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Sample records for citrus hsp70 molecular

  1. 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.

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

    OpenAIRE

    Fietto, Luciano G.; Maximiller D.L. Costa; Cosme D Cruz; De Souza, Alessandra A.; Machado, Marcos A; Fontes, Elizabeth P. B.

    2007-01-01

    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 enc...

  3. The nucleotide exchange factors of Hsp70 molecular chaperone

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    Andreas eBracher

    2015-04-01

    Full Text Available Molecular chaperones of the Hsp70 family form an important hub in the cellular protein folding networks in bacteria and eukaryotes, connecting translation with the downstream machineries of protein folding and degradation. The Hsp70 folding cycle is driven by two types of cochaperones: J-domain proteins stimulate ATP hydrolysis by Hsp70, while nucleotide exchange factors (NEFs promote replacement of Hsp70-bound ADP with ATP. Bacteria and organelles of bacterial origin have only one known NEF type for Hsp70, GrpE. In contrast, a large diversity of Hsp70 NEFs has been discovered in the eukaryotic cell. These NEFs belong to the Hsp110/Grp170, HspBP1/Sil1 and BAG domain protein families. In this short review we compare the structures and molecular mechanisms of nucleotide exchange factors for Hsp70 and discuss how these cochaperones contribute to protein folding and quality control in the cell.

  4. The assembly and intermolecular properties of the Hsp70-Tomm34-Hsp90 molecular chaperone complex.

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    Trcka, Filip; Durech, Michal; Man, Petr; Hernychova, Lenka; Muller, Petr; Vojtesek, Borivoj

    2014-04-01

    Maintenance of protein homeostasis by molecular chaperones Hsp70 and Hsp90 requires their spatial and functional coordination. The cooperation of Hsp70 and Hsp90 is influenced by their interaction with the network of co-chaperone proteins, some of which contain tetratricopeptide repeat (TPR) domains. Critical to these interactions are TPR domains that target co-chaperone binding to the EEVD-COOH motif that terminates Hsp70/Hsp90. Recently, the two-TPR domain-containing protein, Tomm34, was reported to bind both Hsp70 and Hsp90. Here we characterize the structural basis of Tomm34-Hsp70/Hsp90 interactions. Using multiple methods, including pull-down assays, fluorescence polarization, hydrogen/deuterium exchange, and site-directed mutagenesis, we defined the binding activities and specificities of Tomm34 TPR domains toward Hsp70 and Hsp90. We found that Tomm34 TPR1 domain specifically binds Hsp70. This interaction is partly mediated by a non-canonical TPR1 two-carboxylate clamp and is strengthened by so far unidentified additional intermolecular contacts. The two-carboxylate clamp of the isolated TPR2 domain has affinity for both chaperones, but as part of the full-length Tomm34 protein, the TPR2 domain binds specifically Hsp90. These binding properties of Tomm34 TPR domains thus enable simultaneous binding of Hsp70 and Hsp90. Importantly, we provide evidence for the existence of an Hsp70-Tomm34-Hsp90 tripartite complex. In addition, we defined the basic conformational demands of the Tomm34-Hsp90 interaction. These results suggest that Tomm34 represents a novel scaffolding co-chaperone of Hsp70 and Hsp90, which may facilitate Hsp70/Hsp90 cooperation during protein folding.

  5. Zinc-L-carnosine binds to molecular chaperone HSP70 and inhibits the chaperone activity of the protein.

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    Haga, Asami; Okamoto, Tomoya; Yamada, Shintaroh; Kubota, Toshihiko; Sanpei, Ann; Takahashi, Shota; Nakayama, Masahiro; Nagai, Miki; Otaka, Michiro; Miyazaki, Toshio; Nunomura, Wataru; Grave, Ewa; Itoh, Hideaki

    2013-09-01

    In this study, we have investigated the specific binding proteins of Zinc-L-carnosine (Polaprezinc) using Polaprezinc-affinity column chromatography in vitro. A protein having a 70-kDa molecular mass was eluted by the linear gradient of 0-1.0 mM Polaprezinc from the affinity column and the protein was identified as the molecular chaperone HSP70 by immunoblotting. The chaperone activity of HSP70 was completely suppressed by Polaprezinc. The ATPase activity of HSP70 was affected to some extent by the reagent. In the circular dichroism (CD) spectrum, the secondary structure of HSP70 was changed in the presence of Polaprezinc, i.e. it decreased in the α-helix. We have determined the Polaprezinc-binding domain of HSP70 by using recombinant HSP70N- and C-domains. Although Polaprezinc could bind to both the N-terminal and the C-terminal of HSP70, the HSP70N-domain has a high affinity to the drug. Regarding the peptide cleavage of the HSP70N- and C-domains with proteinase K, the intact HSP70N still remained in the presence of Polaprezinc. On the other hand, the quantity of the intact C-domain slightly decreased under the same conditions along with the newly digested small peptides appeared. It has been suggested that Polaprezinc binds to HSP70 especially in the N-domains, suppresses the chaperone activity and delays an ATPase activities of HSP70. PMID:23687308

  6. Molecular mechanism of mutant p53 stabilization: the role of HSP70 and MDM2.

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    Milena Wiech

    Full Text Available Numerous p53 missense mutations possess gain-of-function activities. Studies in mouse models have demonstrated that the stabilization of p53 R172H (R175H in human mutant protein, by currently unknown factors, is a prerequisite for its oncogenic gain-of-function phenotype such as tumour progression and metastasis. Here we show that MDM2-dependent ubiquitination and degradation of p53 R175H mutant protein in mouse embryonic fibroblasts is partially inhibited by increasing concentration of heat shock protein 70 (HSP70/HSPA1-A. These phenomena correlate well with the appearance of HSP70-dependent folding intermediates in the form of dynamic cytoplasmic spots containing aggregate-prone p53 R175H and several molecular chaperones. We propose that a transient but recurrent interaction with HSP70 may lead to an increase in mutant p53 protein half-life. In the presence of MDM2 these pseudoaggregates can form stable amyloid-like structures, which occasionally merge into an aggresome. Interestingly, formation of folding intermediates is not observed in the presence of HSC70/HSPA8, the dominant-negative K71S variant of HSP70 or HSP70 inhibitor. In cancer cells, where endogenous HSP70 levels are already elevated, mutant p53 protein forms nuclear aggregates without the addition of exogenous HSP70. Aggregates containing p53 are also visible under conditions where p53 is partially unfolded: 37°C for temperature-sensitive variant p53 V143A and 42°C for wild-type p53. Refolding kinetics of p53 indicate that HSP70 causes transient exposure of p53 aggregate-prone domain(s. We propose that formation of HSP70- and MDM2-dependent protein coaggregates in tumours with high levels of these two proteins could be one of the mechanisms by which mutant p53 is stabilized. Moreover, sequestration of p73 tumour suppressor protein by these nuclear aggregates may lead to gain-of-function phenotypes.

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

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    Ishida, Kenji; Jolly, Emmitt R

    2016-09-01

    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 behavior transitional

  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

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    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.

  9. Thermal tolerance of the crab Pachygrapsus marmoratus: intraspecific differences at a physiological (CTMax) and molecular level (Hsp70).

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    Madeira, D; Narciso, L; Cabral, H N; Diniz, M S; Vinagre, C

    2012-11-01

    Temperature is one of the most important variables influencing organisms, especially in the intertidal zone. This work aimed to test physiological and molecular intraspecific differences in thermal tolerance of the crab Pachygrapsus marmoratus (Fabricius, 1787). The comparisons made focused on sex, size, and habitat (estuary and coast) differences. The physiological parameter was upper thermal limit, tested via the critical thermal maximum (CTMax) and the molecular parameter was total heat shock protein 70 (Hsp70 and Hsp70 plus Hsc70) production, quantified via an enzyme-linked imunosorbent assay. Results showed that CTMax values and Hsp70 production are higher in females probably due to different microhabitat use and potentially due to different hormonal regulation in males and females. Among females, non-reproducing ones showed a higher CTMax value, but no differences were found in Hsp70, even though reproducing females showed higher variability in Hsp70 amounts. As reproduction takes up a lot of energy, its allocation for other activities, including stress responses, is lower. Juveniles also showed higher CTMax and Hsp70 expression because they occur in greater shore heights and ageing leads to alterations in protein synthesis. Comparing estuarine and coastal crabs, no differences were found in CTMax but coastal crabs produce more Hsp70 than estuarine crabs because they occur in drier and hotter areas than estuarine ones, which occur in moister environments. This work shows the importance of addressing intraspecific differences in the stress response at different organizational levels. This study shows that these differences are key factors in stress research, climate research, and environmental monitoring. PMID:22619030

  10. Molecular cloning of HSP70 in Mycoplasma ovipneumoniae and comparison with that of other mycoplasmas.

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    Li, M; Ma, C J; Liu, X M; Zhao, D; Xu, Q C; Wang, Y J

    2011-05-10

    Mycoplasma ovipneumoniae, a bacterial species that specifically affects ovine and goat, is the cause of ovine infectious pleuropneumonia. We cloned, sequenced and analyzed heat shock protein 70 (HSP70) (dnaK) gene of M. ovipneumoniae. The full length open reading frame of the M. ovipneumoniae HSP70 gene consists of 1812 nucleotides, with a G+C content of 34.16%, encoding 604 amino acids. Comparative analysis with the HSP70 sequences of 15 Mycoplasma species revealed 59 to 87% DNA sequence identity, with an amino acid sequence identity range of 58 to 94%. M. ovipneumoniae and M. hyopneumoniae shared the highest DNA and amino acid sequence identity (87 and 94%, respectively). Based on phylogenetic analysis, both the DNA and amino acid identities of M. ovipneumoniae with other mycoplasmal HSP70 were correlated with the degree of relationship between the species. The C-terminus of the HSP70 was cloned into a bacterial expression vector and expressed in Escherichia coli cells. The recombinant C-terminal portion of HSP70 protein strongly reacted with convalescent sera from M. ovipneumoniae-infected sheep, based on an immunoblotting assay. This indicates that HSP70 is immunogenic in a natural M. ovipneumoniae infection and may be a relevant antigen for vaccine development.

  11. Molecular characterization and expression of HSP70, HSF and HSBP genes inOctopus vulgaris during thermal stress

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    HONG Jingni; MAO Yong; NIU Sufang; SUN Tiantian; SU Yongquan

    2015-01-01

    Temperature is an important environmental factor that affects the growth and survival ofOctopus vulgaris, the common octopus. To understand the protective mechanism thatO.vulgaris exhibits under heat stress, we used rapid amplification of cDNA ends (RACE) to obtain full-length sequences of three heat stress response related genes: (1) the heat shock protein 70 (OvHSP70), (2) the heat shock transcription factor (OvHSF) , and (3) the heat shock factor-binding protein (OvHSBP) ofO.vulgaris. The OvHSP70, OvHSF, and OvHSBP proteins contained 2 222 bp, 2 264 bp, 841 bp that encoded for 635, 458 and 90 amino acids, respectively. The results of multiple sequence alignment showed that the amino acid sequences of OvHSP70 were highly conserved with respect to other species. Similarly, the DNA binding domain, the trimerization domain of OvHSF, and the coiled coil region of OvHSBP also had highly conserved regions. The real-time polymerase chain reaction (PCR) results indicated that OvHSP70 was temperature-dependent and time-dependent, showing a positive response to heat stress. On exposure to 28°C and to 30°C, the mRNA expression levels of OvHSF and OvHSBP were higher than those in the control group at 24°C. The mRNA expression of OvHSBP significantly increased with heat treatment at 26°C, while the mRNA expression of OvHSF decreased. The experimental results indicated that the expression of OvHSP70, OvHSF and OvHSBP were all sensitive to heat stress, which suggests that these three genes may play an important role forO.vulgaris in responding to environmental stress. Thus, this study sets a theoretical foundation for further in-depth studies on the molecular protective mechanisms of the heat response inO.vulgaris.

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

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    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.

