase, glutathione, and superoxide dismutase associated with attenuation in gastric acidity, and compensatory effect on the loss of gastric wall mucus. In addition, pretreatment of rats with EEAM caused significant reduction in the level of malondialdehyde, as a marker for oxidative stress, associated with an increase in prostaglandin E2 activity. Immunohistochemical staining also demonstrated that EEAM induced the downregulation of Bax and upregulation of Hsp70 proteins after pretreatment. Collectively, the present results suggest that EEAM has a promising antiulcer potential, which could be attributed to its suppressive effect against oxidative damage and preservative effect toward gastric wall mucus.

  2. Curcumin treatment enhances islet recovery by induction of heat shock response proteins, Hsp70 and heme oxygenase-1, during cryopreservation.

    Science.gov (United States)

    Kanitkar, Meghana; Bhonde, Ramesh R

    2008-01-16

    Limited recovery of islets post-cryopreservation influences graft survival and transplantation efficiency during diabetes treatment. As curcumin, a potent antioxidant/radical scavenging compound, protects islets against beta cell toxins, we hypothesized that inclusion of curcumin during cryopreservation or during post-thaw culture or both may rescue islets from cryoinjury. To test the effect of curcumin inclusion on islet recovery murine islets were isolated by the collagenase digestion, cultured for 48 h, cryopreserved using dimethylsulphoxide as cryoprotectant -- with or without curcumin (10 microM) -- and then slow cooled to -40 degrees C before immersing them in liquid nitrogen for 7 days. Following rapid thawing with sucrose gradient and 24 h post-thaw culture -- in presence or absence of curcumin (10 microM) -- islet viability and functionality were determined. Islet recovery in curcumin treated groups was significantly higher than in groups where islets were cryopreserved without curcumin. Islets cryopreserved with curcumin also showed more intact islets as well as better morphology as compared to islets cryopreserved without curcumin. Curcumin treated islets also showed significant inhibition of ROS generation as compared to islets cryopreserved without curcumin. Glucose responsiveness and insulin secretion in islets cryopreserved with curcumin was equal to that of the freshly isolated islets as against islets cryopreserved without curcumin. Elevated level of Hsp 70 and HO-1 were observed in islets cryopreserved with curcumin and may contribute to curcumin-induced islet rescue. Hence, we conclude that inclusion of curcumin into cryopreservation medium inhibits ROS generation and corresponding islet damage and dysfunction.

  3. Engagement of cellular prion protein with the co-chaperone Hsp70/90 organizing protein regulates the proliferation of glioblastoma stem-like cells.

    Science.gov (United States)

    Iglesia, Rebeca Piatniczka; Prado, Mariana Brandão; Cruz, Lilian; Martins, Vilma Regina; Santos, Tiago Góss; Lopes, Marilene Hohmuth

    2017-04-17

    Glioblastoma (GBM), a highly aggressive brain tumor, contains a subpopulation of glioblastoma stem-like cells (GSCs) that play roles in tumor maintenance, invasion, and therapeutic resistance. GSCs are therefore a promising target for GBM treatment. Our group identified the cellular prion protein (PrP C ) and its partner, the co-chaperone Hsp70/90 organizing protein (HOP), as potential target candidates due to their role in GBM tumorigenesis and in neural stem cell maintenance. GSCs expressing different levels of PrP C were cultured as neurospheres with growth factors, and characterized with stem cells markers and adhesion molecules markers through immunofluorescence and flow cytometry. We than evaluated GSC self-renewal and proliferation by clonal density assays and BrdU incorporation, respectively, in front of recombinant HOP treatment, combined or not with a HOP peptide which mimics the PrP C binding site. Stable silencing of HOP was also performed in parental and/or PrP C -depleted cell populations, and proliferation in vitro and tumor growth in vivo were evaluated. Migration assays were performed on laminin-1 pre-coated glass. We observed that, when GBM cells are cultured as neurospheres, they express specific stemness markers such as CD133, CD15, Oct4, and SOX2; PrP C is upregulated compared to monolayer culture and co-localizes with CD133. PrP C silencing downregulates the expression of molecules associated with cancer stem cells, upregulates markers of cell differentiation and affects GSC self-renewal, pointing to a pivotal role for PrP C in the maintenance of GSCs. Exogenous HOP treatment increases proliferation and self-renewal of GSCs in a PrP C -dependent manner while HOP knockdown disturbs the proliferation process. In vivo, PrP C and/or HOP knockdown potently inhibits the growth of subcutaneously implanted glioblastoma cells. In addition, disruption of the PrP C -HOP complex by a HOP peptide, which mimics the PrP C binding site, affects GSC self

  4. Embryo development, stress protein (Hsp70) responses, and histopathology in zebrafish (Danio rerio) following exposure to nickel chloride, chlorpyrifos, and binary mixtures of them.

    Science.gov (United States)

    Scheil, Volker; Zürn, Alexandra; Köhler, Heinz-R; Triebskorn, Rita

    2010-02-01

    Two different classes of chemicals were tested in a multilevel approach in this study: NiCl(2) as a representative for heavy metals and chlorpyrifos, a pesticide. Both, the single substances and mixtures of them were investigated for their effects on embryonic development, histological alterations, and the stress protein (Hsp70) response in the zebrafish Danio rerio. Fishes were exposed from fertilization of eggs up to a maximum of 168 h post fertilization, depending on the investigated endpoint. NiCl(2) led to effects in all tests which, however, were less severe at the histopathological level than in developmental (hatching success) and stress protein studies. Chlorpyrifos did not lead to developmental alterations but it was found to induce the Hsp70 response as well as histopathological damages. Mixtures of both substances resulted in similar results as the single substances; the results suggest an independent mode of action of these two substances and additivity of their effects.

  5. Heterologous minor coat proteins of Citrus tristeza virus strains affect encapsidation, but the coexpression of HSP70h and p61 restores encapsidation to wild-type levels.

    Science.gov (United States)

    Tatineni, Satyanarayana; Gowda, Siddarame; Dawson, William O

    2010-07-05

    The long flexuous bipolar virions of Citrus tristeza virus (CTV), a Closterovirus, are encapsidated with two capsid proteins at opposite ends: the minor coat protein (CPm) encapsidates the 5' 630 nts of the genomic RNA and the major coat protein encapsidates the remainder of the genome. In this study, we found encapsidation of CTV CPm in the absence of other assembly-related proteins is highly specific in contrast to most plant viruses that allow virion assembly by a range of heterologous coat proteins. Heterologous CPms with 95-96% amino acid identity from related strains in CTV-CPm, a replicon with CPm as the only assembly-related ORF, either failed to initiate encapsidation or reduced encapsidation substantially. Substitution of subsets of amino acids revealed that the amino acids that differ between positions 121 and 180 of the VT strain, and 61 and 120 of the T3 strain were involved in specific encapsidation. We further mapped the specific encapsidation to a single amino acid: mutation of methionine(165) to threonine (VT type) or serine(105) to proline (T3 type) in CTV-CPm failed to form nucleocapsids. However, the heterologous CPm in combination with both HSP70h and p61 proteins, but not HSP70h or p61 alone, encapsidated at wild-type levels, suggesting that specific encapsidation by CPm was mitigated by the combination of HSP70h and p61. Thus, in addition to the previously described functions of HSP70h and p61 of greatly enhanced virion formation and restriction of CPm encapsidation to the 5' 630 nts of the genomic RNA, these proteins facilitate encapsidation by heterologous CPms. Published by Elsevier Inc.

  6. Cortisol levels and expression of selected stress- and apoptosis-related genes in the embryos of Atlantic cod, Gadus morhua following short-term exposure to air

    DEFF Research Database (Denmark)

    Marlowe, Christopher; Caipang, A.; Fagutao, Ferdinand F.

    2015-01-01

    Embryos (morula stage) of Atlantic cod, Gadus morhua L., were collected and subjected to air exposure for 2 min. followed by recovery at ambient conditions in the rearing container. Total immunoreactive cortisol and transcription of selected stress- and apoptosis-related genes of the embryos were...... determined before the application of the stressor and at 0.5, 1 and 24 h post-exposure. There was no significant difference in the total cortisol levels of the fertilized eggs before and after handling stress. There was high expression level of hsp70 and sod before application of the stressor...... and significantly increased at 0.5 h postexposure. The expression levels of cat and gpx were weak to moderate and were not affected by the stressor. The apoptotic genes, mcl1 and NR-13 were highly expressed and significantly increased after exposure to air. Bcl-X1 and Bcl-X2 were moderately expressed in the control...

  7. Gene expression changes in response to aging compared to heat stress, oxidative stress and ionizing radiation in Drosophila melanogaster

    Science.gov (United States)

    Landis, Gary; Shen, Jie; Tower, John

    2012-01-01

    Gene expression changes in response to aging, heat stress, hyperoxia, hydrogen peroxide, and ionizing radiation were compared using microarrays. A set of 18 genes were up-regulated across all conditions, indicating a general stress response shared with aging, including the heat shock protein (Hsp) genes Hsp70, Hsp83 and l(2)efl, the glutathione-S-transferase gene GstD2, and the mitochondrial unfolded protein response (mUPR) gene ref(2)P. Selected gene expression changes were confirmed using quantitative PCR, Northern analysis and GstD-GFP reporter constructs. Certain genes were altered in only a subset of the conditions, for example, up-regulation of numerous developmental pathway and signaling genes in response to hydrogen peroxide. While aging shared features with each stress, aging was more similar to the stresses most associated with oxidative stress (hyperoxia, hydrogen peroxide, ionizing radiation) than to heat stress. Aging is associated with down-regulation of numerous mitochondrial genes, including electron-transport-chain (ETC) genes and mitochondrial metabolism genes, and a sub-set of these changes was also observed upon hydrogen peroxide stress and ionizing radiation stress. Aging shared the largest number of gene expression changes with hyperoxia. The extensive down-regulation of mitochondrial and ETC genes during aging is consistent with an aging-associated failure in mitochondrial maintenance, which may underlie the oxidative stress-like and proteotoxic stress-like responses observed during aging. PMID:23211361

  8. Gene expression changes in response to aging compared to heat stress, oxidative stress and ionizing radiation in Drosophila melanogaster.

    Science.gov (United States)

    Landis, Gary; Shen, Jie; Tower, John

    2012-11-01

    Gene expression changes in response to aging, heat stress, hyperoxia, hydrogen peroxide, and ionizing radiation were compared using microarrays. A set of 18 genes were up-regulated across all conditions, indicating a general stress response shared with aging, including the heat shock protein (Hsp) genes Hsp70, Hsp83 and l(2)efl, the glutathione-S-transferase gene GstD2, and the mitochondrial unfolded protein response (mUPR) gene ref(2)P. Selected gene expression changes were confirmed using quantitative PCR, Northern analysis and GstD-GFP reporter constructs. Certain genes were altered in only a subset of the conditions, for example, up-regulation of numerous developmental pathway and signaling genes in response to hydrogen peroxide. While aging shared features with each stress, aging was more similar to the stresses most associated with oxidative stress (hyperoxia, hydrogen peroxide, ionizing radiation) than to heat stress. Aging is associated with down-regulation of numerous mitochondrial genes, including electron-transport-chain (ETC) genes and mitochondrial metabolism genes, and a sub-set of these changes was also observed upon hydrogen peroxide stress and ionizing radiation stress. Aging shared the largest number of gene expression changes with hyperoxia. The extensive down-regulation of mitochondrial and ETC genes during aging is consistent with an aging-associated failure in mitochondrial maintenance, which may underlie the oxidative stress-like and proteotoxic stress-like responses observed during aging.

  9. Temperature effects on gene expression and morphological development of European eel, Anguilla anguilla larvae.

    Directory of Open Access Journals (Sweden)

    Sebastian N Politis

    Full Text Available Temperature is important for optimization of rearing conditions in aquaculture, especially during the critical early life history stages of fish. Here, we experimentally investigated the impact of temperature (16, 18, 20, 22 and 24°C on thermally induced phenotypic variability, from larval hatch to first-feeding, and the linked expression of targeted genes [heat shock proteins (hsp, growth hormone (gh and insulin-like growth factors (igf] associated to larval performance of European eel, Anguilla anguilla. Temperature effects on larval morphology and gene expression were investigated throughout early larval development (in real time from 0 to 18 days post hatch and at specific developmental stages (hatch, jaw/teeth formation, and first-feeding. Results showed that hatch success, yolk utilization efficiency, survival, deformities, yolk utilization, and growth rates were all significantly affected by temperature. In real time, increasing temperature from 16 to 22°C accelerated larval development, while larval gene expression patterns (hsp70, hsp90, gh and igf-1 were delayed at cold temperatures (16°C or accelerated at warm temperatures (20-22°C. All targeted genes (hsp70, hsp90, gh, igf-1, igf-2a, igf-2b were differentially expressed during larval development. Moreover, expression of gh was highest at 16°C during the jaw/teeth formation, and the first-feeding developmental stages, while expression of hsp90 was highest at 22°C, suggesting thermal stress. Furthermore, 24°C was shown to be deleterious (resulting in 100% mortality, while 16°C and 22°C (~50 and 90% deformities respectively represent the lower and upper thermal tolerance limits. In conclusion, the high survival, lowest incidence of deformities at hatch, high yolk utilization efficiency, high gh and low hsp expression, suggest 18°C as the optimal temperature for offspring of European eel. Furthermore, our results suggest that the still enigmatic early life history stages of

  10. Computational Analysis of Residue Interaction Networks and Coevolutionary Relationships in the Hsp70 Chaperones: A Community-Hopping Model of Allosteric Regulation and Communication.

    Directory of Open Access Journals (Sweden)

    Gabrielle Stetz

    2017-01-01

    Full Text Available Allosteric interactions in the Hsp70 proteins are linked with their regulatory mechanisms and cellular functions. Despite significant progress in structural and functional characterization of the Hsp70 proteins fundamental questions concerning modularity of the allosteric interaction networks and hierarchy of signaling pathways in the Hsp70 chaperones remained largely unexplored and poorly understood. In this work, we proposed an integrated computational strategy that combined atomistic and coarse-grained simulations with coevolutionary analysis and network modeling of the residue interactions. A novel aspect of this work is the incorporation of dynamic residue correlations and coevolutionary residue dependencies in the construction of allosteric interaction networks and signaling pathways. We found that functional sites involved in allosteric regulation of Hsp70 may be characterized by structural stability, proximity to global hinge centers and local structural environment that is enriched by highly coevolving flexible residues. These specific characteristics may be necessary for regulation of allosteric structural transitions and could distinguish regulatory sites from nonfunctional conserved residues. The observed confluence of dynamics correlations and coevolutionary residue couplings with global networking features may determine modular organization of allosteric interactions and dictate localization of key mediating sites. Community analysis of the residue interaction networks revealed that concerted rearrangements of local interacting modules at the inter-domain interface may be responsible for global structural changes and a population shift in the DnaK chaperone. The inter-domain communities in the Hsp70 structures harbor the majority of regulatory residues involved in allosteric signaling, suggesting that these sites could be integral to the network organization and coordination of structural changes. Using a network-based formalism of

  11. Computational Analysis of Residue Interaction Networks and Coevolutionary Relationships in the Hsp70 Chaperones: A Community-Hopping Model of Allosteric Regulation and Communication.