  13. Molecular Mechanism of Allosteric Communication in Hsp70 Revealed by Molecular Dynamics Simulations

    OpenAIRE

    Chiappori, Federica; Merelli, Ivan; Colombo, Giorgio; Milanesi, Luciano; Morra, Giulia

    2012-01-01

    Author Summary Allostery, or the capability of proteins to respond to ligand binding events with a variation in structure or dynamics at a distant site, is a common feature for biomolecular function and regulation in a large number of proteins. Intra-protein connections and inter-residue coordinations underlie allosteric mechanisms and react to binding primarily through a finely tuned modulation of motions and structures at the microscopic scale. Hence, all-atom molecular dynamics simulations...

  14. Molecular cloning, characterization, and expression analysis of a heat shock protein (HSP) 70 gene from Paphia undulata.

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    Wu, Xiangwei; Tan, Jing; Cai, Mingyi; Liu, Xiande

    2014-06-15

    In this study, a full-length HSP70 cDNA from Paphia undulata was cloned using reverse transcriptase polymerase chain reaction (RT-PCR) coupled with rapid amplification of cDNA ends (RACE). The full-length cDNA is 2,351 bp, consisting of a 5'-untranslated region (UTR) of 83 bp, a 3'-UTR of 315 bp, and an open reading frame (ORF) of 1,953 bp. This cDNA encodes 650 amino acids with an estimated molecular weight of 71.3 kDa and an isoelectric point of 5.51. Based on the amino acid sequence analysis and phylogenetic analysis, this HSP70 gene was identified as a member of the cytoplasmic HSP70 family, being the constitutive expression, and it was designated as PuHSC70. The distribution of PuHSC70 mRNA in the mantle, digestive gland, adductor muscle, gonad, gill, heart, and hemocytes suggested that PuHSC70 is ubiquitously expressed. The mRNA levels of PuHSC70 under high temperature and high salinity stresses were analyzed by real-time PCR. Under high temperature stress of 32°C, PuHSC70 mRNA in the mantle, digestive gland, gill, and heart was significantly up-regulated at 1h and 2h, and it was then progressively down-regulated. In the adductor muscle, the level of PuHSC70 mRNA gradually increased throughout the study period; the mRNA levels in the gonad and hemocytes increased significantly at 4h and 8h (Padductor muscle, hemocytes, and heart were significantly increased, reaching a maximum at 24h, and then they gradually decreased; moreover, in the heart, the mRNA expression recovered to the pretreatment level at 50h; while in the adductor muscle and hemocytes, the expression level remained higher than that of the control. The cloning and expression analyses of PuHSC70 provide theoretical basis to further study the mechanism of physiological response to thermal and high salinity stresses. PMID:24726551

  15. The molecular chaperone Hsp70 activates protein phosphatase 5 (PP5) by binding the tetratricopeptide repeat (TPR) domain.

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    Connarn, Jamie N; Assimon, Victoria A; Reed, Rebecca A; Tse, Eric; Southworth, Daniel R; Zuiderweg, Erik R P; Gestwicki, Jason E; Sun, Duxin

    2014-01-31

    Protein phosphatase 5 (PP5) is auto-inhibited by intramolecular interactions with its tetratricopeptide repeat (TPR) domain. Hsp90 has been shown to bind PP5 to activate its phosphatase activity. However, the functional implications of binding Hsp70 to PP5 are not yet clear. In this study, we find that both Hsp90 and Hsp70 bind to PP5 using a luciferase fragment complementation assay. A fluorescence polarization assay shows that Hsp90 (MEEVD motif) binds to the TPR domain of PP5 almost 3-fold higher affinity than Hsp70 (IEEVD motif). However, Hsp70 binding to PP5 stimulates higher phosphatase activity of PP5 than the binding of Hsp90. We find that PP5 forms a stable 1:1 complex with Hsp70, but the interaction appears asymmetric with Hsp90, with one PP5 binding the dimer. Solution NMR studies reveal that Hsc70 and PP5 proteins are dynamically independent in complex, tethered by a disordered region that connects the Hsc70 core and the IEEVD-TPR contact area. This tethered binding is expected to allow PP5 to carry out multi-site dephosphorylation of Hsp70-bound clients with a range of sizes and shapes. Together, these results demonstrate that Hsp70 recruits PP5 and activates its phosphatase activity which suggests dual roles for PP5 that might link chaperone systems with signaling pathways in cancer and development.

  16. Sulfatide-Hsp70 Interaction Promotes Hsp70 Clustering and Stabilizes Binding to Unfolded Protein

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    Yoichiro Harada

    2015-05-01

    Full Text Available The 70-kDa heat shock protein (Hsp70, one of the major stress-inducible molecular chaperones, is localized not only in the cytosol, but also in extracellular milieu in mammals. Hsp70 interacts with various cell surface glycolipids including sulfatide (3'-sulfogalactosphingolipid. However, the molecular mechanism, as well as the biological relevance, underlying the glycolipid-Hsp70 interaction is unknown. Here we report that sulfatide promotes Hsp70 oligomerization through the N-terminal ATPase domain, which stabilizes the binding of Hsp70 to unfolded protein in vitro. We find that the Hsp70 oligomer has apparent molecular masses ranging from 440 kDa to greater than 669 kDa. The C-terminal peptide-binding domain is dispensable for the sulfatide-induced oligomer formation. The oligomer formation is impaired in the presence of ATP, while the Hsp70 oligomer, once formed, is unable to bind to ATP. These results suggest that sulfatide locks Hsp70 in a high-affinity state to unfolded proteins by clustering the peptide-binding domain and blocking the binding to ATP that induces the dissociation of Hsp70 from protein substrates.

  17. 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

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    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.

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

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    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. PMID:26894764

  19. 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) 5.0 g/dL] is a leading 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...... 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....... Treatment of cultured peripheral blood mononuclear cells (PBMCs) with PfHz caused a time-dependent decrease in Hsp70 transcripts/protein, and NF-κB activation. Gln treatment of PBMCs overcame PfHz-induced suppression of HSP70 transcripts/protein, reduced NF-κB activation, and suppressed over...

  20. The Complex Function of Hsp70 in Metastatic Cancer

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    Kata Juhasz

    2013-12-01

    Full Text Available Elevated expression of the inducible heat shock protein 70 (Hsp70 is known to correlate with poor prognosis in many cancers. Hsp70 confers survival advantage as well as resistance to chemotherapeutic agents, and promotes tumor cell invasion. At the same time, tumor-derived extracellular Hsp70 has been recognized as a “chaperokine”, activating antitumor immunity. In this review we discuss localization dependent functions of Hsp70 in the context of invasive cancer. Understanding the molecular principles of metastasis formation steps, as well as interactions of the tumor cells with the microenvironment and the immune system is essential for fighting metastatic cancer. Although Hsp70 has been implicated in different steps of the metastatic process, the exact mechanisms of its action remain to be explored. Known and potential functions of Hsp70 in controlling or modulating of invasion and metastasis are discussed.

  1. Molecular cloning, characterization and tissue profiles of a HSP70 cDNA in Siberian sturgeon Acipenser baerii%西伯利亚鲟热休克蛋白HSP70 cDNA的克隆、序列分析和组织分布

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    田照辉; 徐绍刚; 王巍; 胡红霞; 董颖; 宋超

    2012-01-01

    A full length cDNA encoding heat shock protein 70 (HSP70) was cloned from liver RNA in Siberian sturgeon (Acipenser baerii) ,and was found to contain 2 343 bp, including 5'untranslated region of 140 bp, 3'un-translated region of 256 bp and an open reading frame of 1 947 bp. The deduced 648 amino acids sequence with estimated molecular of 71 000 and theoretical isoelectric point of 5. 21 had high similarity with other vertebrate HSP70 genes in NCBI. Sequence analysis indicated that the HSP70 cDNA was consisted of three signature sequences IDL-GTTYS, IFDLGGGTFDVSrL and IVLVGGSTRIPKIQK belonging to HSP70 family. Phylogenetic tree revealed that Alligator mississippiensis, Xenopus laevis ,Ambystoma mexicanum and Sebrian Sturgeon showed a close genetic relationship. Using fluorescent real-time quantitative PCR.the HSP70 was expressed in the liver, gill, spleen, heart, intestine, muscle, gonad and brain of Siberian sturgeon, with the peak in the spleen and the minimum in the liver (P<0.05) , indicating that the HSP70 cDNA could be used in understanding of anti-stress mechanism in Siberian sturgeon and in genetic breeding improvement.%采用普通PCR和RACE技术克隆了西伯利亚鲟Acipenser baerii热休克蛋白HSP70 cDNA的全序列,该序列全长为2343 bp,其中5'非编码区为140 bp,3'非编码区为256 bp,可读编码框(ORF)为1947bp,编码为648个氨基酸.该氨基酸序列中含有HSP70家族的3个特征序列——IDLGTTYS、IFDLGGGTFDVSIL和ⅣLVGGSTRIPKIQK,细胞质特征性保守序列为EEVD,C端重复序列为GGMP.该cDNA序列与其它生物的HSP70 cDNA序列一样具有很高的相似性.系统发育树显示,西伯利亚鲟与非洲爪蛙蟾Xenopus laevis、密西西比短吻鳄Alligator mississippiensis、美西螈Ambystoma mexicanum的亲缘关系较近.实时定量分析结果表明,水温为17.5℃时,西伯利亚鲟肝脏、鳃、脾脏、心脏、肌肉、中肠、性腺、脑8种组织中均有HSP70表达,其中HSP70在脾脏中的表达

  2. Targeting Hsp70: A possible therapy for cancer.

    Science.gov (United States)

    Kumar, Sanjay; Stokes, James; Singh, Udai P; Scissum Gunn, Karyn; Acharya, Arbind; Manne, Upender; Mishra, Manoj

    2016-04-28

    In all organisms, heat-shock proteins (HSPs) provide an ancient defense system. These proteins act as molecular chaperones by assisting proper folding and refolding of misfolded proteins and aid in the elimination of old and damaged cells. HSPs include Hsp100, Hsp90, Hsp70, Hsp40, and small HSPs. Through its substrate-binding domains, Hsp70 interacts with wide spectrum of molecules, ranging from unfolded to natively folded and aggregated proteins, and provides cytoprotective role against various cellular stresses. Under pathophysiological conditions, the high expression of Hsp70 allows cells to survive with lethal injuries. Increased Hsp70, by interacting at several points on apoptotic signaling pathways, leads to inhibition of apoptosis. Elevated expression of Hsp70 in cancer cells may be responsible for tumorigenesis and for tumor progression by providing resistance to chemotherapy. In contrast, inhibition or knockdown of Hsp70 reduces the size of tumors and can cause their complete regression. Moreover, extracellular Hsp70 acts as an immunogen that participates in cross presentation of MHC-I molecules. The goals of this review are to examine the roles of Hsp70 in cancer and to present strategies targeting Hsp70 in the development of cancer therapeutics. PMID:26898980

  3. Effect of environmental impact to molecular expression of heat-shock protein (HSP70) in oyster Crassostrea gigas from Gamak bay, Korea.

    Science.gov (United States)

    Cho, Eun-Seob; Jeong, Hee-Dong

    2012-05-01

    The cDNA of Crassostrea gigas HSP70 was cloned and rapid amplification of cDNA (RACE) techniques were used. The full length of HSP70 cDNA was 2045 bp, consisting of a 5' terminal untranslated region (UTR) of 80 bp, a 3' terminal UTR 146 bp, and an open reading frame (ORF) of 1829 bp encoding deduced 620 amino acids. The HSP70 cDNA contained HSP70 family signatures, ATP-GTP binding site motif, tetrapeptide (GGMP) and conserved carboxyl terminal region (EEVD) at C-terminal of deduced amino acid sequence. BLAST analysis revealed that the HSP70 gene has an extreme similarity of 98.9% with C. gigas (AF144646). Northern blotting was used to examine the expression of HSP70 mRNA in the gill tissue of the oyster obtained from surface, middle and bottom layers. The HSP70 mRNA observed the samples taken from middle and bottom layers in September and February, but samples from the surface layer did not find a signal intensity of HSP70 mRNA transcript. Consequently, it seems that the oyster occurring middle and bottom layer have been stressed during the period of summer and winter, which is associated with the massive mortality in Gamak bay. PMID:23029911

  4. Stimulation of TLR4 by recombinant HSP70 requires structural integrity of the HSP70 protein itself

    Directory of Open Access Journals (Sweden)

    Luong Michael

    2012-03-01

    Full Text Available Abstract Background Toll-like receptor 4 (TLR4 is activated by bacterial endotoxin, a prototypical pathogen-associated molecular pattern (PAMP. It has been suggested that TLR4 can also be activated by damage-associated molecular pattern (DAMP proteins such as HSP70. It remains a challenge to provide unequivocal evidence that DAMP proteins themselves play a role in TLR4 activation, as the DAMP proteins used are often contaminated with endotoxin and other TLR ligands introduced during protein expression and/or purification. Results Here we report that the activation of TLR4 on primary human macrophage cultures by recombinant HSP70 is not solely due to contaminating endotoxin. Polymyxin B pretreatment of HSP70 preparations to neutralize contaminating endotoxin caused significant reductions in the amount of TNF-α induced by the recombinant protein as determined by ELISA. However, digestion of HSP70 with Proteinase K-agarose beads also dramatically reduced the TNF-α response of macrophages to HSP70, while leaving levels of contaminating endotoxin largely unchanged relative to controls. Conclusions These results indicate that the stimulatory effect of recombinant HSP70 requires both the presence of endotoxin and structural integrity of the heat shock protein itself.

  5. 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

    The hsp70 (dnaK) locus of the moderate thermophilic archaeon Methanosarcina thermophila TM-1 was cloned, sequenced, and tested in vitro to measure gene induction by heat and ammonia, i.e., stressors pertinent to the biotechnological ecosystem of this methanogen that plays a key role in anaerobic......-negative bacteria - first described in the S-6 molecule and later found to be present in all homologs from archaea and Gram positives. The genes responded to a temperature elevation in a manner that demonstrated that they are heat-shock genes, functionally active in vivo. Ammonia also induced a heat-shock type...

  6. Roles of Hsp70s in Stress Responses of Microorganisms, Plants, and Animals

    Directory of Open Access Journals (Sweden)

    Anmin Yu

    2015-01-01

    Full Text Available Hsp70s (heat shock protein 70s are a class of molecular chaperones that are highly conserved and ubiquitous in organisms ranging from microorganisms to plants and humans. Most research on Hsp70s has focused on the mechanisms of their functions as molecular chaperones, but recently, studies on stress responses are coming to the forefront. Hsp70s play key roles in cellular development and protecting living organisms from environmental stresses such as heat, drought, salinity, acidity, and cold. Moreover, functions of human Hsp70s are related to diseases including neurological disorders, cancer, and virus infection. In this review, we provide an overview of the specific roles of Hsp70s in response to stress, particularly abiotic stress, in all living organisms.

  7. 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

  8. Expression and Purification of Goose HSP70 and Compound Formation with Virus Polypeptide

    Institute of Scientific and Technical Information of China (English)

    WANG Bao-wei; WU Xiao-ping; ZHANG Xu-hui; JIA Xiao-hui; ZHANG Ming-ai; LONG Fang-yu; YANG Zhi-gang; WANG Lei

    2008-01-01

    flow of the molecular sieve preliminary separation,ConA-agarose chromatography and the ADP-agarose chromatography.Under certain condition in vitro,the synthetic peptide could combine with HSP70 to form the compound,thereby providing a further experimental foundation for the immunity function of the complex.

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

    Science.gov (United States)

    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.

  10. Regulation of Hsp70 function by nucleotide-exchange factors

    OpenAIRE

    Gowda, Naveen Kumar Chandappa

    2016-01-01

    Protein folding is the process in which polypeptides in their non-native states attain the unique folds of their native states. Adverse environmental conditions and genetic predisposition challenge the folding process and accelerate the production of proteotoxic misfolded proteins. Misfolded proteins are selectively recognized and removed from the cell by processes of protein quality control (PQC). In PQC molecular chaperones of the Heat shock protein 70 kDa (Hsp70) family play important role...

  11. Detection of irradiation-induced, membrane heat shock protein 70 (Hsp70) in mouse tumors using Hsp70 Fab fragment

    International Nuclear Information System (INIS)

    Background and purpose: The major stress-inducible heat shock protein 70 (Hsp70) is frequently overexpressed in highly aggressive tumors, and elevated intracellular Hsp70 levels mediate protection against apoptosis. Following therapeutic intervention, such as ionizing irradiation, translocation of cytosolic Hsp70 to the plasma membrane is selectively increased in tumor cells and therefore, membrane Hsp70 might serve as a therapy-inducible, tumor-specific target structure. Materials and methods: Based on the IgG1 mouse monoclonal antibody (mAb) cmHsp70.1, we produced the Hsp70-specific recombinant Fab fragment (Hsp70 Fab), as an imaging tool for the detection of membrane Hsp70 positive tumor cells in vitro and in vivo. Results: The binding characteristics of Hsp70 Fab towards mouse colon (CT26) and pancreatic (1048) carcinoma cells at 4 deg. C were comparable to that of cmHsp70.1 mAb, as determined by flow cytometry. Following a temperature shift to 37 deg. C, Hsp70 Fab rapidly translocates into subcellular vesicles of mouse tumor cells. Furthermore, in tumor-bearing mice Cy5.5-conjugated Hsp70 Fab, but not unrelated IN-1 control Fab fragment (IN-1 ctrl Fab), gradually accumulates in CT26 tumors between 12 and 55 h after i.v. injection. Conclusions: In summary, the Hsp70 Fab provides an innovative, low immunogenic tool for imaging of membrane Hsp70 positive tumors, in vivo.

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

    Science.gov (United States)

    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...

  13. Molecular characterization of three heat shock protein 70 genes and their expression profiles under thermal stress in the citrus red mite.

    Science.gov (United States)

    Yang, Li-Hong; Jiang, Hong-Bo; Liu, Yong-Hua; Dou, Wei; Wang, Jin-Jun

    2012-04-01

    Three heat shock protein 70 family transcripts, named PcHsp70-1, PcHsp70-2 and PcHsp70-3, were isolated from the citrus red mite, Panonychus citri. PcHsp70-1, PcHsp70-2, and PcHsp70-3 contained an open reading frame of 1977, 1968, and 2028 nucleotides that encoded 658, 655 and 675 amino acid residues, respectively. Comparison of deduced amino acid sequences of PcHsp70-1 and PcHsp70-2 showed 86.34% identity, while the amino acid sequence of PcHsp70-3 was only 57.39 and 58.75% identical to that of PcHsp70-1 and PcHsp70-2, respectively. Sequences and phylogenetic analyses suggested that PcHsp70-1 and PcHsp70-2 were cytosolic Hsps, whereas PcHsp70-3 was located in ER (endoplasmic reticulum). To accurately validate mRNA expression profiles of the three Hsp70s under thermal stress conditions, seven housekeeping genes were evaluated. Alpha-tubulin and RpII were selected as optimal endogenous references for cold shock and heat shock conditions, respectively. Real-time quantitative RT-PCR revealed that only the mRNA expression of PcHsp70-2 was up-regulated under heat shocks, and all of the three Hsp70s were constitutively expressed under cold shocks. The results suggest that the three Hsp70s were more critical to coping with heat than cold shocks. PMID:21717055

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

    Directory of Open Access Journals (Sweden)

    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.

  15. Species- and Strain-Specific Adaptation of the HSP70 Super Family in Pathogenic Trypanosomatids.

    Science.gov (United States)

    Drini, Sima; Criscuolo, Alexis; Lechat, Pierre; Imamura, Hideo; Skalický, Tomáš; Rachidi, Najma; Lukeš, Julius; Dujardin, Jean-Claude; Späth, Gerald F

    2016-01-01

    All eukaryotic genomes encode multiple members of the heat shock protein 70 (HSP70) family, which evolved distinctive structural and functional features in response to specific environmental constraints. Phylogenetic analysis of this protein family thus can inform on genetic and molecular mechanisms that drive species-specific environmental adaptation. Here we use the eukaryotic pathogen Leishmania spp. as a model system to investigate the evolution of the HSP70 protein family in an early-branching eukaryote that is prone to gene amplification and adapts to cytotoxic host environments by stress-induced and chaperone-dependent stage differentiation. Combining phylogenetic and comparative analyses of trypanosomatid genomes, draft genome of Paratrypanosoma and recently published genome sequences of 204 L. donovani field isolates, we gained unique insight into the evolutionary dynamics of the Leishmania HSP70 protein family. We provide evidence for (i) significant evolutionary expansion of this protein family in Leishmania through gene amplification and functional specialization of highly conserved canonical HSP70 members, (ii) evolution of trypanosomatid-specific, non-canonical family members that likely gained ATPase-independent functions, and (iii) loss of one atypical HSP70 member in the Trypanosoma genus. Finally, we reveal considerable copy number variation of canonical cytoplasmic HSP70 in highly related L. donovani field isolates, thus identifying this locus as a potential hot spot of environment-genotype interaction. Our data draw a complex picture of the genetic history of HSP70 in trypanosomatids that is driven by the remarkable plasticity of the Leishmania genome to undergo massive intra-chromosomal gene amplification to compensate for the absence of regulated transcriptional control in these parasites. PMID:27371955

  16. Targeting membrane heat-shock protein 70 (Hsp70) on tumors by cmHsp70.1 antibody

    Science.gov (United States)

    Stangl, Stefan; Gehrmann, Mathias; Riegger, Julia; Kuhs, Kristin; Riederer, Isabelle; Sievert, Wolfgang; Hube, Kathrin; Mocikat, Ralph; Dressel, Ralf; Kremmer, Elisabeth; Pockley, Alan G.; Friedrich, Lars; Vigh, Laszlo; Skerra, Arne; Multhoff, Gabriele

    2011-01-01

    Immunization of mice with a 14-mer peptide TKDNNLLGRFELSG, termed “TKD,” comprising amino acids 450–461 (aa450–461) in the C terminus of inducible Hsp70, resulted in the generation of an IgG1 mouse mAb cmHsp70.1. The epitope recognized by cmHsp70.1 mAb, which has been confirmed to be located in the TKD sequence by SPOT analysis, is frequently detectable on the cell surface of human and mouse tumors, but not on isogenic cells and normal tissues, and membrane Hsp70 might thus serve as a tumor-specific target structure. As shown for human tumors, Hsp70 is associated with cholesterol-rich microdomains in the plasma membrane of mouse tumors. Herein, we show that the cmHsp70.1 mAb can selectively induce antibody-dependent cellular cytotoxicity (ADCC) of membrane Hsp70+ mouse tumor cells by unstimulated mouse spleen cells. Tumor killing could be further enhanced by activating the effector cells with TKD and IL-2. Three consecutive injections of the cmHsp70.1 mAb into mice bearing CT26 tumors significantly inhibited tumor growth and enhanced the overall survival. These effects were associated with infiltrations of NK cells, macrophages, and granulocytes. The Hsp70 specificity of the ADCC response was confirmed by preventing the antitumor response in tumor-bearing mice by coinjecting the cognate TKD peptide with the cmHsp70.1 mAb, and by blocking the binding of cmHsp70.1 mAb to CT26 tumor cells using either TKD peptide or the C-terminal substrate-binding domain of Hsp70. PMID:21187371

  17. Targeting membrane heat-shock protein 70 (Hsp70) on tumors by cmHsp70.1 antibody

    OpenAIRE

    Stangl, Stefan; Gehrmann, Mathias; Riegger, Julia; Kuhs, Kristin; Riederer, Isabelle; Sievert, Wolfgang; Hube, Kathrin; Mocikat, Ralph; Dressel, Ralf; Kremmer, Elisabeth; Pockley, Alan G.; Friedrich, Lars; Vigh, Laszlo; Skerra, Arne; Multhoff, Gabriele

    2010-01-01

    Immunization of mice with a 14-mer peptide TKDNNLLGRFELSG, termed “TKD,” comprising amino acids 450–461 (aa450–461) in the C terminus of inducible Hsp70, resulted in the generation of an IgG1 mouse mAb cmHsp70.1. The epitope recognized by cmHsp70.1 mAb, which has been confirmed to be located in the TKD sequence by SPOT analysis, is frequently detectable on the cell surface of human and mouse tumors, but not on isogenic cells and normal tissues, and membrane Hsp70 might thus serve as a tumor-s...