    Science.gov (United States)

    Stetz, Gabrielle; Verkhivker, Gennady M

    2017-01-01

    Allosteric interactions in the Hsp70 proteins are linked with their regulatory mechanisms and cellular functions. Despite significant progress in structural and functional characterization of the Hsp70 proteins fundamental questions concerning modularity of the allosteric interaction networks and hierarchy of signaling pathways in the Hsp70 chaperones remained largely unexplored and poorly understood. In this work, we proposed an integrated computational strategy that combined atomistic and coarse-grained simulations with coevolutionary analysis and network modeling of the residue interactions. A novel aspect of this work is the incorporation of dynamic residue correlations and coevolutionary residue dependencies in the construction of allosteric interaction networks and signaling pathways. We found that functional sites involved in allosteric regulation of Hsp70 may be characterized by structural stability, proximity to global hinge centers and local structural environment that is enriched by highly coevolving flexible residues. These specific characteristics may be necessary for regulation of allosteric structural transitions and could distinguish regulatory sites from nonfunctional conserved residues. The observed confluence of dynamics correlations and coevolutionary residue couplings with global networking features may determine modular organization of allosteric interactions and dictate localization of key mediating sites. Community analysis of the residue interaction networks revealed that concerted rearrangements of local interacting modules at the inter-domain interface may be responsible for global structural changes and a population shift in the DnaK chaperone. The inter-domain communities in the Hsp70 structures harbor the majority of regulatory residues involved in allosteric signaling, suggesting that these sites could be integral to the network organization and coordination of structural changes. Using a network-based formalism of allostery, we

  12. Resistência à Síndrome Ascítica, Competência Homeotérmica e Níveis de Hsp70 no Coração e Pulmão de Frangos de Corte Resistance to Ascites Syndrome, Homoeothermic Competence and Levels of Hsp70 in the Heart and Lung of Broilers

    Directory of Open Access Journals (Sweden)

    Renata Hernandes

    2002-06-01

    Full Text Available Como em outros seres vivos, também nas células das aves ocorre a síntese das proteínas de baixo peso molecular (Hsp, cujo aumento é induzido sob condições de estresse. As Hsps têm um papel importante na manutenção da integridade celular, questiona-se o seu envolvimento no mecanismo de proteção celular de órgãos alvos na ocorrência da síndrome ascítica (SA. Este trabalho objetivou avaliar a temperatura corporal e os níveis da Hsp70 no coração e pulmão de frangos de corte Hubbard (sensível à SA e caipira de pescoço-pelado (resistente, criados em termoneutralidade (25°C e frio (16°C entre 10 e 45 dias de idade. Foram utilizados 192 pintos machos, 96 de cada linhagem. Não houve mortalidade por SA nas aves caipiras. Nas aves Hubbard, a mortalidade devida à SA foi de 4% e 41% em ambiente termoneutro e frio, respectivamente. Em ambiente frio, a temperatura corporal das aves Hubbard foi menor que a das caipiras. A temperatura corporal e o nível de Hsp70 do coração das aves Hubbard diminuíram com o aumento da idade, mas não nas aves caipiras, os quais se mantiveram constantes, inclusive a Hsp70 do pulmão. Independente da idade ou da temperatura, o nível de Hsp70 no pulmão das aves caipiras era superior ao das aves Hubbard. Em relação às aves Hubbard, as caipiras são homeotérmicas mais competentes e apresentam uma maior indução de Hsp70 nos órgãos primariamente afetados na SA, mas este não parece ser o sistema de proteção contra SA, a qual as aves de pescoço pelado são resistentes.Similar to other living animals, the cells of the birds also synthesize small proteins (heat shock protein, Hsp, which increasing levels can be induce by stress. The Hsp have a relevant function in maintaining the integrity of the cell, and we question if they are involved in the mechanism of the cellular protection of target organs affected by ascites syndrome (AS. The objective of this study was to evaluate the body temperature

  13. Changes in gene expression by 193- and 248-nm excimer laser radiation in cultured human fibroblasts.

    Science.gov (United States)

    Rimoldi, D; Flessate, D M; Samid, D

    1992-09-01

    Tissue ablation by ultraviolet excimer lasers results in exposure of viable cells to subablative doses of radiation. To understand the potential biological consequences better, we have studied changes in gene expression in cultured human skin fibroblasts exposed to either 193- or 248-nm laser light. Northern blot analyses revealed that both treatments up-regulate a common set of genes, including interstitial collagenase, tissue inhibitor of metalloprotease, metallothionein, and the proto-oncogene c-fos. Dose-response and kinetic studies of collagenase induction by 193-nm radiation showed a maximal effect with 60 J/m2 and at approximately 24 h. The induction was still persistent 96 h later. In addition to the commonly affected genes, known to be activated also by conventional UV light (254 nm) and tumor-promoting phorbol esters, other genes were found to be selectively induced by the 193-nm radiation. The heat-shock hsp70 mRNA, undetectable in controls and in cultures irradiated at 248 nm, was transiently induced 8 h after exposure to 193-nm radiation. Furthermore, a selective up-regulation of collagen type I expression was observed. The results indicate that the 193- and 248-nm radiations by excimer lasers elicit specific and different cellular responses, in addition to an overlapping pathway of gene activation common also to UV radiation by germicidal lamps. The laser-induced genes could serve as molecular markers in evaluating cell injury in situ.

  14. Expression of genes related to antioxidant activity in Nile tilapia kept under salinity stress and fed diets containing different levels of vitamin C.

    Science.gov (United States)

    Caxico Vieira, Caio Alexandre Santos; Vieira, Jodnes Sobreira; Bastos, Marisa Silva; Zancanela, Vittor; Barbosa, Leandro Teixeira; Gasparino, Eliane; Del Vesco, Ana Paula

    2018-01-01

    The aim of this study was to examine whether (1) severe changes in salinity produced increased stress, and (2) vitamin C supplementation might reduce the observed damage in Nile tilapia. The parameters measured included condition factor, survival rate, and gene expression of catalase (CAT), heat shock protein 70 (HSP70), glutathione reductase (GSR), glutathione synthase (GSS), and glutathione peroxidase (GPx). The investigation was conducted with 160 Nile tilapia divided into four treatment groups: freshwater; 7 or 21 parts per thousand (‰) salinity, all fed a basal diet; as well as a fourth treatment group consisting of fish kept at 21‰ salinity fed a diet supplemented with vitamin C (1500 mg/kg). For gene expression analysis, liver samples were collected after 24 h or after 14 d. After 24 h, fish raised in 21‰ salinity and fed with the diet supplemented with vitamin C showed similar GPx expression as the control freshwater group. GSS expression in 21‰ salinity was similar to fish exposed to 7‰ salinity. Nile tilapia exposed to 21‰ salinity without vitamin C supplementation exhibited the highest HSP70 gene expression levels after 24 h. After 14-dtreatment, the lowest survival rate was observed in the 21‰ salinity group. After 14 d, the highest expression of GPx and GSR levels was detected in fish in the 21‰ salinity group that received vitamin C. Data indicate that vitamin C supplementation enhanced the expression of genes related to antioxidant capacity in Nile tilapia exposed to higher salinity, thereby increasing protection against the oxidative effects induced by high water salinity..

  15. Effects of predator exposure on Hsp70 expression and survival in tadpoles of the Common Frog (Rana temporaria)

    National Research Council Canada - National Science Library

    Sorensen, Jesper Givskov; Loeschcke, Volker; Merila, Juha; Laurila, Anssi

    2011-01-01

    ...)—a commonly found response to stress—in tadpoles of the Common Frog ( Rana temporaria L., 1758). In another experiment, we tested the survival of tadpoles in the presence of a free-ranging predator...

  16. Molecular Karyotype and Chromosomal Localization of Genes Encoding -tubulin, Cysteine Proteinase, hsp 70 and Actin in Trypanosoma rangeli

    National Research Council Canada - National Science Library

    CB Toaldo; M Steindel; MA Sousa; CC Tavares

    2001-01-01

    The molecular karyotype of nine Trypanosoma rangeli strains was analyzed by contour-clamped homogeneous electric field electrophoresis, followed by the chromosomal localization of -tubulin, cysteine...

  17. Thermotolerance and Heat-Shock Protein Gene Expression Patterns in Bemisia tabaci (Hemiptera: Aleyrodidae) Mediterranean in Relation to Developmental Stage.

    Science.gov (United States)

    Jiang, Rui; Qi, Lan-Da; Du, Yu-Zhou; Li, Yuan-Xi

    2017-10-01

    Temperature plays an important role in the growth, development, and geographic distribution of insects. There is convincing evidence that heat-shock proteins (HSPs) play important roles in helping organisms adapt to thermal stress. To better understand the physiological and ecological influence of thermal stress on the different development stages of Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) Mediterranean species (MED), nymphs and adults were shocked with temperatures of 35, 38, and 41℃ for 1 and 2 h, respectively, and the survival rate, fecundity, and developmental duration were investigated in the laboratory. The expression levels of the hsp40, hsp70, and hsp90 genes were assessed using real-time PCR. The results indicate that the survival rates of the nymphs and adults decreased with increased temperature. A 2-h heat shock at 41℃ induced a significant reduction in fecundity in adults and an increase in developmental duration in young nymphs. Hsp90 showed higher temperature responses to thermal stress than hsp40 or hsp70. The expression levels of the hsps in the adults were significantly down-regulated by a 2-h heat shock at 41℃ compared with that by a 1-h treatment. A significant decrease in the expression levels of the hsps also occurred in the adults when the temperature increased from 38 to 41℃ for the 2-h treatment, whereas no significant decrease occurred in the nymphs. Compared with previous studies, we provide some evidence indicating that MED has the potential to adapt to a wider temperature range than the Middle East-Asia Minor 1 species. © The Author 2017. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  18. Gene Expression Omnibus (GEO)

    Data.gov (United States)

    U.S. Department of Health & Human Services — Gene Expression Omnibus is a public functional genomics data repository supporting MIAME-compliant submissions of array- and sequence-based data. Tools are provided...

  19. Decipher the mechanisms of protein conformational changes induced by nucleotide binding through free-energy landscape analysis: ATP binding to Hsp70.

    Directory of Open Access Journals (Sweden)

    Adrien Nicolaï

    Full Text Available ATP regulates the function of many proteins in the cell by transducing its binding and hydrolysis energies into protein conformational changes by mechanisms which are challenging to identify at the atomic scale. Based on molecular dynamics (MD simulations, a method is proposed to analyze the structural changes induced by ATP binding to a protein by computing the effective free-energy landscape (FEL of a subset of its coordinates along its amino-acid sequence. The method is applied to characterize the mechanism by which the binding of ATP to the nucleotide-binding domain (NBD of Hsp70 propagates a signal to its substrate-binding domain (SBD. Unbiased MD simulations were performed for Hsp70-DnaK chaperone in nucleotide-free, ADP-bound and ATP-bound states. The simulations revealed that the SBD does not interact with the NBD for DnaK in its nucleotide-free and ADP-bound states whereas the docking of the SBD was found in the ATP-bound state. The docked state induced by ATP binding found in MD is an intermediate state between the initial nucleotide-free and final ATP-bound states of Hsp70. The analysis of the FEL projected along the amino-acid sequence permitted to identify a subset of 27 protein internal coordinates corresponding to a network of 91 key residues involved in the conformational change induced by ATP binding. Among the 91 residues, 26 are identified for the first time, whereas the others were shown relevant for the allosteric communication of Hsp70 s in several experiments and bioinformatics analysis. The FEL analysis revealed also the origin of the ATP-induced structural modifications of the SBD recently measured by Electron Paramagnetic Resonance. The pathway between the nucleotide-free and the intermediate state of DnaK was extracted by applying principal component analysis to the subset of internal coordinates describing the transition. The methodology proposed is general and could be applied to analyze allosteric communication in

  20. Use of Heat Stress Responsive Gene Expression Levels for Early Selection of Heat Tolerant Cabbage (Brassica oleracea L.)

    Science.gov (United States)

    Park, Hyun Ji; Jung, Won Yong; Lee, Sang Sook; Song, Jun Ho; Kwon, Suk-Yoon; Kim, HyeRan; Kim, ChulWook; Ahn, Jun Cheul; Cho, Hye Sun

    2013-01-01

    Cabbage is a relatively robust vegetable at low temperatures. However, at high temperatures, cabbage has disadvantages, such as reduced disease tolerance and lower yields. Thus, selection of heat-tolerant cabbage is an important goal in cabbage breeding. Easier or faster selection of superior varieties of cabbage, which are tolerant to heat and disease and have improved taste and quality, can be achieved with molecular and biological methods. We compared heat-responsive gene expression between a heat-tolerant cabbage line (HTCL), “HO”, and a heat-sensitive cabbage line (HSCL), “JK”, by Genechip assay. Expression levels of specific heat stress-related genes were increased in response to high-temperature stress, according to Genechip assays. We performed quantitative RT-PCR (qRT-PCR) to compare expression levels of these heat stress-related genes in four HTCLs and four HSCLs. Transcript levels for heat shock protein BoHsp70 and transcription factor BoGRAS (SCL13) were more strongly expressed only in all HTCLs compared to all HSCLs, showing much lower level expressions at the young plant stage under heat stress (HS). Thus, we suggest that expression levels of these genes may be early selection markers for HTCLs in cabbage breeding. In addition, several genes that are involved in the secondary metabolite pathway were differentially regulated in HTCL and HSCL exposed to heat stress. PMID:23736694

  1. Use of Heat Stress Responsive Gene Expression Levels for Early Selection of Heat Tolerant Cabbage (Brassica oleracea L.

    Directory of Open Access Journals (Sweden)

    Jun Cheul Ahn

    2013-06-01

    Full Text Available Cabbage is a relatively robust vegetable at low temperatures. However, at high temperatures, cabbage has disadvantages, such as reduced disease tolerance and lower yields. Thus, selection of heat-tolerant cabbage is an important goal in cabbage breeding. Easier or faster selection of superior varieties of cabbage, which are tolerant to heat and disease and have improved taste and quality, can be achieved with molecular and biological methods. We compared heat-responsive gene expression between a heat-tolerant cabbage line (HTCL, “HO”, and a heat-sensitive cabbage line (HSCL, “JK”, by Genechip assay. Expression levels of specific heat stress-related genes were increased in response to high-temperature stress, according to Genechip assays. We performed quantitative RT-PCR (qRT-PCR to compare expression levels of these heat stress-related genes in four HTCLs and four HSCLs. Transcript levels for heat shock protein BoHsp70 and transcription factor BoGRAS (SCL13 were more strongly expressed only in all HTCLs compared to all HSCLs, showing much lower level expressions at the young plant stage under heat stress (HS. Thus, we suggest that expression levels of these genes may be early selection markers for HTCLs in cabbage breeding. In addition, several genes that are involved in the secondary metabolite pathway were differentially regulated in HTCL and HSCL exposed to heat stress.

  2. Constitutive or Inducible Protective Mechanisms against UV-B Radiation in the Brown Alga Fucus vesiculosus? A Study of Gene Expression and Phlorotannin Content Responses.

    Science.gov (United States)

    Creis, Emeline; Delage, Ludovic; Charton, Sophie; Goulitquer, Sophie; Leblanc, Catherine; Potin, Philippe; Ar Gall, Erwan

    2015-01-01

    A role as UV sunscreens has been suggested for phlorotannins, the phenolic compounds that accumulate in brown algae in response to a number of external stimuli and take part in cell wall structure. After exposure of the intertidal brown alga Fucus vesiculosus to artificial UV-B radiation, we examined its physiological responses by following the transcript level of the pksIII gene encoding a phloroglucinol synthase, likely to be involved in the first step of phlorotannins biosynthesis. We also monitored the expression of three targeted genes, encoding a heat shock protein (hsp70), which is involved in global stress responses, an aryl sulfotransferase (ast), which could be involved in the sulfation of phlorotannins, and a vanadium bromoperoxidase (vbpo), which can potentially participate in the scavenging of Reactive Oxygen Species (ROS) and in the cross-linking and condensation of phlorotannins. We investigated whether transcriptional regulation of these genes is correlated with an induction of phlorotannin accumulation by establishing metabolite profiling of purified fractions of low molecular weight phlorotannins. Our findings demonstrated that a high dose of UV-B radiation induced a significant overexpression of hsp70 after 12 and 24 hours following the exposure to the UV-B treatment, compared to control treatment. The physiological performance of algae quantified by the photosynthetic efficiency (Fv/Fm) was slightly reduced. However UV-B treatment did not induce the accumulation of soluble phlorotannins in F. vesiculosus during the kinetics of four weeks, a result that may be related to the lack of induction of the pksIII gene expression. Taken together these results suggest a constitutive accumulation of phlorotannins occurring during the development of F.vesiculosus, rather than inducible processes. Gene expression studies and phlorotannin profiling provide here complementary approaches to global quantifications currently used in studies of phenolic compounds

  3. Constitutive or Inducible Protective Mechanisms against UV-B Radiation in the Brown Alga Fucus vesiculosus? A Study of Gene Expression and Phlorotannin Content Responses.

    Directory of Open Access Journals (Sweden)

    Emeline Creis

    Full Text Available A role as UV sunscreens has been suggested for phlorotannins, the phenolic compounds that accumulate in brown algae in response to a number of external stimuli and take part in cell wall structure. After exposure of the intertidal brown alga Fucus vesiculosus to artificial UV-B radiation, we examined its physiological responses by following the transcript level of the pksIII gene encoding a phloroglucinol synthase, likely to be involved in the first step of phlorotannins biosynthesis. We also monitored the expression of three targeted genes, encoding a heat shock protein (hsp70, which is involved in global stress responses, an aryl sulfotransferase (ast, which could be involved in the sulfation of phlorotannins, and a vanadium bromoperoxidase (vbpo, which can potentially participate in the scavenging of Reactive Oxygen Species (ROS and in the cross-linking and condensation of phlorotannins. We investigated whether transcriptional regulation of these genes is correlated with an induction of phlorotannin accumulation by establishing metabolite profiling of purified fractions of low molecular weight phlorotannins. Our findings demonstrated that a high dose of UV-B radiation induced a significant overexpression of hsp70 after 12 and 24 hours following the exposure to the UV-B treatment, compared to control treatment. The physiological performance of algae quantified by the photosynthetic efficiency (Fv/Fm was slightly reduced. However UV-B treatment did not induce the accumulation of soluble phlorotannins in F. vesiculosus during the kinetics of four weeks, a result that may be related to the lack of induction of the pksIII gene expression. Taken together these results suggest a constitutive accumulation of phlorotannins occurring during the development of F.vesiculosus, rather than inducible processes. Gene expression studies and phlorotannin profiling provide here complementary approaches to global quantifications currently used in studies of

  4. Standardized tape stripping prior to patch testing induces upregulation of Hsp90, Hsp70, IL-33, TNF-α and IL-8/CXCL8 mRNA: new insights into the involvement of 'alarmins'.

    Science.gov (United States)

    Dickel, Heinrich; Gambichler, Thilo; Kamphowe, Jeanette; Altmeyer, Peter; Skrygan, Marina

    2010-10-01

    The strip patch test is recommended if patch test results are presumed to be false-negative. It is under discussion whether the cutaneous inflammation induced by tape stripping is a pivotal element in enhancing the allergic contact dermatitis response. To investigate epidermal mRNA expression of mediators implicated in the regulation of inflammation induced by tape stripping according to our standardized protocol in human skin. Thirty-one healthy volunteers (median age 26 years, range 18-62 years) participated in the study. We measured epidermal mRNAs coding for cytokines [tumour necrosis factor-α (TNF-α), interferon-γ (IFN-γ), interleukin (IL)-1α, IL-1β, IL-33, IL-10, IL-8/CXCL8, chemokine (C-C motif) ligand 2 (CCL2)/MCP-1 (monocyte chemotactic protein-1), chemokine (C-C motif) ligand 5 (CCL5)/RANTES (regulated upon activation normal T-cell expressed, and presumably secreted), IL-6, granulocyte-macrophage colony-stimulating factor (GM-CSF)], an adhesion molecule [intercellular adhesion molecule type 1 (ICAM-1)], heat shock proteins (Hsp27, Hsp70 and Hsp90), and the inflammasome complex [NACHT-LRR-PYD-containing protein-3 (NALP3), apoptosis-associated speck-like protein containing a CARD (ASC)/CARD5 (caspase recruitment domain-containing protein-5) and caspase-1] in tape-stripped skin of the upper back relative to untreated skin of the upper back, using quantitative real-time reverse transcription polymerase chain reaction technology. Epidermal mRNA expression levels of TNF-α, Hsp90, Hsp70, IL-33, and IL-8/CXCL8 were significantly upregulated, whereas CCL5/RANTES expression was significantly downregulated 6 hr after standardized tape stripping. Tape stripping the skin according to our standardized protocol shows immunostimulatory effects, with induction of keratinocyte-derived 'alarmins' and cytokines mounting an immune response, which may contribute to the increased sensitivity of the strip patch test versus the patch test. © 2010 John Wiley & Sons A/S.

  5. Variegated expression of Hsp22 transgenic reporters indicates cell-specific patterns of aging in Drosophila oenocytes.

    Science.gov (United States)

    Tower, John; Landis, Gary; Gao, Rebecca; Luan, Albert; Lee, Jonathan; Sun, Yuanyue

    2014-03-01

    The cytoplasmic chaperone gene Hsp70 and the mitochondrial chaperone gene Hsp22 are upregulated during normal aging in Drosophila in tissue-general patterns. In addition, Hsp22 reporters are dramatically upregulated during aging in a subset of the oenocytes (liver-like cells). Hsp22 reporter expression varied dramatically between individual oenocytes and between groups of oenocytes located in adjacent body segments, and was negatively correlated with accumulation of age pigment, indicating cell-specific and cell-lineage-specific patterns of oenocyte aging. Conditional transgenic systems were used to express 88 transgenes to search for trans-regulators of the Hsp70 and Hsp22 reporters during aging. The wingless gene increased tissue-general upregulation of both Hsp70 and Hsp22 reporters. In contrast, the mitochondrial genes MnSOD and Hsp22 increased expression of Hsp22 reporters in the oenocytes and decreased accumulation of age pigment in these cells. The data suggest that cell-specific and cell lineage-specific patterns of mitochondrial malfunction contribute to oenocyte aging.

  6. Th1 stimulatory proteins of Leishmania donovani: comparative cellular and protective responses of rTriose phosphate isomerase, rProtein disulfide isomerase and rElongation factor-2 in combination with rHSP70 against visceral leishmaniasis.

    Science.gov (United States)

    Jaiswal, Anil Kumar; Khare, Prashant; Joshi, Sumit; Kushawaha, Pramod Kumar; Sundar, Shyam; Dube, Anuradha

    2014-01-01

    In visceral leishmaniasis, the recovery from the disease is always associated with the generation of Th1-type of cellular responses. Based on this, we have previously identified several Th1-stimulatory proteins of Leishmania donovani -triose phosphate isomerase (TPI), protein disulfide isomerase (PDI) and elongation factor-2 (EL-2) etc. including heat shock protein 70 (HSP70) which induced Th1-type of cellular responses in both cured Leishmania patients/hamsters. Since, HSPs, being the logical targets for vaccines aimed at augmenting cellular immunity and can be early targets in the immune response against intracellular pathogens; they could be exploited as vaccine/adjuvant to induce long-term immunity more effectively. Therefore, in this study, we checked whether HSP70 can further enhance the immunogenicity and protective responses of the above said Th1-stimulatory proteins. Since, in most of the studies, immunogenicity of HSP70 of L. donovani was assessed in native condition, herein we generated recombinant HSP70 and tested its potential to stimulate immune responses in lymphocytes of cured Leishmania infected hamsters as well as in the peripheral blood mononuclear cells (PBMCs) of cured patients of VL either individually or in combination with above mentioned recombinant proteins. rLdHSP70 alone elicited strong cellular responses along with remarkable up-regulation of IFN-γ and IL-12 cytokines and extremely lower level of IL-4 and IL-10. Among the various combinations, rLdHSP70 + rLdPDI emerged as superior one augmenting improved cellular responses followed by rLdHSP70 + rLdEL-2. These combinations were further evaluated for its protective potential wherein rLdHSP70 + rLdPDI again conferred utmost protection (∼80%) followed by rLdHSP70 + rLdEL-2 (∼75%) and generated a strong cellular immune response with significant increase in the levels of iNOS transcript as well as IFN-γ and IL-12 cytokines which was further supported by the high level of IgG2 antibody

  7. Expression responses of five cold tolerant related genes to two temperature dropping treatments in sea cucumber Apostichopus japonicus

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    Li, Chengze; Chang, Yaqing; Pang, Zhenguo; Ding, Jun; Ji, Nanjing

    2015-03-01

    Environmental conditions, including ambient temperature, play important roles in survival, growth development, and reproduction of the Japanese sea cucumber, Apostichopus japonicus. Low temperatures result in slowed growth and skin ulceration disease. In a previous study, we investigated the effect of low temperature on gene expression profiles in A. japonicus by suppression subtractive hybridization (SSH). Genes encoding Ferritin, Lysozyme, Hsp70, gp96, and AjToll were selected from a subtracted cDNA library of A. japonicus under acute cold stress. The transcriptional expression profiles of these genes were investigated in different tissues (coelomocyte, respiratory tree, intestine, longitudinal muscle) after exposure to acute and mild temperature dropping treatments. The results show that (1) the five cold-tolerance-related genes were found in all four tissues and the highest mRNA levels were observed in coelomocyte and respiratory tree; (2) under the temperature dropping treatments, three types of transcriptional regulation patterns were observed: primary suppression followed by up-regulation at -2°C, suppressed expression throughout the two treatments, and more rarely an initial stimulation followed by suppression; and (3) gene expression suppression was more severe under acute temperature dropping than under mild temperature dropping treatment. The five cold-tolerance-related genes that were distributed mainly in coelomocyte and respiratory tissues were generally down-regulated by low temperature stress but an inverse up-regulation event was found at the extreme temperature (-2°C).

  8. Characterization and gene expression profiles of thermotolerant Saccharomyces cerevisiae isolates from Thai fruits.

    Science.gov (United States)

    Auesukaree, Choowong; Koedrith, Preeyaporn; Saenpayavai, Pornpon; Asvarak, Thipa; Benjaphokee, Suthee; Sugiyama, Minetaka; Kaneko, Yoshinobu; Harashima, Satoshi; Boonchird, Chuenchit

    2012-08-01

    For industrial applications, fermentation of ethanol at high temperature offers advantages such as reduction in cooling costs, reduced risk of microbial contamination and higher efficiency of fermentation processes including saccharification and continuous ethanol stripping. Three thermotolerant Saccharomyces cerevisiae isolates (C3723, C3751 and C3867) from Thai fruits were capable of growing and producing 38 g/L ethanol up to 41°C. Based on genetic analyses, these isolates were prototrophic and homothallic, with dominant homothallic and thermotolerant phenotypes. After short-term (30 min) and long-term (12 h) exposure at 37°C, expression levels increased for the heat stress-response genes HSP26, SSA4, HSP82, and HSP104 encoding the heat shock proteins small HSP, HSP70, HSP90 and the HSP100 family, respectively. In isolates C3723 and C3867, expression was significantly higher than that in reference isolates W303 and TISTR5606 for TPS1 encoding trehalose-6-phosphate synthase, NTH1 encoding neutral trehalase and GSY1 encoding glycogen synthase. The results suggested that continuous high expression of heat stress-response genes was important for the long-term, heat stress tolerance of these thermotolerant isolates. Copyright © 2012 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  9. Immunogenicity and efficacy of single antigen Gp63, polytope and polytopeHSP70 DNA vaccines against visceral Leishmaniasis in experimental mouse model.

    Directory of Open Access Journals (Sweden)

    Rakhee Sachdeva

    Full Text Available Polytope approach of genetic immunization is a promising strategy for the prevention of infectious disease as it is capable of generating effective cell mediated immunity by delivering the T cell epitopes assembled in series. Leishmaniasis is a significant world wide health problem for which no vaccine exists. In this study we have compared immunogenicity and efficacy of three types of DNA vaccines: single antigen Gp63 (Gp63/pcDNA, polytope (Poly/pcDNA and Polytope fused with hsp70 (Poly/hsp/pcDNA against visceral leishmaniasis in susceptible BALB/c mice. Mice vaccinated with these plasmids generated strong Th1 immune response as seen by dominating IFN-gamma over IL-10 cytokine. Interestingly, cytotoxic responses generated by polytope DNA plasmid fused with hsp70 of Leishmania donovani were significantly higher when compared to polytope and single antigen Gp63 vaccine. Challenge studies revealed that the parasite load in liver and spleen was significantly lower with Poly/hsp/pcDNA vaccination compared to other vaccines. Therefore, our study indicates that polytope DNA vaccine is a feasible, practical and effective approach for visceral leishmaniasis.

  10. Immunogenicity and efficacy of single antigen Gp63, polytope and polytopeHSP70 DNA vaccines against visceral Leishmaniasis in experimental mouse model.

    Science.gov (United States)

    Sachdeva, Rakhee; Banerjea, Akhil C; Malla, Nancy; Dubey, Mohan Lal

    2009-12-02

    Polytope approach of genetic immunization is a promising strategy for the prevention of infectious disease as it is capable of generating effective cell mediated immunity by delivering the T cell epitopes assembled in series. Leishmaniasis is a significant world wide health problem for which no vaccine exists. In this study we have compared immunogenicity and efficacy of three types of DNA vaccines: single antigen Gp63 (Gp63/pcDNA), polytope (Poly/pcDNA) and Polytope fused with hsp70 (Poly/hsp/pcDNA) against visceral leishmaniasis in susceptible BALB/c mice. Mice vaccinated with these plasmids generated strong Th1 immune response as seen by dominating IFN-gamma over IL-10 cytokine. Interestingly, cytotoxic responses generated by polytope DNA plasmid fused with hsp70 of Leishmania donovani were significantly higher when compared to polytope and single antigen Gp63 vaccine. Challenge studies revealed that the parasite load in liver and spleen was significantly lower with Poly/hsp/pcDNA vaccination compared to other vaccines. Therefore, our study indicates that polytope DNA vaccine is a feasible, practical and effective approach for visceral leishmaniasis.

  11. Heat shock proteins HSP70 and HSP27 in the cerebral spinal fluid of patients undergoing thoracic aneurysm repair correlate with the probability of postoperative paralysis.

    Science.gov (United States)

    Hecker, James G; Sundram, Hari; Zou, Shaomin; Praestgaard, Amy; Bavaria, Joseph E; Ramchandren, Sindhu; McGarvey, Michael

    2008-12-01

    An understanding of the time course and correlation with injury of heat shock proteins (HSPs) released during brain and/or spinal cord cellular stress (ischemia) is critical in understanding the role of the HSPs in cellular survival, and may provide a clinically useful biomarker of severe cellular stress. We have analyzed the levels of HSPs in the cerebrospinal fluid (CSF) from patients who are undergoing thoracic aneurysm repair. Blood and CSF samples were collected at regular intervals, and CSF was analyzed by enzyme-linked immunosorbent assay for HSP70 and HSP27. These results were correlated with intraoperative somatosensory-evoked potentials measurements and postoperative paralysis. We find that the levels of these proteins in many patients are elevated and that the degree of elevation correlates with the risk of permanent paralysis. We hypothesize that sequential measurement intraoperatively of the levels of the heat shock proteins HSP70 and HSP27 in the CSF can predict those patients who are at greatest risk for paralysis during thoracic aneurysm surgery and will allow us to develop means of preventing or attenuating this severe and often fatal complication.

  12. Solar radiation stress in climbing snails: behavioural and intrinsic features define the Hsp70 level in natural populations of Xeropicta derbentina (Pulmonata).

    Science.gov (United States)

    Di Lellis, Maddalena A; Seifan, Merav; Troschinski, Sandra; Mazzia, Christophe; Capowiez, Yvan; Triebskorn, Rita; Köhler, Heinz-R

    2012-11-01

    Ectotherms from sunny and hot environments need to cope with solar radiation. Mediterranean land snails of the superfamily Helicoidea feature a behavioural strategy to escape from solar radiation-induced excessive soil heating by climbing up vertical objects. The height of climbing, and also other parameters like shell colouration pattern, shell orientation, shell size, body mass, actual internal and shell surface temperature, and the interactions between those factors may be expected to modulate proteotoxic effects in snails exposed to solar radiation and, thus, their stress response. Focussing on natural populations of Xeropicta derbentina, we conducted a 'snapshot' field study using the individual Hsp70 level as a proxy for proteotoxic stress. In addition to correlation analyses, an IT-model selection approach based on Akaike's Information Criterion was applied to evaluate a set of models with respect to their explanatory power and to assess the relevance of each of the above-mentioned parameters for individual stress, by model averaging and parameter estimation. The analysis revealed particular importance of the individuals' shell size, height above ground, the shell colouration pattern and the interaction height × orientation. Our study showed that a distinct set of behavioural traits and intrinsic characters define the Hsp70 level and that environmental factors and individual features strongly interact.

  13. Histopathological alterations and induction of hsp70 in ramshorn snail (Marisa cornuarietis) and zebrafish (Danio rerio) embryos after exposure to PtCl(2).

    Science.gov (United States)

    Osterauer, Raphaela; Köhler, Heinz-R; Triebskorn, Rita

    2010-08-01

    The platinum group metals (PGMs) platinum (Pt), palladium (Pd), and rhodium (Rh) are used in automobile catalytic converters, from which they have been emitted into the environment to an increasing degree during the last 20 years. Despite the bioavailability of these metals to plants and animals, studies determining the effects of PGMs on organisms are extremely rare. In the present study, effects of various concentrations of PtCl(2) (0.1, 1, 10, 50 and 100 microg/L) were investigated with respect to the induction of hsp70 and histopathological alterations in the zebrafish, Danio rerio and the ramshorn snail, Marisa cornuarietis. Histopathological investigations revealed effects of Pt on both species, which varied between slight and strong cellular reactions, depending on the PtCl(2) concentration. The hsp70 level in M. cornuarietis did not show an increase following Pt exposure whereas it was significantly elevated at 100 micorg/L PtCl(2) in D. rerio. Copyright 2010 Elsevier B.V. All rights reserved.