  18. Phosphorylation of an envelope-associated Hsp70 homolog in amyloplasts isolated from cultured cells of sycamore (Acer pseudoplatanus L.).

    Science.gov (United States)

    Checa, S K; Viale, A M

    1998-09-01

    The presence of Hsp70 and Hsp60 molecular chaperones in amyloplasts isolated from cultured sycamore cells was analyzed by immunoblotting. Hsp70 homologs were located in both amyloplast envelope and stromal fractions, but no Hsp60 homologs were detected in any of the different suborganellar fractions. Incubation of whole amyloplasts or their envelope fraction with Mg2+ gamma-32P-ATP resulted in a rapid phosphorylation of the envelope-associated Hsp70 homolog, which constitutes a major target of phosphorylation in these plastids.

  19. SSI1 encodes a novel Hsp70 of the Saccharomyces cerevisiae endoplasmic reticulum.

    OpenAIRE

    Baxter, B K; James, P; Evans, T.(Department of Physics, University of Oxford, Oxford, UK); Craig, E A

    1996-01-01

    The endoplasmic reticulum (ER) of the budding yeast Saccharomyces cerevisiae contains a well-characterized, essential member of the Hsp70 family of molecular chaperones, Kar2p. Kar2p has been shown to be involved in the translocation of proteins into the ER as well as the proper folding of proteins in that compartment. We report the characterization of a novel Hsp70 of the ER, Ssi1p. Ssi1p, which shares 24% of the amino acids of Kar2p, is not essential for growth under normal conditions. Howe...

  20. Imaging of Hsp70-positive tumors with cmHsp70.1 antibody-conjugated gold nanoparticles

    Directory of Open Access Journals (Sweden)

    Gehrmann MK

    2015-09-01

    Full Text Available Mathias K Gehrmann,1 Melanie A Kimm,2 Stefan Stangl,1 Thomas E Schmid,1 Peter B Noël,2 Ernst J Rummeny,2 Gabriele Multhoff11Department of Radiation Oncology, 2Department of Diagnostic and Interventional Radiology, Klinikum rechts der Isar, Technische Universität München, Munich, GermanyAbstract: Real-time imaging of small tumors is still one of the challenges in cancer diagnosis, prognosis, and monitoring of clinical outcome. Targeting novel biomarkers that are selectively expressed on a large variety of different tumors but not normal cells has the potential to improve the imaging capacity of existing methods such as computed tomography. Herein, we present a novel technique using cmHsp70.1 monoclonal antibody-conjugated spherical gold nanoparticles for quantification of the targeted uptake of gold nanoparticles into membrane Hsp70-positive tumor cells. Upon binding, cmHsp70.1-conjugated gold nanoparticles but not nanoparticles coupled to an isotype-matched IgG1 antibody or empty nanoparticles are rapidly taken up by highly malignant Hsp70 membrane-positive mouse tumor cells. After 24 hours, the cmHsp70.1-conjugated gold nanoparticles are found to be enriched in the perinuclear region. Specificity for membrane Hsp70 was shown by using an Hsp70 knockout tumor cell system. Toxic side effects of the cmHsp70.1-conjugated nanoparticles are not observed at a concentration of 1–10 µg/mL. Experiments are ongoing to evaluate whether cmHsp70.1 antibody-conjugated gold nanoparticles are suitable for the detection of membrane-Hsp70-positive tumors in vivo.Keywords: heat shock protein 70, tumor biomarker, theranostics, multimodal CT, multispectral CT, k-edge

  1. Structure and expression of a rice hsp70 gene

    Institute of Scientific and Technical Information of China (English)

    王群; 方荣祥

    1996-01-01

    A genomic hsp70 gene was isolated from a rice IR36 genomic library and 4 794 bp of the gene have been sequenoed. The 5’ flanking region of the gene contained a putative TATA box and a typical heat shock element sequence 5’-CTcgGAAccTTCgAG-3’. The amino acid sequence of the rice HSP70 deduced from the coding region shared 84%-92% homologies with those of HSP70s from other plant species. An intron 1939bp long was identified in the coding region at the codon specifying amino acid 72 (Asp), the similar position introns occurring in other intron-containing hsp70 genes. In addition, another intron of 57 bp was found in the 3’-untranslated region in the rice hsp70 gene. Southern blot hybridization showed that rice hsp70 gene family contained at least three members. Analysis of the RNA leveis with the gene-specific and non-specific probes revealed that the rice hsp70 gene expressed at normal temperature and the expression was enhanced by heat shock treatment.

  2. Inducible HSP70 Protects Radiation-Induced Salivary Gland Damage

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Hae-June; Lee, Yoon-Jin; Kwon, Hee-Choong; Lee, Su-Jae; Bae, Sang-Woo; Lee, Yun-Sil [Korea Institute of Radiological Medical Sciences, Seoul (Korea, Republic of); Kim, Sung-Ho [Chonnam National University, Gwangju (Korea, Republic of)

    2006-07-01

    Irradiation (IR) delivered to the head and neck is a common treatment for malignancies. Salivary glands in the irradiation field are severely damaged, and consequently this resulted in marked salivary hypofunction. While the exact mechanism of salivary gland damage remains enigmatic, fluid secreting acinar cells are lost, and saliva output is dramatically reduced. Previously we have reported that inducible heat shock protein 70 (HSP70i) induced radioresistance in vitro. Moreover, HSP70i localized to salivary glands by gene transfer has great potential for the treatment of salivary gland. Herein, we investigated whether HSP70 can use as radio protective molecules for radiation-induced salivary gland damage in vivo.

  3. Pathways of allosteric regulation in Hsp70 chaperones.

    Science.gov (United States)

    Kityk, Roman; Vogel, Markus; Schlecht, Rainer; Bukau, Bernd; Mayer, Matthias P

    2015-01-01

    Central to the protein folding activity of Hsp70 chaperones is their ability to interact with protein substrates in an ATP-controlled manner, which relies on allosteric regulation between their nucleotide-binding (NBD) and substrate-binding domains (SBD). Here we dissect this mechanism by analysing mutant variants of the Escherichia coli Hsp70 DnaK blocked at distinct steps of allosteric communication. We show that the SBD inhibits ATPase activity by interacting with the NBD through a highly conserved hydrogen bond network, and define the signal transduction pathway that allows bound substrates to trigger ATP hydrolysis. We identify variants deficient in only one direction of allosteric control and demonstrate that ATP-induced substrate release is more important for chaperone activity than substrate-stimulated ATP hydrolysis. These findings provide evidence of an unexpected dichotomic allostery mechanism in Hsp70 chaperones and provide the basis for a comprehensive mechanical model of allostery in Hsp70s. PMID:26383706

  4. The Anti-inflammatory mechanisms of Hsp70

    Directory of Open Access Journals (Sweden)

    Thiago J Borges

    2012-05-01

    Full Text Available Immune responses to heat shock proteins (Hsp develop in virtually all inflammatory diseases; however, the significance of such responses is only now becoming clear. In experimental disease models, Hsp administration can prevent or arrest inflammatory damage, and in initial clinical trials in patients with chronic inflammatory diseases, Hsp peptides have been shown to promote the production of anti-inflammatory cytokines, indicating immunoregulatory potential of Hsp. Therefore, the presence of immune responses to Hsp in inflammatory diseases can be seen as an attempt of the immune system to correct the inflammatory condition. Hsp70 can modulate inflammatory responses in models of arthritis, colitis and graft rejection, and the mechanisms underlying this effect are now being elucidated. Incubation with microbial Hsp70 was seen to induce tolerogenic DCs and to promote a suppressive phenotype in myeloid-derived suppressor cells and monocytes. These DC could induce regulatory T cells (Tregs, independently of the antigens they presented. Some Hsp70 family members are associated with autophagy, leading to a preferential uploading of Hsp70 peptides in MHC class II molecules of stressed cells. Henceforth, conserved Hsp70 peptides may be presented in these situations and constitute targets of Tregs, contributing to downregulation of inflammation. Finally, an interfering effect in multiple intracellular inflammatory signaling pathways is also known for Hsp70. Altogether it seems attractive to use Hsp70, or its derivative peptides, for modulation of inflammation. This is a physiological immunotherapy approach, without the immediate necessity of defining disease specific auto-antigens. In this article, we present the evidence on anti-inflammatory effects of Hsp70 and discuss the need for experiments that will be crucial for the further exploration of the immuno-suppressive potential of this protein.

  5. Pathways of allosteric regulation in Hsp70 chaperones

    OpenAIRE

    Kityk, Roman; Vogel, Markus; Schlecht, Rainer; Bukau, Bernd; Mayer, Matthias P

    2015-01-01

    Central to the protein folding activity of Hsp70 chaperones is their ability to interact with protein substrates in an ATP-controlled manner, which relies on allosteric regulation between their nucleotide-binding (NBD) and substrate-binding domains (SBD). Here we dissect this mechanism by analysing mutant variants of the Escherichia coli Hsp70 DnaK blocked at distinct steps of allosteric communication. We show that the SBD inhibits ATPase activity by interacting with the NBD through a highly ...

  6. Expression of stress response HSP70 gene in Asian paddle crabs, Charybdis japonica, exposure to endocrine disrupting chemicals, bisphenol A (BPA) and 4-nonylphenol (NP)

    Science.gov (United States)

    Park, Kiyun; Kwak, Ihn-Sil

    2013-06-01

    The Asian paddle crab, Charybdis japonica, is a potential bio-indicator reflecting marine sediment toxicity as well as a commercially important species living along coastal areas in Korea. This study investigated its stress response by looking at the heat shock protein (HSP70) gene of C. japonica when the organism is exposed to bisphenol A (BPA) and 4-nonylphenol (NP). We characterized partial sequence of HSP70 as the stressresponse gene of C. japonica. The nucleotide sequence of C. japonica HSP70 is over 90% homologous with the corresponding gene of other crabs. Phylogenetic tree analysis revealed a close relationship between C. japonica HSP70 and HSP70 in other species of lobster and shrimps. HSP70 mRNA transcripts were detected in all the examined tissues of C. japonica, with the highest level in gills, the organ that most frequently came into contact with the external BPA or NP-laden water. As no reference data were available for C. japonica crab exposure, the BPA and NP 24-h LC50 values have not been previously determined. The expression of the C. japonica HSP70 gene to various BPA or NP concentrations during short and longer times was assessed. Gene expression was significantly induced in concentration- and time-dependent manners after BPA or NP exposures. These results support the postulation that crab C. japonica HSP70 could be a potential stress response molecular marker to monitor marine ecosystems.

  7. Molecular cloning of hsf1 and hsbp1 cDNAs, and the expression of hsf1, hsbp1 and hsp70 under heat stress in the sea cucumber Apostichopus japonicus.

    Science.gov (United States)

    Xu, Dongxue; Sun, Lina; Liu, Shilin; Zhang, Libin; Yang, Hongsheng

    2016-08-01

    The heat shock response (HSR) is known for the elevated synthesis of heat shock proteins (HSPs) under heat stress, which is mediated primarily by heat shock factor 1 (HSF1). Heat shock factor binding protein 1 (HSBP1) and feedback control of heat shock protein 70 (HSP70) are major regulators of the activity of HSF1. We obtained full-length cDNA of genes hsf1 and hsbp1 in the sea cucumber Apostichopus japonicus, which are the second available for echinoderm (after Strongylocentrotus purpuratus), and the first available for holothurian. The full-length cDNA of hsf1 was 2208bp, containing a 1326bp open reading frame encoding 441 amino acids. The full-length cDNA of hsbp1 was 2850bp, containing a 225bp open reading frame encoding 74 amino acids. The similarities of A. japonicus HSF1 with other species are low, and much higher similarity identities of A. japonicus HSBP1 were shared. Phylogenetic trees showed that A. japonicus HSF1 and HSBP1 were clustered with sequences from S. purpuratus, and fell into distinct clades with sequences from mollusca, arthropoda and vertebrata. Analysis by real-time PCR showed hsf1 and hsbp1 mRNA was expressed constitutively in all tissues examined. The expression of hsf1, hsbp1 and hsp70 in the intestine at 26°C was time-dependent. The results of this study might provide new insights into the regulation of heat shock response in this species. PMID:26952354

  8. 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.