  14. Expression profiling of heat shock genes in a scuttle fly Megaselia scalaris (Diptera, Phoridae).

    Science.gov (United States)

    Malewski, Tadeusz; Bogdanowicz, Wiesław; Durska, Ewa; Łoś, Marta; Kamiński, Marcin; Kowalewska, Katarzyna

    2015-10-18

    The Phoridae are a family of small, hump-backed flies, dominating in post-fire areas. Some of these flies are probably able to survive a fire as an egg, larva, or pupa, and may be adapted to the fire-altered environment at the genomic level. In this study, we describe the influence of short-term temperature treatment on the expression of seven heat shock protein genes in the third-instar larvae and imagoes of a scuttle fly Megaselia scalaris-one of the cosmopolitan and polyphagous phorids. In terms of the response to temperature treatment, these genes tested against tubulin as a reference split into three classes. The first class consists of hsp22 (larvae), hsp23 (larvae), and hsp26 (both larvae and imagoes), and is upregulated at the lowest temperature (33°C). The second class consists of hsp22 (imagoes), hsp23 (imagoes), hsp40 (larvae and imagoes), and hsp70 (larvae and imagoes), and is upregulated or induced at 37°C. Expression of genes of the third class (hsp27 and hsp83-larvae and imagoes) increased after treatment at 41°C temperature. Expression of the first two classes of genes occurred at a temperature lower than the LT50 of larvae and imagoes. The fact that there is a gap between the temperature upregulating hsp genes and the temperature leading to the loss of viability suggests that not only the level of hsp gene expression but also the temperature at which gene expression increased is important in an adaptation of M. scalaris to harsh environment. J. Exp. Zool. 9999A:XX-XX, 2015. © 2015 Wiley Periodicals, Inc. © 2015 Wiley Periodicals, Inc.

  15. Heritable variation in heat shock gene expression: a potential mechanism for adaptation to thermal stress in embryos of sea turtles.

    Science.gov (United States)

    Tedeschi, J N; Kennington, W J; Tomkins, J L; Berry, O; Whiting, S; Meekan, M G; Mitchell, N J

    2016-01-13

    The capacity of species to respond adaptively to warming temperatures will be key to their survival in the Anthropocene. The embryos of egg-laying species such as sea turtles have limited behavioural means for avoiding high nest temperatures, and responses at the physiological level may be critical to coping with predicted global temperature increases. Using the loggerhead sea turtle (Caretta caretta) as a model, we used quantitative PCR to characterise variation in the expression response of heat-shock genes (hsp60, hsp70 and hsp90; molecular chaperones involved in cellular stress response) to an acute non-lethal heat shock. We show significant variation in gene expression at the clutch and population levels for some, but not all hsp genes. Using pedigree information, we estimated heritabilities of the expression response of hsp genes to heat shock and demonstrated both maternal and additive genetic effects. This is the first evidence that the heat-shock response is heritable in sea turtles and operates at the embryonic stage in any reptile. The presence of heritable variation in the expression of key thermotolerance genes is necessary for sea turtles to adapt at a molecular level to warming incubation environments. © 2016 The Author(s).

  16. Interference with plastome gene expression and Clp protease activity in Arabidopsis triggers a chloroplast unfolded protein response to restore protein homeostasis.

    Science.gov (United States)

    Llamas, Ernesto; Pulido, Pablo; Rodriguez-Concepcion, Manuel

    2017-09-01

    Disruption of protein homeostasis in chloroplasts impairs the correct functioning of essential metabolic pathways, including the methylerythritol 4-phosphate (MEP) pathway for the production of plastidial isoprenoids involved in photosynthesis and growth. We previously found that misfolded and aggregated forms of the first enzyme of the MEP pathway are degraded by the Clp protease with the involvement of Hsp70 and Hsp100/ClpC1 chaperones in Arabidopsis thaliana. By contrast, the combined unfolding and disaggregating actions of Hsp70 and Hsp100/ClpB3 chaperones allow solubilization and hence reactivation of the enzyme. The repair pathway is promoted when the levels of ClpB3 proteins increase upon reduction of Clp protease activity in mutants or wild-type plants treated with the chloroplast protein synthesis inhibitor lincomycin (LIN). Here we show that LIN treatment rapidly increases the levels of aggregated proteins in the chloroplast, unleashing a specific retrograde signaling pathway that up-regulates expression of ClpB3 and other nuclear genes encoding plastidial chaperones. As a consequence, folding capacity is increased to restore protein homeostasis. This sort of chloroplast unfolded protein response (cpUPR) mechanism appears to be mediated by the heat shock transcription factor HsfA2. Expression of HsfA2 and cpUPR-related target genes is independent of GUN1, a central integrator of retrograde signaling pathways. However, double mutants defective in both GUN1 and plastome gene expression (or Clp protease activity) are seedling lethal, confirming that the GUN1 protein is essential for protein homeostasis in chloroplasts.

  17. Interference with plastome gene expression and Clp protease activity in Arabidopsis triggers a chloroplast unfolded protein response to restore protein homeostasis.

    Directory of Open Access Journals (Sweden)

    Ernesto Llamas

    2017-09-01

    Full Text Available Disruption of protein homeostasis in chloroplasts impairs the correct functioning of essential metabolic pathways, including the methylerythritol 4-phosphate (MEP pathway for the production of plastidial isoprenoids involved in photosynthesis and growth. We previously found that misfolded and aggregated forms of the first enzyme of the MEP pathway are degraded by the Clp protease with the involvement of Hsp70 and Hsp100/ClpC1 chaperones in Arabidopsis thaliana. By contrast, the combined unfolding and disaggregating actions of Hsp70 and Hsp100/ClpB3 chaperones allow solubilization and hence reactivation of the enzyme. The repair pathway is promoted when the levels of ClpB3 proteins increase upon reduction of Clp protease activity in mutants or wild-type plants treated with the chloroplast protein synthesis inhibitor lincomycin (LIN. Here we show that LIN treatment rapidly increases the levels of aggregated proteins in the chloroplast, unleashing a specific retrograde signaling pathway that up-regulates expression of ClpB3 and other nuclear genes encoding plastidial chaperones. As a consequence, folding capacity is increased to restore protein homeostasis. This sort of chloroplast unfolded protein response (cpUPR mechanism appears to be mediated by the heat shock transcription factor HsfA2. Expression of HsfA2 and cpUPR-related target genes is independent of GUN1, a central integrator of retrograde signaling pathways. However, double mutants defective in both GUN1 and plastome gene expression (or Clp protease activity are seedling lethal, confirming that the GUN1 protein is essential for protein homeostasis in chloroplasts.

  18. Micronucleus-specific bacterium Holospora elegans irreversibly enhances stress gene expression of the host Paramecium caudatum.

    Science.gov (United States)

    Hori, Manabu; Fujii, Kimiko; Fujishima, Masahiro

    2008-01-01

    The bacterium Holospora is an endonuclear symbiont of the ciliate Paramecium. Previously, we reported that paramecia bearing the macronuclear-specific symbiont Holospora obtusa survived better than symbiont-free paramecia, even under high temperatures unsuitable for growth. The paramecia with symbionts expressed high levels of hsp70 mRNAs even at 25 degrees C, a usual growth temperature. We report herein that paramecia bearing the micronuclear-specific symbiont Holospora elegans also acquire the heat-shock resistance. Even after the removal of the bacteria from the hosts by treatment with penicillin, the resulting aposymbiotic paramecia nevertheless maintained their heat shock-resistant nature for over 1 yr. Like symbiotic paramecia, these aposymbiotic paramecia also expressed high levels of both hsp60 and hsp70 mRNAs even at 25 degrees C. Moreover, analysis by fluorescent in situ hybridization with a probe specific for Holospora 16S rRNA revealed that the 16S rRNA of H. elegans was expressed around the nucleoli of the macronucleus in the aposymbiotic cells. This result suggests the possible transfer of Holospora genomic DNA from the micronucleus into the macronucleus in symbiotic paramecia. Perhaps this exogenous DNA could trigger the aposymbiotic paramecia to induce a stress response, inducing higher expression of Hsp60 and Hsp70, and thus conferring heat-shock resistance.

  19. Review article: Heat shock protein 70 (Hsp70 peptide activated Natural Killer (NK cells for the treatment of patients with non-small cell lung cancer (NSCLC after radiochemotherapy (RCTx – from preclinical studies to a clinical phase II trial

    Directory of Open Access Journals (Sweden)

    Hanno M Specht

    2015-04-01

    Full Text Available Heat shock protein 70 (Hsp70 is frequently overexpressed in tumor cells. An unusual cell surface localization could be demonstrated on a large variety of solid tumors including lung, colorectal, breast, squamous cell carcinomas of the head and neck, prostate and pancreatic carcinomas, glioblastomas, sarcomas and hematological malignancies, but not on corresponding normal tissues. A membrane (mHsp70-positive phenotype can be determined either directly on single cell suspensions of tumor biopsies by flow cytometry using cmHsp70.1 monoclonal antibody or indirectly in the serum of patients using a novel lipHsp70 ELISA. A mHsp70-positive tumor phenotype has been associated with highly aggressive tumors, causing invasion and metastases and resistance to cell death. However, natural killer (NK, but not T cells were found to kill mHsp70-positive tumor cells after activation with a naturally occurring Hsp70 peptide (TKD plus low dose IL-2 (TKD/IL-2. Safety and tolerability of ex vivo TKD/IL-2 stimulated, autologous NK cells has been demonstrated in patients with metastasized colorectal and NSCLC in a phase I clinical trial. Based on promising clinical results of the previous study, a phase II randomized clinical study was initiated in 2015. The primary objective of this multicenter proof-of-concept trial is to examine whether an adjuvant treatment of NSCLC patients after platinum based radiochemotherapy with TKD/IL-2 activated, autologous NK cells is clinically effective. As a mHsp70-positive tumor phenotype is associated with poor clinical outcome only mHsp70-positive tumor patients will be recruited into the trial. The primary endpoint of this study will be the comparison of the progression-free survival of patients treated with ex vivo activated NK cells compared to patients who were treated with radiochemotherapy alone. As secondary endpoints overall survival, toxicity, quality-of-life and biological responses will be determined in both study groups.

  20. Effect of multiple microsporidian infections and temperature stress on the heat shock protein 70 (hsp70) response of the amphipod Gammarus pulex

    Science.gov (United States)

    2014-01-01

    Background Increasing temperatures can be a significant stressor for aquatic organisms. Amphipods are one of the most abundant and functionally important groups of freshwater macroinvertebrates. Therefore, we conducted a laboratory experiment with Gammarus pulex, naturally infected with microsporidians. Methods In each group, 42 gammarids were exposed to 15°C and 25°C for 24 h. Sex of gammarids was determined and microsporidian infections were detected by specific PCR. To quantify stress levels of the amphipods, the 70 kDa heat shock proteins (hsp70) were analyzed by western blot. Results More males than females were detected in the randomized population sample (ratio of females/males: 0.87). No mortality occurred at 15°C, while 42.9% of gammarids died at 25°C. Sequences of three microsporidians (M1, M2, M3) were detected in this G. pulex population (99.7%-100% sequence identity to Microsporidium spp. from GenBank). Previous studies showed that M3 is vertically transmitted, while M1 and M2 are presumably horizontally transmitted. Prevalences, according to PCR, were 27.0%, 37.8% and 64.9% for Microsporidium sp. M1, M2 and M3, respectively. Cumulative prevalence was 82.4%. Multiple infections with all three microsporidians in single gammarids were detected with a prevalence of 8.1%, and bi-infections ranged between 12.2% and 25.7%. In dead gammarids, comparatively low prevalences were noted for M1 (males and females: 11.1%) and M2 (females: 11.1%; males 0%), while prevalence of M3 was higher (females: 66.7%; males: 88.9%). No significant effect of host sex on microsporidian infection was found. Significant effects of temperature and bi-infection with Microsporidium spp. M2 + M3 on hsp70 response were detected by analysis of the whole sample (15°C and 25°C group) and of M2 + M3 bi-infection and gammarid weight when analyzing the 25°C group separately. None of the parameters had a significant effect on hsp70 levels in the 15°C group. Conclusion This

  1. Probing Allosteric Inhibition Mechanisms of the Hsp70 Chaperone Proteins Using Molecular Dynamics Simulations and Analysis of the Residue Interaction Networks.

    Science.gov (United States)

    Stetz, Gabrielle; Verkhivker, Gennady M

    2016-08-22

    Although molecular mechanisms of allosteric regulation in the Hsp70 chaperones have been extensively studied at both structural and functional levels, the current understanding of allosteric inhibition of chaperone activities by small molecules is still lacking. In the current study, using a battery of computational approaches, we probed allosteric inhibition mechanisms of E. coli Hsp70 (DnaK) and human Hsp70 proteins by small molecule inhibitors PET-16 and novolactone. Molecular dynamics simulations and binding free energy analysis were combined with network-based modeling of residue interactions and allosteric communications to systematically characterize and compare molecular signatures of the apo form, substrate-bound, and inhibitor-bound chaperone complexes. The results suggested a mechanism by which the allosteric inhibitors may leverage binding energy hotspots in the interaction networks to stabilize a specific conformational state and impair the interdomain allosteric control. Using the network-based centrality analysis and community detection, we demonstrated that substrate binding may strengthen the connectivity of local interaction communities, leading to a dense interaction network that can promote an efficient allosteric communication. In contrast, binding of PET-16 to DnaK may induce significant dynamic changes and lead to a fractured interaction network and impaired allosteric communications in the DnaK complex. By using a mechanistic-based analysis of distance fluctuation maps and allosteric propensities of protein residues, we determined that the allosteric network in the PET-16 complex may be small and localized due to the reduced communication and low cooperativity of the substrate binding loops, which may promote the higher rates of substrate dissociation and the decreased substrate affinity. In comparison with the significant effect of PET-16, binding of novolactone to HSPA1A may cause only moderate network changes and preserve allosteric

  2. Matrix protein of VSV New Jersey serotype containing methionine to arginine substitutions at positions 48 and 51 allows near-normal host cell gene expression.

    Science.gov (United States)

    Kim, Gyoung Nyoun; Kang, C Yong

    2007-01-05

    The matrix (M) protein of vesicular stomatitis virus (VSV) plays significant roles in the replication of VSV through its involvement in the assembly of virus particles as well as by facilitating the evasion of innate host cell defense mechanisms. The presence of methionine at position 51 (M51) of the matrix (M) protein of the VSV Indiana serotype (VSV(Ind)) has been proven to be crucial for cell rounding and inhibition of host cell gene expression. The M protein of VSV(Ind) with the substitution of M51 with arginine (R:M51R) results in the loss of inhibitory effects on host cell gene expression. The VSV(Ind) expressing the M(M51R) protein became the attractive oncolytic virus which is safer and more tumor-specific because the normal cells can clear the mutant VSV(Ind) easily but tumor cells are susceptible to the virus because a variety of tumor cells lack innate antiviral activities. We have studied the role of the methionines at positions 48 and 51 of the M protein of the New Jersey serotype of VSV (VSV(NJ)) in the induction of cytopathic effects (CPE) and host cell gene expression. We have generated human embryonic kidney 293 cell lines inducibly expressing M proteins with M to R mutations at positions 48 and 51, either separately or together as a double mutant, and examined expression of heat shock protein 70 (HSP70) as an indicator of host cell gene expression. We have also generated recombinant VSV(NJ) encoding the mutant M proteins M(M48R) or M(M48R+M51R) for the first time and tested for the expression of HSP70 in infected cells. Our results demonstrated that the M51 of VSV(NJ) M proteins has a major role in cell rounding and in suppressing the host cell gene expression either when the M protein was expressed alone in inducible cell lines or when expressed together with other VSV proteins by the recombinant VSV(NJ). Amino acid residue M48 may also have some role in cell rounding and in the inhibitory effects of VSV(NJ) M, which was demonstrated by the fact

  3. Evaluation of the toxic potential of aspartame in third instar larvae of transgenic Drosophila melanogaster (hsp70-lacZ Bg9

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    YASIR HASAN SIDDIQUE

    2017-07-01

    Full Text Available Aspartame is commonly used as a sweetener in prepared foods, beverages and in recipes. In the present study the possible toxic effects of aspartame were studied in third instar larvae of transgenic Drosophila melanogaster (hsp70-lacZ Bg9. The third instar larvae were allowed to feed on the diet having different doses of aspartame. The results obtained in the present study suggest that the exposure of third instar larvae to the diet having 20, 40, 80, 160 and 320 mM of aspartame did not show any significant the toxic effects after 24 hours of the exposure compared to control (p<0.05. The results for pupae formation and emergence of flies also did not show any significant difference compared to control (p<0.05.

  4. Two years of combined high-intensity physical training and heat acclimatization affect lymphocyte and serum HSP70 in purebred military working dogs.