  9. Hsp70 enhances presentation of FMDV antigen to bovine CD4+ T cells in vitro

    OpenAIRE

    McLaughlin, Kerry; Seago, Julian; Robinson, Lucy; Kelly, Charles; Charleston, Bryan

    2010-01-01

    International audience Foot-and-mouth disease virus (FMDV) is the causative agent of a highly contagious acute vesicular disease affecting cloven-hoofed animals, including cattle, sheep and pigs. The current vaccine induces a rapid humoral response, but the duration of the protective antibody response is variable, possibly associated with a variable specific CD4+ T cell response. We investigated the use of heat shock protein 70 (Hsp70) as a molecular chaperone to target viral antigen to th...

  10. 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

    mechanisms. One such pathway includes the battery of stress response genes, especially the heat shock protein HSP70 genes. Three such genes, HSPA1A, HSPA1B and HSPA1L, are present within the MHC-III region on the short arm of chromosome 6. We and others have found alleles, genotypes and haplotypes which have...... 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...... to heat shock. Stress response genes, particularly HSP70, are now the major candidates in the gene-longevity association studies....

  11. Activation of phospholipase A2 by Hsp70 in vitro

    DEFF Research Database (Denmark)

    Mahalka, Ajay K; Code, Christian; Rezaijahromi, Behnam;

    2011-01-01

    We recently suggested a novel mechanism for the activation of phospholipase A2 (PLA2), with a (catalytically) highly active oligomeric state, which subsequently becomes inactivated by conversion into amyloid. This process can be activated by lysophosphatidylcholine which promotes both oligomeriza......We recently suggested a novel mechanism for the activation of phospholipase A2 (PLA2), with a (catalytically) highly active oligomeric state, which subsequently becomes inactivated by conversion into amyloid. This process can be activated by lysophosphatidylcholine which promotes both...... oligomerization and amyloid activation/inactivation. The heat shock protein 70 (Hsp70), has been demonstrated to be able to revert the conversion of a-synuclein and Alzheimer ß-peptide to amyloid fibrils in vitro. Accordingly, we would expect Hsp70 to sustain the lifetime of the active state of the enzyme...

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

    Directory of Open Access Journals (Sweden)

    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

  13. 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

    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...

  14. 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

    . In acidic environments Hsp70 binds with high affinity and specificity to BMP, thereby facilitating the BMP binding and activity of acid sphingomyelinase (ASM). The inhibition of the Hsp70-BMP interaction by BMP antibodies or a point mutation in Hsp70 (Trp90Phe), as well as the pharmacological and genetic...

  15. Heat shock protein 70 (Hsp70) stimulates proliferation and cytolytic activity of natural killer cells

    NARCIS (Netherlands)

    Multhoff, G; Mizzen, L; Winchester, CC; Milner, CM; Wenk, S; Eissner, G; Kampinga, HH; Laumbacher, B; Johnson, J

    1999-01-01

    We previously demonstrated that lysis of tumor cells that express Hsp70, the highly stress-inducible member of the HSP70 family, on their plasma membrane is mediated by natural killer (NK) cells. Here, we studied the effects of different proteins of the HSP70 family in combination with interleukin 2

  16. Electrostatic interactions of Hsp-organizing protein tetratricopeptide domains with Hsp70 and Hsp90: computational analysis and protein engineering.

    Science.gov (United States)

    Kajander, Tommi; Sachs, Jonathan N; Goldman, Adrian; Regan, Lynne

    2009-09-11

    The Hsp-organizing protein (HOP) binds to the C termini of the chaperones Hsp70 and Hsp90, thus bringing them together so that substrate proteins can be passed from Hsp70 to Hsp90. Because Hsp90 is essential for the correct folding and maturation of many oncogenic proteins, it has become a significant target for anti-cancer drug design. HOP binds to Hsp70 and Hsp90 via two independent tetratricopeptide (TPR) domains, TPR1 and TPR2A, respectively. We have analyzed ligand binding using Poisson-Boltzmann continuum electrostatic calculations, free energy perturbation, molecular dynamics simulations, and site-directed mutagenesis to delineate the contribution of different interactions to the affinity and specificity of the TPR-peptide interactions. We found that continuum electrostatic calculations could be used to guide protein design by removing unfavorable interactions to increase binding affinity, with an 80-fold increase in affinity for TPR2A. Contributions at buried charged residues, however, were better predicted by free energy perturbation calculations. We suggest using a combination of the two approaches for increasing the accuracy of results, with free energy perturbation calculations used only at selected buried residues of the ligand binding pocket. Finally we present the crystal structure of TPR2A in complex with its non-cognate Hsp70 ligand, which provides insight on the origins of specificity in TPR domain-peptide recognition. PMID:19586912

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

    Directory of Open Access Journals (Sweden)

    Mathias Gehrmann

    Full Text Available BACKGROUND: 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. METHODOLOGY/PRINCIPAL FINDINGS: 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. CONCLUSIONS

  18. Role of membrane Hsp70 in radiation sensitivity of tumor cells

    International Nuclear Information System (INIS)

    The major stress-inducible heat shock protein 70 (Hsp70) is frequently overexpressed in the cytosol and integrated in the plasma membrane of tumor cells via lipid anchorage. Following stress such as non-lethal irradiation Hsp70 synthesis is up-regulated. Intracellular located Hsp70 is known to exert cytoprotective properties, however, less is known about membrane (m)Hsp70. Herein, we investigate the role of mHsp70 in the sensitivity towards irradiation in tumor sublines that differ in their cytosolic and/or mHsp70 levels. The isogenic human colon carcinoma sublines CX+ with stable high and CX− with stable low expression of mHsp70 were generated by fluorescence activated cell sorting, the mouse mammary carcinoma sublines 4 T1 (4 T1 ctrl) and Hsp70 knock-down (4 T1 Hsp70 KD) were produced using the CRISPR/Cas9 system, and the Hsp70 down-regulation in human lung carcinoma sublines H1339 ctrl/H1339 HSF-1 KD and EPLC-272H ctrl/EPLC-272H HSF-1 KD was achieved by small interfering (si)RNA against Heat shock factor 1 (HSF-1). Cytosolic and mHsp70 was quantified by Western blot analysis/ELISA and flow cytometry; double strand breaks (DSBs) and apoptosis were measured by flow cytometry using antibodies against γH2AX and real-time PCR (RT-PCR) using primers and antibodies directed against apoptosis related genes; and radiation sensitivity was determined using clonogenic cell surviving assays. CX+/CX− tumor cells exhibited similar cytosolic but differed significantly in their mHsp70 levels, 4 T1 ctrl/4 T1 Hsp70 KD cells showed significant differences in their cytosolic and mHsp70 levels and H1339 ctrl/H1339 HSF-1 KD and EPLC-272H ctrl/EPLC-272H HSF-1 KD lung carcinoma cell sublines had similar mHsp70 but significantly different cytosolic Hsp70 levels. γH2AX was significantly up-regulated in irradiated CX− and 4 T1 Hsp70 KD with low basal mHsp70 levels, but not in their mHsp70 high expressing counterparts, irrespectively of their cytosolic Hsp70 content. After

  19. 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.

  20. Prokaryotic expression and purification of fusion protein HSP70-EGFP and its application in the study of dendritic cells internalization antigen

    Institute of Scientific and Technical Information of China (English)

    Ping QU; Yanfang SUI; Libing LIU; Jiahai MA; Guangsheng CHEN; Jiankang CHEN; Fang'e LIU

    2008-01-01

    To study the endocytic activity of dendritic cells (DCs) by obtaining fusion protein HSP70-EGFP as exogenous antigen and loading it with DCs derived from human peripheral blood. Fusion protein HSP70-EGFP was prokaryotically expressed, isolated and puri-fied. DCs were isolated and cultured from human peri-pheral blood. The DCs were divided into 3 groups in the endocytic experiment. There were 106 DCs in each group. Group 1 and 2 were respectively incubated for 30 min. with HSP70-EGFP and EGFP. Group 3 was incubated with HSP70 for 30 min, and then incubated for 30 min. with HSP70-EGFP. Subsequently, 3 groups were placed in an incubator at 37℃ for 0.5, 1,2 and 24 h. Flow cytometry (FCM) was adopted to detect the amount of DCs with EGFP inside. IL-12 Eli-spot was adopted to detect the amount of DCs which secreted IL-12. There were 5 types in the experiment: LPS, inactive LPS, HSP70-EGFP, EGFP and no antigen. Fusion pro-tein HSP70-EGFP was successfully obtained and its molecular weight was 97 000. It accounted for 35.32% of the total protein. Under irradiation of an ultraviolet lamp, the protein solution sent out viridescent fluor-escence. The result detected by FCM indicated that after incubation for 0.5 h at 37℃, the positive rate in group 1 was 63%, while the other 2 groups were negative. After incubation for 1, 2 and 24 h at 37℃, the positive rates in the 3 groups were above 80%. The IL-12 Eli-spot exam-ination shows that with HSP70-EGFP being loaded, the amount of DCs secreting IL-12 was 134.09±31.78/105 cells, a little lower than that of DCs with LPS loaded (with the average point of 156.36±15.73). There was no significant difference between the 2 groups (P<0.01). By contrast, both of them were significantly higher than inactive LPS-(33.78±1.40)/105 cells and EGFP-loaded (23.13±4,57)/105 cells DC groups in the amount of DCs secreting IL-12 (P<0.01). The results suggest that receptor-mediated phagocytosis plays a main role in the preliminary stage of DCs

  1. HSP70: a promising target for laryngeal carcinoma radiaotherapy by inhibiting cleavage and degradation of nucleolin

    Directory of Open Access Journals (Sweden)

    Xu Jing

    2010-08-01

    Full Text Available Abstract Previous studies have shown that heat shock proteins (HSPs were upregulated in various types of tumors and were associated with histological grade, recurrence and metastasis of malignant tumors. In this study, we investigated whether heat shock protein 70 kDa (HSP70 was associated with histological grade of laryngeal squamous cell carcinomas (LSCC. We also determine the role of HSP70 in LSCC radiation resistance using a laryngeal carcinoma xenograft model by antisense HSP70 RNA technique. Immunohistochemistry data showed that HSP70 was detected in 96% of LSCC tissues (48 out of 50. The expression level of HSP70 was significantly lower in early stage of LSCC than that in late stage (P = 0.015. Radiation treatment result showed that the volumes and weights of implantation tumors in the group injected with antisense HSP70 oligos were significantly reduced comparing to the group injected with random oligos(p

  2. 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;

    small HSPs). Hsp70 belongs to the HSP70 family and is expressed at low levels in normal non-stressed cells. Its expression is however induced by different cellular stresses, such as heat shock and oxidative stress. The function of Hsp70 depends on its cellular location: Intracellular it has...... 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...... cytoprotective and anti-apoptotic functions, whereas it exerts immunostimulatory functions extracellularly. Secreted Hsp70 is for example involved in cross-presentation of cancer-derived antigenic peptides, a function which is currently explored in immunotherapeutic approaches against cancer. Additionally...

  3. [PCR-RFLP/Hsp70 for identification and tipification of leishmania from the tropical region].

    Science.gov (United States)

    Margarita Montalvo, Ana; Fraga, Jorge; Aylema Romero, Jaqueline; Monzote, Lianet; Ivon Montano, Ing; Dujardin, Jean Claude

    2006-01-01

    The optimization of the PCR conditions for amplification of the gene coding for the 70 kDa (HSp70) heat shock protein as well as the analysis of the restriction fragment length polymorphism (RFLP) were carried out. DNA from a reference strain of Leishmania mexicana was used as template. Analytical sensitivity and specificity, and reproducibility of PCR using DNA from L. mexicana, Lamazonensis, L. guyanensis and L. lainsoni were determined. A 1.3 kp band was obtained, which confirmed gene amplification. The band patterns derived from Haelll enzyme digestion allowed differentiating several species. L. guyanensis and L. lainsoni were different from each other, while L. mexicana and L. amazonensis, which shared a common pattern, were different from the other two species. Analytical sensitivity and specificity were adequate. The enzymatic restriction of the PCR product made it possible to differentiate Leishmania spp. from T. cruzi. The feasibility of identifying and typifying species from the American continent through PCR-RFLP/Hsp70 and of using enzymatic restriction of amplified product to distinguish Leishmania spp. from Trypanosornma cruzi was shown. This was the first step in implementing these molecular methods in the reference laboratory of the Institute.