    Science.gov (United States)

    Bruchim, Yaron; Aroch, Itamar; Eliav, Ady; Abbas, Atallah; Frank, Ilan; Kelmer, Efrat; Codner, Carolina; Segev, Gilad; Epstein, Yoram; Horowitz, Michal

    2014-07-15

    Military working dogs in hot countries undergo exercise training at high ambient temperatures for at least 9 mo annually. Physiological adaptations to these harsh conditions have been extensively studied; however, studies focusing on the underlying molecular adaptations are limited. In the current study, military working dogs were chosen as a model to examine the effects of superimposing endurance exercise on seasonal acclimatization to environmental heat stress. The lymphocyte HSP70 profile and extracellular HSP70 were studied in tandem with physiological performance in the dogs from their recruitment for the following 2 yr. Aerobic power and heat shock proteins were measured at the end of each summer, with physical performance tests (PPTs) in an acclimatized room (22°C). The study shows that together with a profound enhancement of aerobic power and physical performance, hsp72 mRNA induction immediately post-PPT and 45 min later, progressively increased throughout the study period (relative change in median lymphocyte hsp72 mRNA first PPT, 4.22 and 12.82; second PPT, 17.19 and 109.05, respectively), whereas induction of HSP72 protein was stable. These responses suggest that cellular/molecular adaptive tools for maintaining HSP72 homeostasis exist. There was also a significant rise in basal and peak median optical density extracellular HSP at the end of each exercise test (first PPT, 0.13 and 0.15; second PPT, 1.04 and 1.52, respectively). The relationship between these enhancements and improved aerobic power capacity is not yet fully understood. Copyright © 2014 the American Physiological Society.

  5. Gene expression profiling in hepatic tissue of newly weaned pigs fed pharmacological zinc and phytase supplemented diets

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    Link Jane E

    2008-09-01

    Full Text Available Abstract Background Zinc (Zn is an essential trace element. However, Zn bioavailability from commonly consumed plants may be reduced due to phytic acid. Zn supplementation has been used to treat diarrheal disease in children, and in the U.S. swine industry at pharmacological levels to promote growth and fecal consistency, but underlying mechanisms explaining these beneficial effects remain unknown. Moreover, adding supplemental phytase improves Zn bioavailability. Thus, we hypothesized that benefits of pharmacological Zn supplementation result from changes in gene expression that could be further affected by supplemental phytase. The goal of this study was to investigate the effects of feeding newly weaned pigs dietary Zn (150, 1,000, or 2,000 mg Zn/kg as Zn oxide with or without phytase [500 phytase units (FTU/kg] for 14 d on hepatic gene expression. Liver RNA from pigs fed 150, 1,000, or 2,000 mg Zn/kg, or 1,000 mg Zn/kg with phytase (n = 4 per treatment was reverse transcribed and examined using the differential display reverse transcription polymerase chain reaction technique. Liver RNA from pigs fed 150 or 2,000 mg Zn/kg (n = 4 per treatment was also evaluated using a 70-mer oligonucleotide microarray. Results Expressed sequence tags for 61 putatively differentially expressed transcripts were cloned and sequenced. In addition, interrogation of a 13,297 element oligonucleotide microarray revealed 650 annotated transcripts (FDR ≤ 0.05 affected by pharmacological Zn supplementation. Seven transcripts exhibiting differential expression in pigs fed pharmacological Zn with sequence similarities to genes encoding GLO1, PRDX4, ACY1, ORM1, CPB2, GSTM4, and HSP70.2 were selected for confirmation. Relative hepatic GLO1 (P PRDX4 (P ACY1 (P HSP70.2 (P Conclusion Results suggest that feeding pharmacological Zn (1,000 or 2,000 mg Zn/kg affects genes involved in reducing oxidative stress and in amino acid metabolism, which are essential for cell

  6. Liposome-based DNA carriers may induce cellular stress response and change gene expression pattern in transfected cells

    Science.gov (United States)

    2011-01-01

    Background During functional studies on the rat stress-inducible Hspa1b (hsp70.1) gene we noticed that some liposome-based DNA carriers, which are used for transfection, induce its promoter activity. This observation concerned commercial liposome formulations (LA), Lipofectin and Lipofectamine 2000. This work was aimed to understand better the mechanism of this phenomenon and its potential biological and practical consequences. Results We found that a reporter gene driven by Hspa1b promoter is activated both in the case of transient transfections and in the stably transfected cells treated with LA. Using several deletion clones containing different fragments of Hspa1b promoter, we found that the regulatory elements responsible for most efficient LA-driven inducibility were located between nucleotides -269 and +85, relative to the transcription start site. Further studies showed that the induction mechanism was independent of the classical HSE-HSF interaction that is responsible for gene activation during heat stress. Using DNA microarrays we also detected significant activation of the endogenous Hspa1b gene in cells treated with Lipofectamine 2000. Several other stress genes were also induced, along with numerous genes involved in cellular metabolism, cell cycle control and pro-apoptotic pathways. Conclusions Our observations suggest that i) some cationic liposomes may not be suitable for functional studies on hsp promoters, ii) lipofection may cause unintended changes in global gene expression in the transfected cells. PMID:21663599

  7. Gene Expression Profiles of NO- and HNO-donor Treated Breast Cancer Cells: Insights into Tumor Response and Resistance Pathways

    Science.gov (United States)

    Cheng, Robert YS; Basudhar, Debashree; Ridnour, Lisa A.; Heinecke, Julie L.; Kesarwala, Aparna H.; Glynn, Sharon; Switzer, Christopher H.; Ambs, Stefan; Miranda, Katrina M.; Wink, David A.

    2014-01-01

    Nitric oxide (NO) synthase 2 (NOS2), a major inflammatory protein, modulates disease progression via NO in a number of pathologies, including cancer. The role of NOS2-derived NO is not only flux-dependent, which is higher in mouse vs. human cells, but also varies based on spatial and temporal distribution both within tumor cells and in the tumor microenvironment. NO donors have been utilized to mimic NO flux conditions and to investigate the effects of varied NO concentrations. As a wide range of effects mediated by NO and other nitrogen oxides such as nitroxyl (HNO) have been elucidated, multiple NO- and HNO-releasing compounds have been developed as potential therapeutics, including as tumor modulators. One of the challenges is to determine differences in biomarker expression from extracellular vs. intracellular generation of NO or HNO. Taking advantage of new NO and HNO releasing agents, we have characterized the gene expression profile of estrogen receptor-negative human breast cancer (MDA-MB-231) cells following exposure to aspirin, the NO donor DEA/NO, the HNO donor IPA/NO and their intracellularly-activated prodrug conjugates DEA/NO-aspirin and IPA/NO-aspirin. Comparison of the gene expression profiles demonstrated that several genes were uniquely expressed with respect to NO or HNO, such as miR-21, HSP70, cystathionine γ-lyase and IL24. These findings provide insight into targets and pathways that could be therapeutically exploited by the redox related species NO and HNO. PMID:25153034

  8. Expression of heat shock protein70 in oral submucous fibrosis and oral squamous cell carcinoma: An immunohistochemical study

    Directory of Open Access Journals (Sweden)

    M Thubashini

    2011-01-01

    Conclusion: HSP70 is synthesized upon stress situations arising in cells of all living organisms. Expression of HSP70 indicates that stress plays an important role as a predisposing factor in oral submucous fibrosis and its subsequent progression to oral squamous cell carcinoma.

  9. glsA, a Volvox gene required for asymmetric division and germ cell specification, encodes a chaperone-like protein.

    Science.gov (United States)

    Miller, S M; Kirk, D L

    1999-02-01

    The gls genes of Volvox are required for the asymmetric divisions that set apart cells of the germ and somatic lineages during embryogenesis. Here we used transposon tagging to clone glsA, and then showed that it is expressed maximally in asymmetrically dividing embryos, and that it encodes a 748-amino acid protein with two potential protein-binding domains. Site-directed mutagenesis of one of these, the J domain (by which Hsp40-class chaperones bind to and activate specific Hsp70 partners) abolishes the capacity of glsA to rescue mutants. Based on this and other considerations, including the fact that the GlsA protein is associated with the mitotic spindle, we discuss how it might function, in conjunction with an Hsp70-type partner, to shift the division plane in asymmetrically dividing cells.

  10. Selection and assessment of reference genes for quantitative PCR normalization in migratory locust Locusta migratoria (Orthoptera: Acrididae.

    Directory of Open Access Journals (Sweden)

    Qingpo Yang

    Full Text Available Locusta migratoria is a classic hemimetamorphosis insect and has caused widespread economic damage to crops as a migratory pest. Researches on the expression pattern of functional genes in L. migratoria have drawn focus in recent years, especially with the release of genome information. Real-time quantitative PCR is the most reproducible and sensitive approach for detecting transcript expression levels of target genes, but optimal internal standards are key factors for its accuracy and reliability. Therefore, it's necessary to provide a systematic stability assessment of internal control for well-performed tests of target gene expression profile. In this study, twelve candidate genes (Ach, Act, Cht2, EF1α, RPL32, Hsp70, Tub, RP49, SDH, GAPDH, 18S, and His were analyzed with four statistical methods: the delta Ct approach, geNorm, Bestkeeper and NormFinder. The results from these analyses aimed to choose the best suitable reference gene across different experimental situations for gene profile study in L. migratoria. The result demonstrated that for different developmental stages, EF1α, Hsp70 and RPL32 exhibited the most stable expression status for all samples; EF1α and RPL32 were selected as the best reference genes for studies involving embryo and larvae stages, while SDH and RP49 were identified for adult stage. The best-ranked reference genes across different tissues are RPL32, Hsp70 and RP49. For abiotic treatments, the most appropriate genes we identified were as follows: Act and SDH for larvae subjected to different insecticides; RPL32 and Ach for larvae exposed to different temperature treatments; and Act and Ach for larvae suffering from starvation. The present report should facilitate future researches on gene expression in L. migratoria with accessibly optimal reference genes under different experimental contexts.

  11. Quantitative expression profiling of immune response genes in rainbow trout following infectious haematopoietic necrosis virus (IHNV) infection or DNA vaccination

    Science.gov (United States)

    Purcell, Maureen K.; Kurath, Gael; Garver, Kyle A.; Herwig, Russell P.; Winton, James R.

    2004-01-01

    Infectious haematopoietic necrosis virus (IHNV) is a well-studied virus of salmonid fishes. A highly efficacious DNA vaccine has been developed against this virus and studies have demonstrated that this vaccine induces both an early and transient non-specific anti-viral phase as well as long-term specific protection. The mechanisms of the early anti-viral phase are not known, but previous studies noted changes in Mx gene expression, suggesting a role for type I interferon. This study used quantitative real-time reverse transcriptase PCR methodology to compare expression changes over time of a number of cytokine or cytokine-related genes in the spleen of rainbow trout following injection with poly I:C, live IHNV, the IHNV DNA vaccine or a control plasmid encoding the non-antigenic luciferase gene. The target genes included Mx-1, viral haemorrhagic septicaemia virus induced gene 8 (Vig-8), TNF-α1, TNF-α2, IL-1β1, IL-8, TGF-β1 and Hsp70. Poly I:C stimulation induced several genes but the strongest and significant response was observed in the Mx-1 and Vig-8 genes. The live IHN virus induced a significant response in all genes examined except TGF-β1. The control plasmid construct and the IHNV DNA vaccine marginally induced a number of genes, but the main difference between these two groups was a statistically significant induction of the Mx-1 and Vig-8 genes by the IHNV vaccine only. The gene expression profiles elicited by the live virus and the IHNV DNA vaccine differed in a number of aspects but this study confirms the clear role for a type I interferon-like response in early anti-viral defence.

  12. Sample size matters in dietary gene expression studies—A case study in the gilthead sea bream (Sparus aurata L.

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    Fotini Kokou

    2016-05-01

    Full Text Available One of the main concerns in gene expression studies is the calculation of statistical significance which in most cases remains low due to limited sample size. Increasing biological replicates translates into more effective gains in power which, especially in nutritional experiments, is of great importance as individual variation of growth performance parameters and feed conversion is high. The present study investigates in the gilthead sea bream Sparus aurata, one of the most important Mediterranean aquaculture species. For 24 gilthead sea bream individuals (biological replicates the effects of gradual substitution of fish meal by plant ingredients (0% (control, 25%, 50% and 75% in the diets were studied by looking at expression levels of four immune-and stress-related genes in intestine, head kidney and liver. The present results showed that only the lowest substitution percentage is tolerated and that liver is the most sensitive tissue to detect gene expression variations in relation to fish meal substituted diets. Additionally the usage of three independent biological replicates were evaluated by calculating the averages of all possible triplets in order to assess the suitability of selected genes for stress indication as well as the impact of the experimental set up, thus in the present work the impact of FM substitution. Gene expression was altered depending of the selected biological triplicate. Only for two genes in liver (hsp70 and tgf significant differential expression was assured independently of the triplicates used. These results underlined the importance of choosing the adequate sample number especially when significant, but minor differences in gene expression levels are observed.

  13. Gastroprotective activity of Annona muricata leaves against ethanol-induced gastric injury in rats via Hsp70/Bax involvement

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    Moghadamtousi SZ

    2014-10-01

    downregulation of Bax and upregulation of Hsp70 proteins after pretreatment. Collectively, the present results suggest that EEAM has a promising antiulcer potential, which could be attributed to its suppressive effect against oxidative damage and preservative effect toward gastric wall mucus. Keywords: Annona muricata, annonaceae, gastric injury, antioxidants, Hsp70/Bax

  14. Vitamin A Oral Supplementation Induces Oxidative Stress and Suppresses IL-10 and HSP70 in Skeletal Muscle of Trained Rats

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    Lyvia Lintzmaier Petiz

    2017-04-01

    Full Text Available Exercise training intensity is the major variant that influences the relationship between exercise, redox balance, and immune response. Supplement intake is a common practice for oxidative stress prevention; the effects of vitamin A (VA on exercise training are not yet described, even though this molecule exhibits antioxidant properties. We investigated the role of VA supplementation on redox and immune responses of adult Wistar rats subjected to swimming training. Animals were divided into four groups: sedentary, sedentary + VA, exercise training, and exercise training + VA. Over eight weeks, animals were submitted to intense swimming 5 times/week and a VA daily intake of 450 retinol equivalents/day. VA impaired the total serum antioxidant capacity acquired by exercise, with no change in interleukin-1β and tumor necrosis factor-α levels. In skeletal muscle, VA caused lipid peroxidation and protein damage without differences in antioxidant enzyme activities; however, Western blot analysis showed that expression of superoxide dismutase-1 was downregulated, and upregulation of superoxide dismutase-2 induced by exercise was blunted by VA. Furthermore, VA supplementation decreased anti-inflammatory interleukin-10 and heat shock protein 70 expression, important factors for positive exercise adaptations and tissue damage prevention. Our data showed that VA supplementation did not confer any antioxidative and/or protective effects, attenuating exercise-acquired benefits in the skeletal muscle.

  15. Involvement of NF-ΚB and HSP70 signaling pathways in the apoptosis of MDA-MB-231 cells induced by a prenylated xanthone compound, α-mangostin, from Cratoxylum arborescens

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    Ibrahim MY

    2014-11-01

    the role of the central apoptosis-related proteins, a protein array followed by immunoblot analysis was conducted. Moreover, the involvement of nuclear factor-kappa B (NF-ΚB was also analyzed. Results: Apoptosis was confirmed by the apoptotic cells stained with annexin V and increase in chromatin condensation in nucleus. Treatment of cells with AM promoted cell death-transducing signals that reduced MMP by downregulation of Bcl-2 and upregulation of Bax, triggering cytochrome c release from the mitochondria to the cytosol. The released cytochrome c triggered the activation of caspase-9 followed by the executioner caspase-3/7 and then cleaved the PARP protein. Increase of caspase-8 showed the involvement of extrinsic pathway. AM treatment significantly arrested the cells at the S phase (P<0.05 concomitant with an increase in reactive oxygen species. The protein array and Western blotting demonstrated the expression of HSP70. Moreover, AM significantly blocked the induced translocation of NF-ΚB from cytoplasm to nucleus. Conclusion: Together, the results demonstrate that the AM isolated from C. arborescens inhibited the proliferation of MDA-MB-231 cells, leading to cell cycle arrest and programmed cell death, which was suggested to occur through both the extrinsic and intrinsic apoptosis pathways with involvement of the NF-ΚB and HSP70 signaling pathways. Keywords: mitochondria, protein array, caspase-3/7

  16. Novel UBQLN2 mutations linked to amyotrophic lateral sclerosis and atypical hereditary spastic paraplegia phenotype through defective HSP70-mediated proteolysis.