  4. Members of the Hsp70 family of proteins in the cell wall of Saccharomyces cerevisiae.

    OpenAIRE

    López-Ribot, J L; Chaffin, W L

    1996-01-01

    Western blot (immunoblot) analysis of cell wall and cytosolic extracts obtained from parental and ssa1 and ssa2 single- and double-mutant strains of Saccharomyces cerevisiae showed that the heat shock protein 70 (Hsp70) products of these genes, previously thought to be restricted to the cell interior, are also present in the cell wall. A cell wall location was further confirmed by indirect immunofluorescence with intact cells and biotinylation of extracellular Hsp70. Hsp70s have been implicat...

  5. Expression and Localization of an Hsp70 Protein in the Microsporidian Encephalitozoon cuniculi

    OpenAIRE

    Jolly, Carrie E.; Leonard, Cory A.; J. Russell Hayman

    2010-01-01

    Microsporidia spore surface proteins are an important, under investigated aspect of spore/host cell attachment and infection. For comparison analysis of surface proteins, we required an antibody control specific for an intracellular protein. An endoplasmic reticulum-associated heat shock protein 70 family member (Hsp70; ECU02_0100; “C1”) was chosen for further analysis. DNA encoding the C1 hsp70 was amplified, cloned and used to heterologously express the C1 Hsp70 protein, and...

  6. The role of HSP70 in mediating age-dependent mortality in sepsis

    Science.gov (United States)

    McConnell, Kevin W.; Fox, Amy C.; Clark, Andrew T.; Chang, Nai-Yuan Nicholas; Dominguez, Jessica A.; Farris, Alton B.; Buchman, Timothy G.; Hunt, Clayton R.; Coopersmith, Craig M.

    2011-01-01

    Sepsis is primarily a disease of the aged, with increased incidence and mortality occurring in aged hosts. Heat shock protein (HSP) 70 plays an important role in both healthy aging and the stress response to injury. The purpose of this study was to determine the role of HSP70 in mediating mortality and the host inflammatory response in aged septic hosts. Sepsis was induced in both young (6–12week old) and aged (16–17 month old) HSP70−/− and wild type (WT) mice to determine if HSP70 modulated outcome in an age-dependent fashion. Young HSP70−/− and WT mice subjected to cecal ligation and puncture (CLP), Pseudomonas aeruginosa pneumonia or Streptococcus pneumoniae pneumonia had no differences in mortality, suggesting HSP70 does not mediate survival in young septic hosts. In contrast, mortality was higher in aged HSP70−/− mice than aged WT mice subjected to CLP (p=0.01), suggesting HSP70 mediates mortality in sepsis in an age-dependent fashion. Compared to WT mice, aged septic HSP70−/− mice had increased gut epithelial apoptosis and pulmonary inflammation. In addition, HSP70−/−mice had increased systemic levels of TNF-α, IL-6, IL-10 and IL-1β compared to WT mice. These data demonstrate that HSP70 is a key determinant of mortality in aged but not young hosts in sepsis. HSP70 may play a protective role in an age-dependent response to sepsis by preventing excessive gut apoptosis and both pulmonary and systemic inflammation. PMID:21296977

  7. Hsp70 Knockdown by siRNA Decreased Collagen Production in Keloid Fibroblasts

    OpenAIRE

    Shin, Jung U; Lee, Won Jai; Tran, Thanh-Nga; Jung, Inhee; Lee, Ju Hee

    2015-01-01

    Purpose There are currently no consistently effective treatments for the excessive collagen produced by keloid fibroblasts. Previously, we reported that heat shock protein 70 (Hsp70) is up-regulated in keloid fibroblasts and keloid tissue. We, therefore, investigated whether Hsp70 is related to excessive collagen production in keloid fibroblasts. Materials and Methods We inhibited Hsp70 in keloid fibroblasts by RNA interference and examined the resulting collagen expression. Thus, we selected...

  8. Tumor-specific Hsp70 plasma membrane localization is enabled by the glycosphingolipid Gb3.

    Directory of Open Access Journals (Sweden)

    Mathias Gehrmann

    Full Text Available BACKGROUND: Human tumors differ from normal tissues in their capacity to present Hsp70, the major stress-inducible member of the HSP70 family, on their plasma membrane. Membrane Hsp70 has been found to serve as a prognostic indicator of overall patient survival in leukemia, lower rectal and non small cell lung carcinomas. Why tumors, but not normal cells, present Hsp70 on their cell surface and the impact of membrane Hsp70 on cancer progression remains to be elucidated. METHODOLOGY/PRINCIPAL FINDINGS: Although Hsp70 has been reported to be associated with cholesterol rich microdomains (CRMs, the partner in the plasma membrane with which Hsp70 interacts has yet to be identified. Herein, global lipid profiling demonstrates that Hsp70 membrane-positive tumors differ from their membrane-negative counterparts by containing significantly higher amounts of globotriaoslyceramide (Gb3, but not of other lipids such as lactosylceramide (LacCer, dodecasaccharideceramide (DoCer, galactosylceramide (GalCer, ceramide (Cer, or the ganglioside GM1. Apart from germinal center B cells, normal tissues are Gb3 membrane-negative. Co-localization of Hsp70 and Gb3 was selectively determined in Gb3 membrane-positive tumor cells, and these cells were also shown to bind soluble Hsp70-FITC protein from outside in a concentration-dependent manner. Given that the latter interaction can be blocked by a Gb3-specific antibody, and that the depletion of globotriaosides from tumors reduces the amount of membrane-bound Hsp70, we propose that Gb3 is a binding partner for Hsp70. The in vitro finding that Hsp70 predominantly binds to artificial liposomes containing Gb3 (PC/SM/Chol/Gb3, 17/45/33/5 confirms that Gb3 is an interaction partner for Hsp70. CONCLUSIONS/SIGNIFICANCE: These data indicate that the presence of Gb3 enables anchorage of Hsp70 in the plasma membrane of tumors and thus they might explain tumor-specific membrane localization of Hsp70.

  9. Hsp70 chaperone systems: diversity of cellular functions and mechanism of action.

    Science.gov (United States)

    Mayer, M P; Bukau, B

    1998-03-01

    Hsp70 chaperone systems play an essential role in the life cycle of many proteins not only in an hostile environment but also under normal growth conditions. In the course of evolution the diversification of functions was accompanied by an amplification of components of the Hsp70 system. Here strategies are reviewed how different Hsp70 systems work independently or cooperate with each other in a functional network to perform their housekeeping tasks even under stress conditions. We further discuss how co-chaperones which act as targeting factors regulate the cycle of substrate binding and release upon which the Hsp70 chaperone activity depends.

  10. Proteomics shows Hsp70 does not bind peptide sequences indiscriminately in vivo

    International Nuclear Information System (INIS)

    Heat shock protein 70 (Hsp70) binds peptide and has several functions that include protein folding, protein trafficking, and involvement with immune function. However, endogenous Hsp70-binding peptides had not previously been identified. Therefore, we eluted and identified several hundred endogenously bound peptides from Hsp70 using liquid chromatography ion trap mass spectrophotometry (LC-ITMS). Our work shows that the peptides are capable of binding Hsp70 as previously described. They are generally 8-26 amino acids in length and correspond to specific regions of many proteins. Through computationally assisted analysis of peptides eluted from Hsp70 we determined variable amino acid sequences, including a 5 amino acid core sequence that Hsp70 favorably binds. We also developed a computer algorithm that predicts Hsp70 binding within proteins. This work helps to define what peptides are bound by Hsp70 in vivo and suggests that Hsp70 facilitates peptide selection by aiding a funneling mechanism that is flexible but allows only a limited number of peptides to be processed

  11. Yeast Two-Hybrid and One-Hybrid Screenings Identify Regulators of hsp70 Gene Expression.

    Science.gov (United States)

    Saito, Youhei; Nakagawa, Takanobu; Kakihana, Ayana; Nakamura, Yoshia; Nabika, Tomomi; Kasai, Michihiro; Takamori, Mai; Yamagishi, Nobuyuki; Kuga, Takahisa; Hatayama, Takumi; Nakayama, Yuji

    2016-09-01

    The mammalian stress protein Hsp105β, which is specifically expressed during mild heat shock and localizes to the nucleus, induces the major stress protein Hsp70. In the present study, we performed yeast two-hybrid and one-hybrid screenings to identify the regulators of Hsp105β-mediated hsp70 gene expression. Six and two proteins were detected as Hsp105β- and hsp70 promoter-binding proteins, respectively. A luciferase reporter gene assay revealed that hsp70 promoter activation is enhanced by the transcriptional co-activator AF9 and splicing mediator SNRPE, but suppressed by the coiled-coil domain-containing protein CCDC127. Of these proteins, the knockdown of SNRPE suppressed the expression of Hsp70 irrespective of the presence of Hsp105β, indicating that SNRPE essentially functions as a transcriptional activator of hsp70 gene expression. The overexpression of HSP70 in tumor cells has been associated with cell survival and drug resistance. We here identified novel regulators of Hsp70 expression in stress signaling and also provided important insights into Hsp70-targeted anti-cancer therapy. J. Cell. Biochem. 117: 2109-2117, 2016. © 2016 Wiley Periodicals, Inc. PMID:26873636

  12. Low resolution structural characterization of the Hsp70-interacting protein - Hip - from Leishmania braziliensis emphasizes its high asymmetry.

    Science.gov (United States)

    Dores-Silva, P R; Silva, E R; Gomes, F E R; Silva, K P; Barbosa, L R S; Borges, J C

    2012-04-15

    The Hsp70 is an essential molecular chaperone in protein metabolism since it acts as a pivot with other molecular chaperone families. Several co-chaperones act as regulators of the Hsp70 action cycle, as for instance Hip (Hsp70-interacting protein). Hip is a tetratricopeptide repeat protein (TPR) that interacts with the ATPase domain in the Hsp70-ADP state, stabilizing it and preventing substrate dissociation. Molecular chaperones from protozoans, which can cause some neglected diseases, are poorly studied in terms of structure and function. Here, we investigated the structural features of Hip from the protozoa Leishmania braziliensis (LbHip), one of the causative agents of the leishmaniasis disease. LbHip was heterologously expressed and purified in the folded state, as attested by circular dichroism and intrinsic fluorescence emission techniques. LbHip forms an elongated dimer, as observed by analytical gel filtration chromatography, analytical ultracentrifugation and small angle X-ray scattering (SAXS). With the SAXS data a low resolution model was reconstructed, which shed light on the structure of this protein, emphasizing its elongated shape and suggesting its domain organization. We also investigated the chemical-induced unfolding behavior of LbHip and two transitions were observed. The first transition was related to the unfolding of the TPR domain of each protomer and the second transition of the dimer dissociation. Altogether, LbHip presents a similar structure to mammalian Hip, despite their low level of conservation, suggesting that this class of eukaryotic protein may use a similar mechanism of action. PMID:22387434

  13. INDUCEMENT OF ANTITUMOR-IMMUNITY BY DC ACTIVATED BY HSP70-H22 TUMOR ANTIGEN PEPTIDE

    Institute of Scientific and Technical Information of China (English)

    冯作化; 黄波; 张桂梅; 李东; 王洪涛

    2003-01-01

    Objective: To investigate the feasibility of decreasing the dosage of tumor antigen peptides by dendritic cell (DC)-presenting and the characteristics of modification of DC by heat shock protein (Hsp70) and antigen peptides. Methods: Peptides were bound to Hsp70 and used to modify DC in vitro. The metabolism of the modified DC and the cytokines secreted by the modified DC were determined. The activation of lymphocytes by the modified DC and Hsp70-H22 peptides was tested. The cytotoxicity of the activated lymphocytes to H22 tumor cells was analyzed. The inhibitory effect of tumor in mice by the injection of DC and Hsp70-H22 peptides was tested. Results: 0.15μg of H22 peptides bound with Hsp70 could make 2×105 DC mature. 4×103 matured DC could activate 2×106 lymphocytes. The same amount of lymphocytes could be activated to produce similar cytotoxicity to tumor cells by either DC modified by 0.003μg of peptides bound with Hsp70 or by direct stimulation with 0.15μg of peptides bound with Hsp70. The dosage of peptides could be reduced by about 50 folds if the modified DC was used for injection instead of Hsp70-peptides. Peptides from normal hepatocytes, bound with Hsp70, could not make DC mature, nor activate lymphocytes through DC. Conclusion: The dosage of Hsp70-H22 peptides can be reduced significantly by DC-presenting to activate lymphocytes. Peptides from normal cells could not activate lymphocytes by either Hsp70-presenting or DC-presenting and they have little chance to induce autoimmunity.