    Science.gov (United States)

    Teyssou, Elisa; Chartier, Laura; Amador, Maria-Del-Mar; Lam, Roselina; Lautrette, Géraldine; Nicol, Marie; Machat, Selma; Da Barroca, Sandra; Moigneu, Carine; Mairey, Mathilde; Larmonier, Thierry; Saker, Safaa; Dussert, Christelle; Forlani, Sylvie; Fontaine, Bertrand; Seilhean, Danielle; Bohl, Delphine; Boillée, Séverine; Meininger, Vincent; Couratier, Philippe; Salachas, François; Stevanin, Giovanni; Millecamps, Stéphanie

    2017-10-01

    Mutations in UBQLN2 have been associated with rare cases of X-linked juvenile and adult forms of amyotrophic lateral sclerosis (ALS) and ALS linked to frontotemporal dementia (FTD). Here, we report 1 known (c.1489C>T, p.Pro497Ser, P497S) and 3 novel (c.1481C>T, p.Pro494Leu, P494L; c.1498C>T, p.Pro500Ser, P500S; and c.1516C>G, p.Pro506Ala, P506A) missense mutations in the PXX domain of UBQLN2 in familial motor neuron diseases including ALS and spastic paraplegia (SP). A novel missense mutation (c.1462G>A, p.Ala488Thr, A488T) adjacent to this hotspot UBQLN2 domain was identified in a sporadic case of ALS. These mutations are conserved in mammals, are absent from ExAC and gnomAD browsers, and are predicted to be deleterious by SIFT in silico analysis. Patient lymphoblasts carrying a UBQLN2 mutation showed absence of ubiquilin-2 accumulation, disrupted binding with HSP70, and impaired autophagic pathway. Our results confirm the role of PXX repeat in ALS pathogenesis, show that UBQLN2-linked disease can manifest like a SP phenotype, evidence a highly reduced disease penetrance in females carrying UBQLN2 mutations, which is important information for genetic counseling, and underline the pivotal role of ubiquilin-2 in proteolysis regulation pathways. Copyright © 2017 Elsevier Inc. All rights reserved.

  17. CHIP, a carboxy terminus HSP-70 interacting protein, prevents cell death induced by endoplasmic reticulum stress in the central nervous system

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    Felipe eCabral Miranda

    2015-01-01

    Full Text Available Endoplasmic reticulum (ER stress and protein misfolding are associated with various neurodegenerative diseases. ER stress activates Unfolded Protein Response (UPR, an adaptative response. However, severe ER stress can induce cell death. Here we show that the E3 ubiquitin ligase and co-chaperone Carboxyl Terminus HSP70/90 Interacting Protein (CHIP prevents neuron death in the hippocampus induced by severe ER stress. Organotypic hippocampal slice cultures (OHSCs were exposed to Tunicamycin, a pharmacological ER stress inducer, to trigger cell death. Overexpression of CHIP was achieved with a recombinant adeno-associated viral vector (rAAV and significantly diminished ER stress-induced cell death, as shown by analysis of propidium iodide (PI uptake, condensed chromatin, TUNEL and cleaved caspase 3 in the CA1 region of OHSCs. In addition, overexpression of CHIP prevented upregulation of both CHOP and p53 both pro-apoptotic pathways induced by ER stress. We also detected an attenuation of eIF2a phosphorylation promoted by ER stress. However, CHIP did not prevent upregulation of BiP/GRP78 induced by UPR. These data indicate that overexpression of CHIP attenuates ER-stress death response while maintain ER stress adaptative response in the central nervous system. These results indicate a neuroprotective role for CHIP upon UPR signalling. CHIP emerge as a candidate for clinical intervention in neurodegenerative diseases associated with ER stress.

  18. Evolution of gene expression after gene amplification.

    Science.gov (United States)

    Garcia, Nelson; Zhang, Wei; Wu, Yongrui; Messing, Joachim

    2015-04-24

    We took a rather unique approach to investigate the conservation of gene expression of prolamin storage protein genes across two different subfamilies of the Poaceae. We took advantage of oat plants carrying single maize chromosomes in different cultivars, called oat-maize addition (OMA) lines, which permitted us to determine whether regulation of gene expression was conserved between the two species. We found that γ-zeins are expressed in OMA7.06, which carries maize chromosome 7 even in the absence of the trans-acting maize prolamin-box-binding factor (PBF), which regulates their expression. This is likely because oat PBF can substitute for the function of maize PBF as shown in our transient expression data, using a γ-zein promoter fused to green fluorescent protein (GFP). Despite this conservation, the younger, recently amplified prolamin genes in maize, absent in oat, are not expressed in the corresponding OMAs. However, maize can express the oldest prolamin gene, the wheat high-molecular weight glutenin Dx5 gene, even when maize Pbf is knocked down (through PbfRNAi), and/or another maize transcription factor, Opaque-2 (O2) is knocked out (in maize o2 mutant). Therefore, older genes are conserved in their regulation, whereas younger ones diverged during evolution and eventually acquired a new repertoire of suitable transcriptional activators. © The Author(s) 2015. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  19. Assessment of nuclear and mitochondrial genes in precise identification and analysis of genetic polymorphisms for the evaluation of Leishmania parasites.

    Science.gov (United States)

    Fotouhi-Ardakani, Reza; Dabiri, Shahriar; Ajdari, Soheila; Alimohammadian, Mohammad Hossein; AlaeeNovin, Elnaz; Taleshi, Neda; Parvizi, Parviz

    2016-12-01

    The polymorphism and genetic diversity of Leishmania genus has status under discussion depending on many items such as nuclear and/or mitochondrial genes, molecular tools, Leishmania species, geographical origin, condition of micro-environment of Leishmania parasites and isolation of Leishmania from clinical samples, reservoir host and vectors. The genetic variation of Leishmania species (L. major, L. tropica, L. tarentolae, L. mexicana, L. infantum) were analyzed and compared using mitochondrial (COII and Cyt b) and nuclear (nagt, ITS-rDNA and HSP70) genes. The role of each enzymatic (COII, Cyt b and nagt) or housekeeping (ITS-rDNA, HSP70) gene was employed for accurate identification of Leishmania parasites. After DNA extractions and amplifying of native, natural and reference strains of Leishmania parasites, polymerase chain reaction (PCR) products were sequenced and evaluation of genetic proximity and phylogenetic analysis were performed using MEGA6 and DnaSP5 software. Among the 72 sequences of the five genes, the number of polymorphic sites was significantly lower as compared to the monomorphic sites. Of the 72 sequences, 54 new haplotypes (five genes) of Leishmania species were submitted in GenBank (Access number: KU680818 - KU680871). Four genes had a remarkable number of informative sites (P=0.00), except HSP70 maybe because of its microsatellite regions. The non-synonymous (dN) variants of nagt gene were more than that of other expression genes (47.4%). The synonymous (dS)/dN ratio in three expression genes showed a significant variation between five Leishmania species (P=0.001). The highest and lowest levels of haplotype diversity were observed in L. tropica (81.35%) and L. major (28.38%) populations, respectively. Tajima's D index analyses showed that Cyt b gene in L. tropica species was significantly negative (Tajima's D=-2.2, PLeishmania parasites. Copyright © 2016 Elsevier B.V. All rights reserved.

  20. Stable Reference Gene Selection for RT-qPCR Analysis in Nonviruliferous and Viruliferous Frankliniella occidentalis.

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    Chunxiao Yang

    Full Text Available Reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR is a reliable technique for measuring and evaluating gene expression during variable biological processes. To facilitate gene expression studies, normalization of genes of interest relative to stable reference genes is crucial. The western flower thrips Frankliniella occidentalis (Pergande (Thysanoptera: Thripidae, the main vector of tomato spotted wilt virus (TSWV, is a destructive invasive species. In this study, the expression profiles of 11 candidate reference genes from nonviruliferous and viruliferous F. occidentalis were investigated. Five distinct algorithms, geNorm, NormFinder, BestKeeper, the ΔCt method, and RefFinder, were used to determine the performance of these genes. geNorm, NormFinder, BestKeeper, and RefFinder identified heat shock protein 70 (HSP70, heat shock protein 60 (HSP60, elongation factor 1 α, and ribosomal protein l32 (RPL32 as the most stable reference genes, and the ΔCt method identified HSP60, HSP70, RPL32, and heat shock protein 90 as the most stable reference genes. Additionally, two reference genes were sufficient for reliable normalization in nonviruliferous and viruliferous F. occidentalis. This work provides a foundation for investigating the molecular mechanisms of TSWV and F. occidentalis interactions.

  1. Homeobox gene expression in Brachiopoda

    DEFF Research Database (Denmark)

    Altenburger, Andreas; Martinez, Pedro; Wanninger, Andreas

    2011-01-01

    The molecular control that underlies brachiopod ontogeny is largely unknown. In order to contribute to this issue we analyzed the expression pattern of two homeobox containing genes, Not and Cdx, during development of the rhynchonelliform (i.e., articulate) brachiopod Terebratalia transversa....... Not is a homeobox containing gene that regulates the formation of the notochord in chordates, while Cdx (caudal) is a ParaHox gene involved in the formation of posterior tissues of various animal phyla. The T. transversa homolog, TtrNot, is expressed in the ectoderm from the beginning of gastrulation until...... formation. TtrNot expression is absent in unfertilized eggs, in embryos prior to gastrulation, and in settled individuals during and after metamorphosis. Comparison with the expression patterns of Not genes in other metazoan phyla suggests an ancestral role for this gene in gastrulation and germ layer...

  2. Effect of the militarily-relevant heavy metals, depleted uranium and heavy metal tungsten-alloy on gene expression in human liver carcinoma cells (HepG2).

    Science.gov (United States)

    Miller, Alexandra C; Brooks, Kia; Smith, Jan; Page, Natalie

    2004-01-01

    Depleted uranium (DU) and heavy-metal tungsten alloys (HMTAs) are dense heavy-metals used primarily in military applications. Chemically similar to natural uranium, but depleted of the higher activity 235U and 234U isotopes, DU is a low specific activity, high-density heavy metal. In contrast, the non-radioactive HMTAs are composed of a mixture of tungsten (91-93%), nickel (3-5%), and cobalt (2-4%) particles. The use of DU and HMTAs in military munitions could result in their internalization in humans. Limited data exist however, regarding the long-term health effects of internalized DU and HMTAs in humans. Both DU and HMTAs possess a tumorigenic transforming potential and are genotoxic and mutagenic in vitro. Using insoluble DU-UO2 and a reconstituted mixture of tungsten, nickel, cobalt (rWNiCo), we tested their ability to induce stress genes in thirteen different recombinant cell lines generated from human liver carcinoma cells (HepG2). The commercially available CAT-Tox (L) cellular assay consists of a panel of cell lines stably transfected with reporter genes consisting of a coding sequence for chloramphenicol acetyl transferase (CAT) under transcriptional control by mammalian stress gene regulatory sequences. DU, (5-50 microg/ml) produced a complex profile of activity demonstrating significant dose-dependent induction of the hMTIIA FOS, p53RE, Gadd153, Gadd45, NFkappaBRE, CRE, HSP70, RARE, and GRP78 promoters. The rWNiCo mixture (5-50 microg/ml) showed dose-related induction of the GSTYA, hMTIIA, p53RE, FOS, NFkappaBRE, HSP70, and CRE promoters. An examination of the pure metals, tungsten (W), nickel (Ni), and cobalt (Co), comprising the rWNiCo mixture, demonstrated that each metal exhibited a similar pattern of gene induction, but at a significantly decreased magnitude than that of the rWNiCo mixture. These data showed a synergistic activation of gene expression by the metals in the rWNiCo mixture. Our data show for the first time that DU and rWNiCo can

  3. Molecular Characterization of Heat Shock Protein 70-1 Gene of Goat (Capra hircus

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    Nitin Gade

    2010-01-01

    Full Text Available Heat shock protein 70 (HSP 70 plays a vital role by bestowing cytoprotection against diverse kinds of stresses. The ubiquitous HSP 70 proteins are the most abundant and temperature sensitive among all the HSPs. The present paper has characterized HSP70-1 cDNA in goat (Capra hircus. Total RNA isolated from goat peripheral blood mononuclear cells was reverse transcribed to cDNA that was used for amplification of HSP 70-1 gene. PCR product (1926 bp was cloned in pGEM-T easy vector and sequenced. Sequence analysis revealed 1926-bp-long open reading frame of HSP 70-1 gene encoding 641 amino acids in goat, as reported in cattle. At nucleotide level, goat HSP 70-1 was found to be 96–99% similar to that of sheep (partial, cattle, and buffalo whereas the similarity at amino acid level was 95–100%. Nonsynonymous substitutions exceeding synonymous substitutions indicate the evolution of this protein through positive selection among domestic animals. Goat and sheep appear to have diverged from a common ancestor in phylogenetic analysis. Predicted protein structures of goat HSP 70 protein obtained from deduced amino acid sequence indicated that the functional amino acids involved in chaperoning through ATPase hydrolytic cycle and in uncoating of clathrin coated vesicles are highly conserved.

  4. Human Lacrimal Gland Gene Expression.

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    Vinay Kumar Aakalu

    Full Text Available The study of human lacrimal gland biology and development is limited. Lacrimal gland tissue is damaged or poorly functional in a number of disease states including dry eye disease. Development of cell based therapies for lacrimal gland diseases requires a better understanding of the gene expression and signaling pathways in lacrimal gland. Differential gene expression analysis between lacrimal gland and other embryologically similar tissues may be helpful in furthering our understanding of lacrimal gland development.We performed global gene expression analysis of human lacrimal gland tissue using Affymetrix ® gene expression arrays. Primary data from our laboratory was compared with datasets available in the NLM GEO database for other surface ectodermal tissues including salivary gland, skin, conjunctiva and corneal epithelium.The analysis revealed statistically significant difference in the gene expression of lacrimal gland tissue compared to other ectodermal tissues. The lacrimal gland specific, cell surface secretory protein encoding genes and critical signaling pathways which distinguish lacrimal gland from other ectodermal tissues are described.Differential gene expression in human lacrimal gland compared with other ectodermal tissue types revealed interesting patterns which may serve as the basis for future studies in directed differentiation among other areas.

  5. Phylogenetic analysis of gene expression.

    Science.gov (United States)

    Dunn, Casey W; Luo, Xi; Wu, Zhijin

    2013-11-01

    Phylogenetic analyses of gene expression have great potential for addressing a wide range of questions. These analyses will, for example, identify genes that have evolutionary shifts in expression that are correlated with evolutionary changes in morphological, physiological, and developmental characters of interest. This will provide entirely new opportunities to identify genes related to particular phenotypes. There are, however, 3 key challenges that must be addressed for such studies to realize their potential. First, data on gene expression must be measured from multiple species, some of which may be field-collected, and parameterized in such a way that they can be compared across species. Second, it will be necessary to develop comparative phylogenetic methods suitable for large multidimensional datasets. In most phylogenetic comparative studies to date, the number n of independent observations (independent contrasts) has been greater than the number p of variables (characters). The behavior of comparative methods for these classic problems is now well understood under a wide variety of conditions. In studies of gene expression, and in studies based on other high-throughput tools, the number n of samples is dwarfed by the number p of variables. The estimated covariance matrices will be singular, complicating their analysis and interpretation, and prone to spurious results. Third, new approaches are needed to investigate the expression of the many genes whose phylogenies are not congruent with species phylogenies due to gene loss, gene duplication, and incomplete lineage sorting. Here we outline general considerations of project design for phylogenetic analyses of gene expression and suggest solutions to these three categories of challenges. These topics are relevant to high-throughput phenotypic data well beyond gene expression.

  6. Resistência à Síndrome Ascítica, Competência Homeotérmica e Níveis de Hsp70 no Coração e Pulmão de Frangos de Corte

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    Hernandes Renata

    2002-01-01

    Full Text Available Como em outros seres vivos, também nas células das aves ocorre a síntese das proteínas de baixo peso molecular (Hsp, cujo aumento é induzido sob condições de estresse. As Hsps têm um papel importante na manutenção da integridade celular, questiona-se o seu envolvimento no mecanismo de proteção celular de órgãos alvos na ocorrência da síndrome ascítica (SA. Este trabalho objetivou avaliar a temperatura corporal e os níveis da Hsp70 no coração e pulmão de frangos de corte Hubbard (sensível à SA e caipira de pescoço-pelado (resistente, criados em termoneutralidade (25degreesC e frio (16degreesC entre 10 e 45 dias de idade. Foram utilizados 192 pintos machos, 96 de cada linhagem. Não houve mortalidade por SA nas aves caipiras. Nas aves Hubbard, a mortalidade devida à SA foi de 4% e 41% em ambiente termoneutro e frio, respectivamente. Em ambiente frio, a temperatura corporal das aves Hubbard foi menor que a das caipiras. A temperatura corporal e o nível de Hsp70 do coração das aves Hubbard diminuíram com o aumento da idade, mas não nas aves caipiras, os quais se mantiveram constantes, inclusive a Hsp70 do pulmão. Independente da idade ou da temperatura, o nível de Hsp70 no pulmão das aves caipiras era superior ao das aves Hubbard. Em relação às aves Hubbard, as caipiras são homeotérmicas mais competentes e apresentam uma maior indução de Hsp70 nos órgãos primariamente afetados na SA, mas este não parece ser o sistema de proteção contra SA, a qual as aves de pescoço pelado são resistentes.

  7. The Specialized Hsp70 (HscA) Interdomain Linker Binds to Its Nucleotide-Binding Domain and Stimulates ATP Hydrolysis in Both cis and trans Configurations

    Science.gov (United States)

    2015-01-01

    Proteins from the isc operon of Escherichia coli constitute the machinery used to synthesize iron–sulfur (Fe–S) clusters for delivery to recipient apoproteins. Efficient and rapid [2Fe-2S] cluster transfer from the holo-scaffold protein IscU depends on ATP hydrolysis in the nucleotide-binding domain (NBD) of HscA, a specialized Hsp70-type molecular chaperone with low intrinsic ATPase activity (0.02 min−1 at 25 °C, henceforth reported in units of min–1). HscB, an Hsp40-type cochaperone, binds to HscA and stimulates ATP hydrolysis to promote cluster transfer, yet while the interactions between HscA and HscB have been investigated, the role of HscA’s interdomain linker in modulating ATPase activity has not been explored. To address this issue, we created three variants of the 40 kDa NBD of HscA: NBD alone (HscA386), NBD with a partial linker (HscA389), and NBD with the full linker (HscA395). We found that the rate of ATP hydrolysis of HscA395 (0.45 min–1) is nearly 15-fold higher than that of HscA386 (0.035 min–1), although their apparent affinities for ATP are equivalent. HscA395, which contains the full covalently linked linker peptide, exhibited intrinsic tryptophan fluorescence emission and basal thermostability that were higher than those of HscA386. Furthermore, HscA395 displayed narrower 1HN line widths in its two-dimensional 1H–15N TROSY-HSQC spectrum in comparison to HscA386, indicating that the peptide in the cis configuration binds to and stabilizes the structure of the NBD. The addition to HscA386 of a synthetic peptide with a sequence identical to that of the interdomain linker (L387LLDVIPLS395) stimulated its ATPase activity and induced widespread NMR chemical shift perturbations indicative of a binding interaction in the trans configuration. PMID:25372495

  8. Idiomatic (gene) expressions.

    Science.gov (United States)

    Rockman, Matthew V

    2003-05-01

    Hidden among the myriad nucleotide variants that constitute each species' gene pool are a few variants that contribute to phenotypic variation. Many of these differences that make a difference are non-coding cis-regulatory variants, which, unlike coding variants, can only be identified through laborious experimental analysis. Recently, Cowles et al.1 described a screening method that does an end-run around this problem by searching for genes whose cis regulation varies without having to find the polymorphic nucleotides that influence transcription. While we will continue to require a diverse arsenal of experimental methods, this versatile method will speed the identification of functional genetic variation. Copyright 2003 Wiley Periodicals, Inc.

  9. Freezing of body fluids induces metallothionein gene expression in earthworms (Dendrobaena octaedra).

    Science.gov (United States)

    Fisker, Karina Vincents; Holmstrup, Martin; Sørensen, Jesper Givskov

    2016-01-01

    The molecular mechanisms activated by environmental contaminants and natural stressors such as freezing need to be investigated in order to better understand the mechanisms of interaction and potential effects that combined stressors may have on organisms. Using the freeze-tolerant earthworm Dendrobaena octaedra as model species, we exposed worms to freezing and exposure to sublethal copper in a factorial design and investigated the transcription of candidate genes for metal and cold stress. We hypothesised that both freezing and copper would induce transcription of genes coding for heat shock proteins (hsp10 and hsp70), metallothioneins (mt1 and mt2), and glutathione-S-transferase (gst), and that the combined effects of these two stressors would be additive. The gene transcripts hsp10, hsp70, and gst were significantly upregulated by freezing, but only hsp10 was upregulated by copper. We found that copper at the time of sampling had no effect on transcription of two metallothionein genes whereas transcription was strongly upregulated by freezing. Moreover, there was a significant interaction causing more than additive transcription rates of mt1 in the copper/freezing treatment suggesting that freeze-induced cellular dehydration increases the concentration of free copper ions in the cytosol. This metallothionein response to freezing is likely adaptive and possibly provides protection against freeze-induced elevated metal concentrations in the cytosol and excess ROS levels due to hypoxia during freezing. Copyright © 2015 Elsevier Inc. All rights reserved.

  10. 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

  11. Correction of gene expression data

    DEFF Research Database (Denmark)

    Darbani Shirvanehdeh, Behrooz; Stewart, C. Neal, Jr.; Noeparvar, Shahin

    2014-01-01

    This report investigates for the first time the potential inter-treatment bias source of cell number for gene expression studies. Cell-number bias can affect gene expression analysis when comparing samples with unequal total cellular RNA content or with different RNA extraction efficiencies....... For maximal reliability of analysis, therefore, comparisons should be performed at the cellular level. This could be accomplished using an appropriate correction method that can detect and remove the inter-treatment bias for cell-number. Based on inter-treatment variations of reference genes, we introduce...

  12. Sleep deprivation and gene expression.

    Science.gov (United States)

    da Costa Souza, Annie; Ribeiro, Sidarta

    2015-01-01

    Sleep occurs in a wide range of animal species as a vital process for the maintenance of homeostasis, metabolic restoration, physiological regulation, and adaptive cognitive functions in the central nervous system. Long-term perturbations induced by the lack of sleep are mostly mediated by changes at the level of transcription and translation. This chapter reviews studies in humans, rodents, and flies to address the various ways by which sleep deprivation affects gene expression in the nervous system, with a focus on genes related to neuronal plasticity, brain function, and cognition. However, the effects of sleep deprivation on gene expression and the functional consequences of sleep loss are clearly not restricted to the cognitive domain but may include increased inflammation, expression of stress-related genes, general impairment of protein translation, metabolic imbalance, and thermal deregulation.

  13. Pogostick: a new versatile piggyBac vector for inducible gene over-expression and down-regulation in emerging model systems.

    Directory of Open Access Journals (Sweden)

    Bin Chen

    2011-04-01

    Full Text Available Non-traditional model systems need new tools that will enable them to enter the field of functional genetics. These tools should enable the exploration of gene function, via knock-downs of endogenous genes, as well as over-expression and ectopic expression of transgenes.We constructed a new vector called Pogostick that can be used to over-express or down-regulate genes in organisms amenable to germ line transformation by the piggyBac transposable element. Pogostick can be found at www.addgene.org, a non-profit plasmid repository. The vector currently uses the heat-shock promoter Hsp70 from Drosophila to drive transgene expression and, as such, will have immediate applicability to organisms that can correctly interpret this promotor sequence. We detail how to clone candidate genes into this vector and test its functionality in Drosophila by targeting a gene coding for the fluorescent protein DsRed. By cloning a single DsRed copy into the vector, and generating transgenic lines, we show that DsRed mRNA and protein levels are elevated following heat-shock. When cloning a second copy of DsRed in reverse orientation into a flanking site, and transforming flies constitutively expressing DsRed in the eyes, we show that endogenous mRNA and protein levels drop following heat-shock. We then test the over-expression vector, containing the complete cDNA of Ultrabithorax (Ubx gene, in an emerging model system, Bicyclus anynana. We produce a transgenic line and show that levels of Ubx mRNA expression rise significantly following a heat-shock. Finally, we show how to obtain genomic sequence adjacent to the Pogostick insertion site and to estimate transgene copy number in genomes of transformed individuals.This new vector will allow emerging model systems to enter the field of functional genetics with few hurdles.

  14. ASPM gene expression in medulloblastoma.

    Science.gov (United States)

    Vulcani-Freitas, Tânia M; Saba-Silva, Najsla; Cappellano, Andréa; Cavalheiro, Sérgio; Marie, Sueli K N; Oba-Shinjo, Sueli M; Malheiros, Suzana M F; de Toledo, Sílvia Regina Caminada

    2011-01-01

    Medulloblastomas are the most common malignant tumors of the central nervous system in childhood. The incidence is about 19-20% between children younger than 16 years old with peak incidence between 4 and 7 years. Despite its sensibility to no specific therapeutic means like chemotherapy and radiotherapy, the treatment is very aggressive and frequently results in regression, growth deficit, and endocrine dysfunction. From this point of view, new treatment approaches are needed such as molecular targeted therapies. Studies in glioblastoma demonstrated that ASPM gene was overexpressed when compared to normal brain and ASPM inhibition by siRNA-mediated inhibits tumor cell proliferation and neural stem cell proliferation, supporting ASPM gene as a potential molecular target in glioblastoma. The aim of this work was to evaluate ASPM expression in medulloblastoma fragment samples, and to compare the results with the patient clinical features. Analysis of gene expression was performed by quantitative PCR real time using SYBR Green system in tumor samples from 37 children. The t test was used to analyze the gene expression, and Mann-Whitney test was performed to analyze the relationship between gene expressions and clinical characteristics. Kaplan-Meier test evaluated curve survival. All samples overexpressed ASPM gene more than 40-fold. However, we did not find any association between the overexpressed samples and the clinical parameters. ASPM overexpression may modify the ability of stem cells to differentiate during the development of the central nervous system, contributing to the development of medulloblastoma, a tumor of embryonic origin from cerebellar progenitor cells.

  15. Gene expression based cancer classification

    OpenAIRE

    Sara Tarek; Reda Abd Elwahab; Mahmoud Shoman

    2017-01-01

    Cancer classification based on molecular level investigation has gained the interest of researches as it provides a systematic, accurate and objective diagnosis for different cancer types. Several recent researches have been studying the problem of cancer classification using data mining methods, machine learning algorithms and statistical methods to reach an efficient analysis for gene expression profiles. Studying the characteristics of thousands of genes simultaneously offered a deep in...

  16. Transgenic Arabidopsis Gene Expression System

    Science.gov (United States)

    Ferl, Robert; Paul, Anna-Lisa

    2009-01-01

    The Transgenic Arabidopsis Gene Expression System (TAGES) investigation is one in a pair of investigations that use the Advanced Biological Research System (ABRS) facility. TAGES uses Arabidopsis thaliana, thale cress, with sensor promoter-reporter gene constructs that render the plants as biomonitors (an organism used to determine the quality of the surrounding environment) of their environment using real-time nondestructive Green Fluorescent Protein (GFP) imagery and traditional postflight analyses.

  17. Gene expression in colorectal cancer

    DEFF Research Database (Denmark)

    Birkenkamp-Demtroder, Karin; Christensen, Lise Lotte; Olesen, Sanne Harder

    2002-01-01

    Understanding molecular alterations in colorectal cancer (CRC) is needed to define new biomarkers and treatment targets. We used oligonucleotide microarrays to monitor gene expression of about 6,800 known genes and 35,000 expressed sequence tags (ESTs) on five pools (four to six samples in each......%). Fifteen nuclear encoded mitochondrial proteins were all down-regulated in CRC. We identified several chromosomal locations with clusters of either potential oncogenes or potential tumor suppressors. Some of these, such as aminopeptidase N/CD13 and sigma B3 protein on chromosome 15q25, coincided...

  18. Inhibition of Salmonella-induced IL-8 synthesis and expression of Hsp70 in enterocyte-like Caco-2 cells after exposure to non-starter lactobacilli

    NARCIS (Netherlands)

    Nemeth, Edina; Fajdiga, Sana; Malago, Joshua; Koninkx, Jos; Tooten, Peter; van Dijk, Jaap

    2006-01-01

    Oral administration of lactobacilli as probiotics is gaining importance in the treatment of intestinal inflammations. We investigated the effect of non-starter lactobacilli Lactobacillus casei subsp casei 2756, Lactobacillus curvatus 2775, and Lactobacillus plantarum 2142 as well as their spent

  19. Heat Shock Protein 70 Expression is Increased in the Liver of Neonatal Intrauterine Growth Retardation Piglets

    Directory of Open Access Journals (Sweden)

    Wei Li

    2012-08-01

    Full Text Available Intrauterine growth retardation (IUGR leads to the dysfunction in digestive system, as well as the alteration in the expression of some functional proteins. Heat shock protein 70 (Hsp70 could be induced by various stress factors, but whether Hsp70 expression is changed in neonatal IUGR infants has not been demonstrated. This study was conducted to explore the expression of Hsp70 in the liver by using the IUGR piglet model. Liver and plasma samples were obtained from IUGR and normal birth weight (NBW piglets at birth. The neonatal IUGR piglets had significantly lower liver weight than their counterparts. The activities of aspartate aminotransferase and alanine aminotransferase in serum were enhanced significantly in IUGR indicating liver dysfunction. The activities of superoxide dismutase (p0.05 were lower and the level of malondialdehybe was higher (p<0.05 in IUGR liver compared with in NBW. According to the results of histological tests, fatty hepatic infiltrates and cytoplasmic vacuolization were present in the liver of IUGR piglets, but not in NBW liver. The expression of Hsp70 protein was significantly higher (p<0.05 in IUGR piglet liver than in NBW. Similar to where the hepatic injuries were observed, location of Hsp70 was mostly in the midzonal hepatic lobule indicating that oxidative stress might be responsible for the increased expression of Hsp70.

  20. Heat Shock Protein 70 Expression is Increased in the Liver of Neonatal Intrauterine Growth Retardation Piglets.

    Science.gov (United States)

    Li, Wei; Zhong, Xiang; Zhang, Lili; Wang, Yuanxiao; Wang, Tian

    2012-08-01

    Intrauterine growth retardation (IUGR) leads to the dysfunction in digestive system, as well as the alteration in the expression of some functional proteins. Heat shock protein 70 (Hsp70) could be induced by various stress factors, but whether Hsp70 expression is changed in neonatal IUGR infants has not been demonstrated. This study was conducted to explore the expression of Hsp70 in the liver by using the IUGR piglet model. Liver and plasma samples were obtained from IUGR and normal birth weight (NBW) piglets at birth. The neonatal IUGR piglets had significantly lower liver weight than their counterparts. The activities of aspartate aminotransferase and alanine aminotransferase in serum were enhanced significantly in IUGR indicating liver dysfunction. The activities of superoxide dismutase (p0.05) were lower and the level of malondialdehybe was higher (ppiglets, but not in NBW liver. The expression of Hsp70 protein was significantly higher (ppiglet liver than in NBW. Similar to where the hepatic injuries were observed, location of Hsp70 was mostly in the midzonal hepatic lobule indicating that oxidative stress might be responsible for the increased expression of Hsp70.

  1. Heat Shock Protein 70 Expression is Increased in the Liver of Neonatal Intrauterine Growth Retardation Piglets

    Science.gov (United States)

    Li, Wei; Zhong, Xiang; Zhang, Lili; Wang, Yuanxiao; Wang, Tian

    2012-01-01

    Intrauterine growth retardation (IUGR) leads to the dysfunction in digestive system, as well as the alteration in the expression of some functional proteins. Heat shock protein 70 (Hsp70) could be induced by various stress factors, but whether Hsp70 expression is changed in neonatal IUGR infants has not been demonstrated. This study was conducted to explore the expression of Hsp70 in the liver by using the IUGR piglet model. Liver and plasma samples were obtained from IUGR and normal birth weight (NBW) piglets at birth. The neonatal IUGR piglets had significantly lower liver weight than their counterparts. The activities of aspartate aminotransferase and alanine aminotransferase in serum were enhanced significantly in IUGR indicating liver dysfunction. The activities of superoxide dismutase (p0.05) were lower and the level of malondialdehybe was higher (ppiglets, but not in NBW liver. The expression of Hsp70 protein was significantly higher (ppiglet liver than in NBW. Similar to where the hepatic injuries were observed, location of Hsp70 was mostly in the midzonal hepatic lobule indicating that oxidative stress might be responsible for the increased expression of Hsp70. PMID:25049668

  2. Sequence characterization of heat shock protein gene of Cyclospora cayetanensis isolates from Nepal, Mexico, and Peru.

    Science.gov (United States)

    Sulaiman, Irshad M; Torres, Patricia; Simpson, Steven; Kerdahi, Khalil; Ortega, Ynes

    2013-04-01

    We have described the development of a 2-step nested PCR protocol based on the characterization of the 70-kDa heat shock protein (HSP70) gene for rapid detection of the human-pathogenic Cyclospora cayetanensis parasite. We tested and validated these newly designed primer sets by PCR amplification followed by nucleotide sequencing of PCR-amplified HSP70 fragments belonging to 16 human C. cayetanensis isolates from 3 different endemic regions that include Nepal, Mexico, and Peru. No genetic polymorphism was observed among the isolates at the characterized regions of the HSP70 locus. This newly developed HSP70 gene-based nested PCR protocol provides another useful genetic marker for the rapid detection of C. cayetanensis in the future.

  3. Gene Expression Analysis of Breast Cancer Progression

    National Research Council Canada - National Science Library

    Gerald, Wiliam L

    2004-01-01

    ... to identify genes, gene expression profiles and molecular pathways associated with metastatic BC we have performed genome-wide gene expression analysis of a large number of breast cancer samples...