  14. 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

    We show that inhibition of HDAC activity leads to surface expression of Hsp70 on various hematopoietic cancer cells, an occurance that was not observed on naïve or activated peripheral blood cells. HDAC inhibitor-mediated Hsp70 surface expression was confined to the apoptotic Annexin V-positive c......We show that inhibition of HDAC activity leads to surface expression of Hsp70 on various hematopoietic cancer cells, an occurance that was not observed on naïve or activated peripheral blood cells. HDAC inhibitor-mediated Hsp70 surface expression was confined to the apoptotic Annexin V......-positive cells and blocked by inhibition of apoptosis. Other chemotherapeutic inducers of apoptosis such as etoposide and camptothecin also led to a robust induction of Hsp70 surface expression. Hsp70 expression was, however, not caused by induction of apoptosis per se, as activated CD4 T cells remained Hsp70...... 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...

  15. 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.

  16. Could Upregulated Hsp70 Protein Compensate for the Hsp90-Silence-Induced Cell Death in Glioma Cells?

    Directory of Open Access Journals (Sweden)

    Chinmay Munje

    2014-01-01

    Full Text Available The molecular chaperone heat shock protein 90 alpha (Hsp90α has been recognized in various tumours including glioma. This pilot study using a proteomic approach analyses the downstream effects of Hsp90 inhibition using 17-allylamino-17-demethoxygeldanamycin (17AAG and a short hairpin RNA (shRNA oligonucleotide targeting hsp90α (shhsp90α in the U87-MG glioma cell line. Preliminary data coupled with bioinformatic analysis identified several known and unknown Hsp90 client proteins that demonstrated a change in their protein expression after Hsp90 inhibition, signifying an alteration in the canonical pathways of cell cycle progression, apoptosis, cell invasion, angiogenesis, and metastasis. Members of the glycolysis pathway were upregulated, demonstrating increased dependency on glycolysis for energy source by the treated glioma cells. Upregulated proteins also include Hsp70 and members of its family such as Hsp27 and gp96, thereby suggesting the role of Hsp90 co-chaperones in compensating for Hsp90 function after Hsp90 inhibition. Considering Hsp70’s role in antiapoptosis, it was postulated that a combination therapy involving a multitarget approach could be carried out. Consequently inhibition of both Hsp90 and Hsp70 in U87-MG glioma cells resulted in 60% cell death indicating the importance of combination therapy for glioma therapeutics.

  17. Dynamic Analysis of the Expression of HSP70 during Experimental Tooth Movement in Rats

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    In this study, the expression of HSP70 during experimental tooth movement was dynamically observed and the relationship between HSP70 and orthodontic periodontal tissue remodeling were observed. The orthodontic appliance was placed between the right maxillary first molar and maxillary central incisors of adult SD rats to establish a rat molar movement model. Immunohistochemistry was performed 1, 3, 5, 7 and 14 day(s) after orthodontic force application to observe the expression and localization of inducible HSP70. The expression of HSP70 was strongly positive in the early stage of the tooth movement, became gradually less positive, and was weakly positive in the restoration stage. There was difference in staining pattern between different parts of PDL during the same period. These results suggest that the expression of HSP70 and difference in staining pattern among different parts of PDL during orthodontic tooth movement in rats may be implicated in stress response and remodeling of periodontal tissue.

  18. Expression of HSP70 in Peripheral Lymphocytes of the Patients with Allergic Rhinitis

    Institute of Scientific and Technical Information of China (English)

    刘立思; 肖成峰; 张明; 程磊; 王鄂芬; 邬堂春

    2003-01-01

    The expression levels of heat shock protein 70 (HSP70) from peripheral lymphocytes ofthe patients with allergic rhinitis (AR) and the clinical implication were investigated. In the morn-ing, 3 ml of fasting venous blood was taken out. The lymphocytes were isolated by using Ficoll-Hypaque and the expression of HSP70 in the lymphocytes was detected by using Western blot. Inthe AR patients the HSP70 level (41.49± 15.77 integrated optical density, IOD) were significantlyhigher than that in the control group (23.89±10. 13 IOD, P<0.05). Western blot demonstratedthat HSP70 bands in AR patients were more intensive than those in the control group. It was con-cluded that the elevated HSP70 level in peripheral lymphocytes of the AR patients might contributeto the development of AR.

  19. 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...

  20. Glial Hsp70 Protects K+ Homeostasis in the Drosophila Brain during Repetitive Anoxic Depolarization

    Science.gov (United States)

    Armstrong, Gary A. B.; Xiao, Chengfeng; Krill, Jennifer L.; Seroude, Laurent; Dawson-Scully, Ken; Robertson, R. Meldrum

    2011-01-01

    Neural tissue is particularly vulnerable to metabolic stress and loss of ion homeostasis. Repetitive stress generally leads to more permanent dysfunction but the mechanisms underlying this progression are poorly understood. We investigated the effects of energetic compromise in Drosophila by targeting the Na+/K+-ATPase. Acute ouabain treatment of intact flies resulted in subsequent repetitive comas that led to death and were associated with transient loss of K+ homeostasis in the brain. Heat shock pre-conditioned flies were resistant to ouabain treatment. To control the timing of repeated loss of ion homeostasis we subjected flies to repetitive anoxia while recording extracellular [K+] in the brain. We show that targeted expression of the chaperone protein Hsp70 in glial cells delays a permanent loss of ion homeostasis associated with repetitive anoxic stress and suggest that this is a useful model for investigating molecular mechanisms of neuroprotection. PMID:22174942

  1. Glial Hsp70 protects K+ homeostasis in the Drosophila brain during repetitive anoxic depolarization.

    Directory of Open Access Journals (Sweden)

    Gary A B Armstrong

    Full Text Available Neural tissue is particularly vulnerable to metabolic stress and loss of ion homeostasis. Repetitive stress generally leads to more permanent dysfunction but the mechanisms underlying this progression are poorly understood. We investigated the effects of energetic compromise in Drosophila by targeting the Na(+/K(+-ATPase. Acute ouabain treatment of intact flies resulted in subsequent repetitive comas that led to death and were associated with transient loss of K(+ homeostasis in the brain. Heat shock pre-conditioned flies were resistant to ouabain treatment. To control the timing of repeated loss of ion homeostasis we subjected flies to repetitive anoxia while recording extracellular [K(+] in the brain. We show that targeted expression of the chaperone protein Hsp70 in glial cells delays a permanent loss of ion homeostasis associated with repetitive anoxic stress and suggest that this is a useful model for investigating molecular mechanisms of neuroprotection.

  2. Hsp70 as an indicator of stress in the cells after contact with nanoparticles

    Science.gov (United States)

    Hardilová, Šárka; Havrdová, Markéta; Panáček, Aleš; Kvítek, Libor; Zbořil, Radek

    2015-05-01

    In recent years, production of nanoparticles is increased and thus grows our contact with them too. Question of safety is closely related to the issue of use nanoparticles. There are a number of tests that monitor the viability, ROS production, the effect on the DNA and cell cycle, however, rarely encountered studies on stress in the cells after contact with nanoparticles. Heat shock proteins (HSP) are among the substances that can be used for monitoring stress in cells. HSP are structures with a chaperone activity. They are evolutionarily very old, conservative and they are found with a high degree of homology in prokaryotes and eukaryotes including humans. They exist at low concentrations under physiological conditions, while in the denaturing conditions e.g. high or low temperature, radiation, exposure to chemicals, heavy metals, or nanoparticles their expression is changed. HSPs are involved in maintaining homeostasis in the cell that the denatured protein conformations allow recovery to the original stage. One of the most common proteins from HSP family is Hsp70 - protein with a molecular weight of 70 kDa. The level of Hsp70 in a cell after exposure to the stress changes depending on the stress level to which the cell is exposed to and a time period during which lasted stressful conditions. Our research monitors stress levels of cells manifesting by Hsp70 production after contact with silver nanoparticles. Nanoparticles show different toxicity towards different types of target cells, which is reflected in the values of IC50 - concentration that kills 50% tested cells. Concentration of test substance toxic to one cell type may be innocuous to cells of another type. IC50 obtained from the MTT assay provides a suitable default data and if multiples of IC50 values are used, we can compare and generalize. Studies can be used to compare stress levels in cells that show different sensitivity to the tested nanoparticles compared with cells under optimal growth

  3. Compound A, a selective glucocorticoid receptor modulator, enhances heat shock protein Hsp70 gene promoter activation.

    Directory of Open Access Journals (Sweden)

    Ilse M Beck

    Full Text Available Compound A possesses glucocorticoid receptor (GR-dependent anti-inflammatory properties. Just like classical GR ligands, Compound A can repress NF-κB-mediated gene expression. However, the monomeric Compound A-activated GR is unable to trigger glucocorticoid response element-regulated gene expression. The heat shock response potently activates heat shock factor 1 (HSF1, upregulates Hsp70, a known GR chaperone, and also modulates various aspects of inflammation. We found that the selective GR modulator Compound A and heat shock trigger similar cellular effects in A549 lung epithelial cells. With regard to their anti-inflammatory mechanism, heat shock and Compound A are both able to reduce TNF-stimulated IκBα degradation and NF-κB p65 nuclear translocation. We established an interaction between Compound A-activated GR and Hsp70, but remarkably, although the presence of the Hsp70 chaperone as such appears pivotal for the Compound A-mediated inflammatory gene repression, subsequent novel Hsp70 protein synthesis is uncoupled from an observed CpdA-induced Hsp70 mRNA upregulation and hence obsolete in mediating CpdA's anti-inflammatory effect. The lack of a Compound A-induced increase in Hsp70 protein levels in A549 cells is not mediated by a rapid proteasomal degradation of Hsp70 or by a Compound A-induced general block on translation. Similar to heat shock, Compound A can upregulate transcription of Hsp70 genes in various cell lines and BALB/c mice. Interestingly, whereas Compound A-dependent Hsp70 promoter activation is GR-dependent but HSF1-independent, heat shock-induced Hsp70 expression alternatively occurs in a GR-independent and HSF1-dependent manner in A549 lung epithelial cells.