  4. Exposure of the gilthead seabream (Sparus aurata to sediments contaminated with heavy metals down-regulates the gene expression of stress biomarkers

    Directory of Open Access Journals (Sweden)

    Said Benhamed

    2016-01-01

    Full Text Available Heavy metals incidence in the aquatic environment and its accumulation in fish are under constant review. Gilthead seabream (Sparus aurata specimens were exposed for two weeks to sediments highly concentrated in metals, collected at the Portman Bay (Murcia, Spain. The metals bioaccumulation was tested in liver, muscle and skin. The potential of the sediment exposure to induce variation of the stress biomarkers genes was conducted in liver and skin. Results revealed that sediments were highly contaminated with metals. However, following 2 weeks exposure to the sediments, Cd accumulates only in liver. Interestingly, the expression of the genes mta, hsp 70 and hsp 90 were significantly down-regulated in skin. Nevertheless, cyp1a1 gene was up-regulated only in liver. Results uphold that the stress response magnitude was organ-dependent and the skin was the most responsive tissue to metal stress conditions. These results suggest that skin should be considered as target organ for biomarkers analysis in fishes.

  5. [Searching Radiation Countermeasures using the Model of Prolonged Irradiation of Mice with Low Dose Rate and Evaluation of Their Influence on Heat Shock Protein Genes Expression].

    Science.gov (United States)

    Rozhdestvensky, L M; Mikhailov, V F; Schlyakova, T G; Shagirova, J M; Shchegoleva, R A; Raeva, N F; Lisina, N I; Shulenina, L V; Zorin, V V; Pchelka, A V; Trubitsina, K Y

    2015-01-01

    Different radiomodificators (cytokine betaleukine, antioxidant phenoxan, antigipoksant limontar and nucleoside riboxin) were investigated on mice for evaluating their radiation protective capacity against prolonged (21 h) exposure at a dose of 12.6 Gy at a low dose rate of 10 mGy/min. Bone marrow cellularity and endogenic CFUs were used as evaluation criteria 9 days after exposure. Simultaneously, expression of the heat shock proteins of 25, 70 and 90 kDa in unexposed mice bone marrow was studied 2, 24 and 48 h after injections. Betaleukine only had a positive significant effect in both tests in the variants of 50 mcg/kg and 3 mcg/kg when administered 2 h and 22 h before exposure, correspondingly. Effects of betaleukine HSPs on expression were both stimulating and inhibiting, that was in contradiction with a constant positive effect in 5 experiments on exposed mice for each betaleukine variant. It argues against the vital role of HSPs in the betaleukine antiradiation effect. In 2 experiments with high temperatures betaleukine administered at a dose of 50 mcg/kg evoked a very high HSP-70 gene expression after 24 h, and mice exposed to irradiation at that time in a parallel experiment showed an increased radiation effect. It corresponds to the idea that HSPs serve a stress indicator.

  6. Vascular Gene Expression: A Hypothesis

    Directory of Open Access Journals (Sweden)

    Angélica Concepción eMartínez-Navarro

    2013-07-01

    Full Text Available The phloem is the conduit through which photoassimilates are distributed from autotrophic to heterotrophic tissues and is involved in the distribution of signaling molecules that coordinate plant growth and responses to the environment. Phloem function depends on the coordinate expression of a large array of genes. We have previously identified conserved motifs in upstream regions of the Arabidopsis genes, encoding the homologs of pumpkin phloem sap mRNAs, displaying expression in vascular tissues. This tissue-specific expression in Arabidopsis is predicted by the overrepresentation of GA/CT-rich motifs in gene promoters. In this work we have searched for common motifs in upstream regions of the homologous genes from plants considered to possess a primitive vascular tissue (a lycophyte, as well as from others that lack a true vascular tissue (a bryophyte, and finally from chlorophytes. Both lycophyte and bryophyte display motifs similar to those found in Arabidopsis with a significantly low E-value, while the chlorophytes showed either a different conserved motif or no conserved motif at all. These results suggest that these same genes are expressed coordinately in non- vascular plants; this coordinate expression may have been one of the prerequisites for the development of conducting tissues in plants. We have also analyzed the phylogeny of conserved proteins that may be involved in phloem function and development. The presence of CmPP16, APL, FT and YDA in chlorophytes suggests the recruitment of ancient regulatory networks for the development of the vascular tissue during evolution while OPS is a novel protein specific to vascular plants.

  7. Variant evidence of three genes of Potato vein yellow virus infecting Solanum tubersosum sp. using single strand conformational polymorphism.

    Directory of Open Access Journals (Sweden)

    Karen Andrea CUBILLOS ABELLO

    2014-10-01

    Full Text Available 1024x768 Potato yellow vein virus (PYVV with tripartite RNA (ss+ genome is classified as Crinivirus within the family Closteoviridae. PYVV is the causal agent of potato yellow vein disease (PYVD with yield effect. Single strand conformational polymorphism (SSCP was reported to estimate variability in eukaryotes and different viruses’ species. In this study, the molecular variability of PYVV was analyzed by SSCP patterns regarding three genes: major capsid protein (CP, minor capsid protein (CPm, and heat shock protein (Hsp70 obtained from isolates of 60 potato plants leaves expressing PYVD. Leaves were collected in Nariño, Colombia from 30 Solanum tuberosum, Phureja Group (PhG and 30 Andigena Group (AG. Genes were amplified by RT-PCR and the purified direct PCR products were used for SSCP. Three SSCP patterns were detected for CP gen, 12 for CPm and 12 for Hsp70 genes. The AG host, most frequent pattern was C (66.6% for the Hsp70 gene and IV (33.3% for CPm gene. Complex SSCP patters with inter and intra variation were detected. CP gene pattern 1 was present in 93.3% of the isolates of both host groups, indicating low number of variants compared to CPm and Hsp70 genes. This is the first attempt to estimate intra e inter PYVV variability by the use of a simple molecular method considering three genes in a large group of potato samples affected by PYVD. SSCP technique was useful to discriminate the migration patterns generated for each gene Normal 0 false false false EN-US X-NONE X-NONE /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Tabla normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-qformat:yes; mso-style-parent:""; mso-padding-alt:0in 5.4pt 0in 5.4pt; mso-para-margin:0in; mso-para-margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:11.0pt; font-family:"Calibri","sans-serif"; mso-ascii-font-family:Calibri; mso-ascii-theme-font:minor-latin; mso

  8. Gene expressions and metabolomic research on the effects of polyphenols from the involucres of Castanea mollissima Blume on heat-stressed broilers chicks.

    Science.gov (United States)

    Xiong, Y; Dong, S; Zhao, X; Guo, K J; Gasco, Laura; Zoccarato, Ivo

    2016-08-01

    To study the effects of polyphenolic extract from involucres of Castanea mollissima Blume ( PICB: ), a novel approach using gene expression by real time polymerase chain reaction ( REAL-TIME PCR: ) coupled with metabolomic profiling technique was established to explain the mechanism of PICB on heat-stressed broiler chicks. Four thousand 28-day-old male Arbor Acres (AA) broilers were randomly assigned to 5 groups (4 replicates / group, 20 chicks / replicate), in which group 1 was normal control group fed with basic ration; groups 2, 3, 4, and 5 were fed with the basic ration with a supplementation of 0.2% Vitamin C ( VC: ), or 0.2%, 0.3%, or 0.4% of PICB respectively. After 1 wk of adaptation, heat stress was applied for 7 consecutive days. On d 3 and d 7 of heat stress, the chicks were sacrificed and sampled. The mRNA expression of heat stress protein 70 (HSP70), glutathione peroxidase ( GSH-PX: ), ornithine decarboxylase ( ODC: ), epidermal growth factor ( EGF: ) and epidermal growth factor receptor ( EGFR: ) were detected by real-time PCR using samples from jejunum mucosa. The serum and jejunum mucosa metabolomic profiles of PICB group showing best antioxidative effects and control group at d 3 were studied using the method of the gas chromatography - time of flight mass spectrometry ( GT-TOF-MS: ), followed by principal component analysis and partial least squares-discriminate analysis. Potential biomarkers were found using Student's t-test. The results showed mRNA expressions of HSP70, GSH-Px, ODC, EGF, and EGFR were altered by the supplementation of PICB. PICB exhibited antioxidative and growth promoting effects, and 0.3% PICB supplementation level exhibited the best. Three metabolites in the serum and 5 in the jejunum mucosa were identified as potential biomarkers. They were considered to be in accordance with antioxidative and growth promoting effects of PICB, which involved in the energy metabolism (sorbitol, palmitic acid), carbohydrate metabolism, amino

  9. Neighboring Genes Show Correlated Evolution in Gene Expression

    Science.gov (United States)

    Ghanbarian, Avazeh T.; Hurst, Laurence D.

    2015-01-01

    When considering the evolution of a gene’s expression profile, we commonly assume that this is unaffected by its genomic neighborhood. This is, however, in contrast to what we know about the lack of autonomy between neighboring genes in gene expression profiles in extant taxa. Indeed, in all eukaryotic genomes genes of similar expression-profile tend to cluster, reflecting chromatin level dynamics. Does it follow that if a gene increases expression in a particular lineage then the genomic neighbors will also increase in their expression or is gene expression evolution autonomous? To address this here we consider evolution of human gene expression since the human-chimp common ancestor, allowing for both variation in estimation of current expression level and error in Bayesian estimation of the ancestral state. We find that in all tissues and both sexes, the change in gene expression of a focal gene on average predicts the change in gene expression of neighbors. The effect is highly pronounced in the immediate vicinity (genes increasing their expression in humans tend to avoid nuclear lamina domains and be enriched for the gene activator 5-hydroxymethylcytosine, we conclude that, most probably owing to chromatin level control of gene expression, a change in gene expression of one gene likely affects the expression evolution of neighbors, what we term expression piggybacking, an analog of hitchhiking. PMID:25743543

  10. Gene expression profile of pulpitis

    Science.gov (United States)

    Galicia, Johnah C.; Henson, Brett R.; Parker, Joel S.; Khan, Asma A.

    2016-01-01

    The cost, prevalence and pain associated with endodontic disease necessitate an understanding of the fundamental molecular aspects of its pathogenesis. This study was aimed to identify the genetic contributors to pulpal pain and inflammation. Inflamed pulps were collected from patients diagnosed with irreversible pulpitis (n=20). Normal pulps from teeth extracted for various reasons served as controls (n=20). Pain level was assessed using a visual analog scale (VAS). Genome-wide microarray analysis was performed using Affymetrix GeneTitan Multichannel Instrument. The difference in gene expression levels were determined by the Significance Analysis of Microarray program using a false discovery rate (q-value) of 5%. Genes involved in immune response, cytokine-cytokine receptor interaction and signaling, integrin cell surface interactions, and others were expressed at relatively higher levels in the in the pulpitis group. Moreover, several genes known to modulate pain and inflammation showed differential expression in asymptomatic and mild pain patients (≥30mm on VAS) compared to those with moderate to severe pain. This exploratory study provides a molecular basis for the clinical diagnosis of pulpitis. With an enhanced understanding of pulpal inflammation, future studies on treatment and management of pulpitis and on pain associated with it can have a biological reference to bridge treatment strategies with pulpal biology. PMID:27052691

  11. Gene Expression in Trypanosomatid Parasites

    Directory of Open Access Journals (Sweden)

    Santiago Martínez-Calvillo

    2010-01-01

    Full Text Available The parasites Leishmania spp., Trypanosoma brucei, and Trypanosoma cruzi are the trypanosomatid protozoa that cause the deadly human diseases leishmaniasis, African sleeping sickness, and Chagas disease, respectively. These organisms possess unique mechanisms for gene expression such as constitutive polycistronic transcription of protein-coding genes and trans-splicing. Little is known about either the DNA sequences or the proteins that are involved in the initiation and termination of transcription in trypanosomatids. In silico analyses of the genome databases of these parasites led to the identification of a small number of proteins involved in gene expression. However, functional studies have revealed that trypanosomatids have more general transcription factors than originally estimated. Many posttranslational histone modifications, histone variants, and chromatin modifying enzymes have been identified in trypanosomatids, and recent genome-wide studies showed that epigenetic regulation might play a very important role in gene expression in this group of parasites. Here, we review and comment on the most recent findings related to transcription initiation and termination in trypanosomatid protozoa.

  12. Gene expression profile of pulpitis.

    Science.gov (United States)

    Galicia, J C; Henson, B R; Parker, J S; Khan, A A

    2016-06-01

    The cost, prevalence and pain associated with endodontic disease necessitate an understanding of the fundamental molecular aspects of its pathogenesis. This study was aimed to identify the genetic contributors to pulpal pain and inflammation. Inflamed pulps were collected from patients diagnosed with irreversible pulpitis (n=20). Normal pulps from teeth extracted for various reasons served as controls (n=20). Pain level was assessed using a visual analog scale (VAS). Genome-wide microarray analysis was performed using Affymetrix GeneTitan Multichannel Instrument. The difference in gene expression levels were determined by the significance analysis of microarray program using a false discovery rate (q-value) of 5%. Genes involved in immune response, cytokine-cytokine receptor interaction and signaling, integrin cell surface interactions, and others were expressed at relatively higher levels in the pulpitis group. Moreover, several genes known to modulate pain and inflammation showed differential expression in asymptomatic and mild pain patients (⩾30 mm on VAS) compared with those with moderate to severe pain. This exploratory study provides a molecular basis for the clinical diagnosis of pulpitis. With an enhanced understanding of pulpal inflammation, future studies on treatment and management of pulpitis and on pain associated with it can have a biological reference to bridge treatment strategies with pulpal biology.

  13. Involvement of NF-κB and HSP70 signaling pathways in the apoptosis of MDA-MB-231 cells induced by a prenylated xanthone compound, α-mangostin, from Cratoxylum arborescens [Retraction

    Directory of Open Access Journals (Sweden)

    Ibrahim MY

    2017-05-01

    Full Text Available Ibrahim MY, Hashim NM, Mohan S, et al. Involvement of NF-κB and HSP70 signaling pathways in the apoptosis of MDA-MB-231 cells induced by a prenylated xanthone compound, α-mangostin, from Cratoxylum arborescens. Drug Design, Development and Therapy. 2014;8:2193–2211 was published subsequent to Ibrahim MY, Hashim NM, Mohan S, et al. α-Mangostin from Cratoxylum arborescens demonstrates apoptogenesis in MCF-7 with regulation of NF-κB and Hsp70 protein modulation in vitro, and tumor reduction in vivo. Drug Design, Development and Therapy. 2014;8:1629–1647.When comparing the papers it becomes apparent that they have an unacceptably high degree of similarity and re-use. Further, there is no clear scientific distinction between the cell lines and the results in both. Accordingly, the Editor-in-Chief and Publisher have issued this Notice of Retraction.This retraction relates to this paperThis retraction relates to this Corrigendum

  14. In Silico Molecular Modeling and Docking Studies on Novel Mutants (E229V, H225P and D230C) of the Nucleotide-Binding Domain of Homo sapiens Hsp70.

    Science.gov (United States)

    Elengoe, Asita; Hamdan, Salehhuddin

    2017-12-01

    In this study, we explored the possibility of determining the synergistic interactions between nucleotide-binding domain (NBD) of Homo sapiens heat-shock 70 kDa protein (Hsp70) and E1A 32 kDa of adenovirus serotype 5 motif (PNLVP) in the efficiency of killing of tumor cells in cancer treatment. At present, the protein interaction between NBD and PNLVP motif is still unknown, but believed to enhance the rate of virus replication in tumor cells. Three mutant models (E229V, H225P and D230C) were built and simulated, and their interactions with PNLVP motif were studied. The PNLVP motif showed the binding energy and intermolecular energy values with the novel E229V mutant at -7.32 and -11.2 kcal/mol. The E229V mutant had the highest number of hydrogen bonds (7). Based on the root mean square deviation, root mean square fluctuation, hydrogen bonds, salt bridge, secondary structure, surface-accessible solvent area, potential energy and distance matrices analyses, it was proved that the E229V had the strongest and most stable interaction with the PNLVP motif among all the four protein-ligand complex structures. The knowledge of this protein-ligand complex model would help in designing Hsp70 structure-based drug for cancer therapy.

  15. Systems Biophysics of Gene Expression

    Science.gov (United States)

    Vilar, Jose M.G.; Saiz, Leonor

    2013-01-01

    Gene expression is a process central to any form of life. It involves multiple temporal and functional scales that extend from specific protein-DNA interactions to the coordinated regulation of multiple genes in response to intracellular and extracellular changes. This diversity in scales poses fundamental challenges to the use of traditional approaches to fully understand even the simplest gene expression systems. Recent advances in computational systems biophysics have provided promising avenues to reliably integrate the molecular detail of biophysical process into the system behavior. Here, we review rece