  4. Use of an enzyme-linked immunosorbent assay (Elisa) for the determination of HSP 70

    International Nuclear Information System (INIS)

    It has been suggested that HSP 70 is a good indicator to quantify thermotolerance. The authors developed an enzyme-linked immunosorbent assay (Elisa) for measuring the total amount of HSP 70 in mammalian cells. Microtitration plates were incubated with various concentrations of cell lysate in PBS. Milk proteins were used to block non-specific protein binding sites in the wells. Monoclonal antibody against purified HSP 70 raised in a mouse was added, followed by peroxidase conjugated goat-anti-mouse antibody. After washing, the bound peroxidase activity was measured colorimetrically to calculate the original amount of HSP 70 antigen. Kinetic studies indicate that the linear range of the assay corresponds to the amount of HSP 70 from 103-104 HA-1 cells (0.1-2.0 μg cytosolic proteins in each well). Elisa is a highly sensitive method for quantifying HSP 70 and can be easily used to measure the total amount of HSP 70 instead of the synthesis rate of HSP 70 by pulse radiolabeling. The authors are currently developing the sensitive Elisa system for measurement of thermotolerance in human tumors. A comparison of measurement of HSP 70 by pulse labeling the cells with 35S-methionine and by Elisa has been done in HA-1 cells. HA-1 cells were exposed to 450C, 10 min, then incubated at 370C for 0-24 h. These results show that the rate of HSP synthesis increased several fold, although the total amount of HSP 70 is increased less than 2 fold during thermotolerance induction. These results are consistent with data in the literature obtained by other techniques

  5. Increased hsp70 of glucocorticoid receptor complex induced by scald and heat stress and its possible effect on the affinity of glucocorticoid receptor

    Institute of Scientific and Technical Information of China (English)

    WANG Xiao-hui; TANG Hong-tai; LU Jian; XIA Zhao-fan

    2010-01-01

    Background Glucocorticoid (GC) insensitivity/GC resistance is an important etiological and prognostic factor in multiple diseases and pathophysiological processes such as scald, shock and asthma. The function of GC was mediated by glucocorticoid receptor (GR). Scald not only decreased the expression of GR but also reduced the affinity of GR, which played an important role in GC resistance in scalded rats. Whereas the molecular mechanism responsible for the decrease of GR affinity resulted from scald remains unclear. Recent studies showed that the changes of heat shock proteins (hsp) especially hsp90 and hsp70 of GR heterocomplex were associated with GR low affinity in vitro. Methods The affinity of GR in hepatic cytosols and in the cytosols of SMMC-7721 cells were determined by radioligand binding assay and scatchard plot. GR heterocomplex in cytosols were captured by coimmunoprecipation and the levels of hsp90 and hsp70 of GR complex were detected by quantitative Western blotting.Results Similar with that of hepatic cytosol of scalded rats, a remarkable decrease of GR affinity was also found in the cytosol of heat stressed SMMC-7721 cells. The level of hsp70 of GR complex in hepatic cytosol of scalded rats (30% total body surface area immersion scald) and in cytosol of heat stressed human hepatocarcinoma cell line SMMC-7721 were both increased by 1.5 fold, whereas no change of hsp90 in GR heterocomplex was found. According to the correlation analysis, there may be a positive relationship between increased hsp70 of GR complex and decreased GR affinity in the cytosols.Conclusions The primary results indicated that the level of hsp70 of GR heterocomplex was increased in the hepatic cytosol of scalded rats and the cytosol of heat stressed SMMC-7721 cells. The increase of hsp70 of GR complex might be associated with the decrease of GR affinity.

  6. 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.

  7. In silico identification of carboxylate clamp type tetratricopeptide repeat proteins in Arabidopsis and rice as putative co-chaperones of Hsp90/Hsp70.

    Directory of Open Access Journals (Sweden)

    Bishun D Prasad

    Full Text Available The essential eukaryotic molecular chaperone Hsp90 operates with the help of different co-chaperones, which regulate its ATPase activity and serve as adaptors to recruit client proteins and other molecular chaperones, such as Hsp70, to the Hsp90 complex. Several Hsp90 and Hsp70 co-chaperones contain the tetratricopeptide repeat (TPR domain, which interacts with the highly conserved EEVD motif at the C-terminal ends of Hsp90 and Hsp70. The acidic side chains in EEVD interact with a subset of basic residues in the TPR binding pocket called a 'carboxylate clamp'. Since the carboxylate clamp residues are conserved in the TPR domains of known Hsp90/Hsp70 co-chaperones, we carried out an in silico search for TPR proteins in Arabidopsis and rice comprising of at least one three-motif TPR domain with conserved amino acid residues required for Hsp90/Hsp70 binding. This approach identified in Arabidopsis a total of 36 carboxylate clamp (CC-TPR proteins, including 24 novel proteins, with potential to interact with Hsp90/Hsp70. The newly identified CC-TPR proteins in Arabidopsis and rice contain additional protein domains such as ankyrin, SET, octicosapeptide/Phox/Bem1p (Phox/PB1, DnaJ-like, thioredoxin, FBD and F-box, and protein kinase and U-box, indicating varied functions for these proteins. To provide proof-of-concept of the newly identified CC-TPR proteins for interaction with Hsp90, we demonstrated interaction of AtTPR1 and AtTPR2 with AtHsp90 in yeast two-hybrid and in vitro pull down assays. These findings indicate that the in silico approach used here successfully identified in a genome-wide context CC-TPR proteins with potential to interact with Hsp90/Hsp70, and further suggest that the Hsp90/Hsp70 system relies on TPR co-chaperones more than it was previously realized.

  8. Enhanced Hsp70 expression protects against acute lung injury by modulating apoptotic pathways.

    Directory of Open Access Journals (Sweden)

    Gabriella Aschkenasy

    Full Text Available The Acute respiratory distress syndrome (ARDS is a highly lethal inflammatory lung disorder. Apoptosis plays a key role in its pathogenesis. We showed that an adenovirus expressing the 70 kDa heat shock protein Hsp70 (AdHSP protected against sepsis-induced lung injury. In this study we tested the hypothesis that AdHSP attenuates apoptosis in sepsis-induced lung injury. Sepsis was induced in rats via cecal ligation and double puncture (2CLP. At the time of 2CLP PBS, AdHSP or AdGFP (an adenoviral vector expressing green fluorescent protein were injected into the tracheas of septic rats. 48 hours later, lungs were isolated. One lung was fixed for TUNEL staining and immunohistochemistry. The other was homogenized to isolate cytosolic and nuclear protein. Immunoblotting, gel filtration and co-immunoprecipitation were performed in these extracts. In separate experiments MLE-12 cells were incubated with medium, AdHSP or AdGFP. Cells were stimulated with TNFα. Cytosolic and nuclear proteins were isolated. These were subjected to immunoblotting, co-immunoprecipitation and a caspase-3 activity assay. TUNEL assay demonstrated that AdHSP reduced alveolar cell apoptosis. This was confirmed by immunohistochemical detection of caspase 3 abundance. In lung isolated from septic animals, immunoblotting, co-immunoprecipitation and gel filtration studies revealed an increase in cytoplasmic complexes containing caspases 3, 8 and 9. AdHSP disrupted these complexes. We propose that Hsp70 impairs apoptotic cellular pathways via interactions with caspases. Disruption of large complexes resulted in stabilization of lower molecular weight complexes, thereby, reducing nuclear caspase-3. Prevention of apoptosis in lung injury may preserve alveolar cells and aid in recovery.

  9. Expression and Significance of HSP70 and MGMT in Breast Neoplasms%HSP70和MGMT在乳腺肿瘤中的表达和意义

    Institute of Scientific and Technical Information of China (English)

    吴荣薇; 薄爱华; 王发亮; 芦广萍

    2008-01-01

    目的:探讨乳腺肿瘤组织中热休克蛋白HSP70和DNA修复酶O6-甲基鸟嘌呤-DNA甲基转移酶(O6-methyguanine-DNA methytransferase,MGMT)的表达及临床意义.方法:采用免疫组织化学SABC法检测65例乳腺肿瘤组织中HSP70和MGMT的表达情况.结果:乳腺肿瘤中HSP70的阳性率为80.00%(52/65),MGMT的阳性率为60.00%(39/65),二者呈中度相关性(r=0.46).其中,有淋巴结转移组的阳性率高于无转移组(P<0.05).结论:HSP70和MGMT的表达率与淋巴结转移情况有关.检测HSP70与MGMT对乳腺肿瘤预后和化疗方案的选择有指导意义.

  10. Stress-induced interaction between p38 MAPK and HSP70

    International Nuclear Information System (INIS)

    Highlights: ► HSP70 interacts to p38 MAPK in vitro and in vivo. ► HSP70 co-localizes with p38 MAPK in the nucleus upon stress. ► HSP70 is involved in the nuclear phosphorylation of MK2 by p38 MAPK. -- Abstract: p38 MAPK, one of the four MAPK subfamilies in mammalian cells, is activated by environmental stresses and pro-inflammatory cytokines, playing fundamental roles in many biological processes. Despite all that is known on the structure and functions of p38, many questions still exist. The coupling of activation and nuclear translocation represents an important aspect of p38 signaling. In our effort in exploring the potential chaperone for p38 translocation, we performed an endogenous pull-down assay and identified HSP70 as a potential interacting protein of p38. We confirmed the interaction between p38 and HSP70 in vitro and in vivo, and identified their interaction domains. We also showed stress-induced nuclear co-localization of these two proteins. Our preliminary result indicated that HSP70 was related to the phosphorylation of MK2, a specific nuclear downstream target of p38, suggesting HSP70 is a potential chaperone for the nuclear translocation of p38.

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

    DEFF Research Database (Denmark)

    Jensen, Helle

    The function of Hsp70 depends on its cellular location: When located intracellularly it exerts cytoprotective and anti-apoptotic functions, whereas it exerts immunostimulatory functions when located extracellularly. Secreted Hsp70 is for example involved in cross-presentation of cancer-derived an......The function of Hsp70 depends on its cellular location: When located intracellularly it exerts cytoprotective and anti-apoptotic functions, whereas it exerts immunostimulatory functions when located extracellularly. Secreted Hsp70 is for example involved in cross-presentation of cancer......-derived antigenic peptides, a function which is currently explored in immunotherapeutic approaches against cancer. Additionally, membrane-bound Hsp70 can stimulate antigen presenting cells to release proinflammatory cytokines and can provide a target structure for NK cell-mediated lysis. Human cancer cells...... frequently express Hsp70 on their cell surface, whereas the corresponding normal tissues do not. In addition, several clinically applied reagents, such as alkyl-lysophospholipides, chemotherapeutic agents, and anti-inflammatory reagents, have been found to enhance Hsp70 cell surface expression on cancer...

  12. A mathematical model of the hsp70 regulation in the cell.

    Science.gov (United States)

    Peper, A; Grimbergen, C A; Spaan, J A; Souren, J E; van Wijk, R

    1998-01-01

    A mathematical model of the regulation process of the heat shock protein hsp70 in the cell is presented. The model describes the damaging effect of elevated temperature on proteins; the interaction of free hsp70 with injured proteins and its chaperone role in nascent protein translation; the relation between the amount of free hsp70 and the formation of the activated trimer form of the heat shock factor protein (HSF); the binding of activated HSF with the heat shock elements on the DNA; the transcription of mRNA of hsp70 and the synthesis of hsp70. The reaction of the model to a temporal rise in temperature shows an initial decline and a subsequent sharp rise to an ultimately increased level of free hsp70 in the cell. The response of the model to both a single and two consecutive heat shocks appears to closely resemble experimental data on hsp70 synthesis. This general agreement demonstrates the structure of the model to be sound and suitable as a basis for further modelling the complex tolerance mechanism of the cell. PMID:9483450

  13. Cloning and expression analysis of a HSP70 gene from Korean rockfish (Sebastes schlegeli).

    Science.gov (United States)

    Mu, Weijie; Wen, Haishen; Li, Jifang; He, Feng

    2013-10-01

    The gene encoding HSP70 was isolated from Korean rockfish Sebastes schlegeli by homologous cloning and rapid amplification of cDNA ends (RACE). The full-length of HSP70 cDNA was composed of 2259 bp and encoded a polypeptide of 639 amino acids. BLAST analysis showed that HSP70 of S. schlegeli shared high identities with those of the Lates calcarifer, Oreochromis niloticus, Seriola quinqueradiata HSP70s (88-89%). Our current study also revealed that HSP70 of Korean rockfish was expressed in many tissues by RT-PCR under unstressed condition. Quantitative real-time PCR showed that the expression patterns of Korean rockfish HSP70 were developmental stage-dependency. The expression of HSP70 was measured by quantitative real-time PCR after different oxygen treatments. The results showed that expression of HSP70 increased significantly after exposure to hypoxia for 30 min in gill and ovary, and then decreased for 60 min, and the level in spleen and liver gradually increased and reached the highest at 60 min. In addition, in gill, spleen and liver, the HSP70 mRNA level reached the maximum in hypoxia group after one hour different oxygen concentration stress. Increased amounts of serum thyroxine (T4), and triiodothyronine (T3) were also found during 30 min hypoxia treatment and 60 min normoxia group in our study. All of the results provide information to further study the mechanism of physiology and immune function under stress conditions of ovoviviparous teleosts. PMID:23877000

  14. The Effects of Venlafaxine and Dexamethasone on the Expression of HSP70 in Rat C6 Glioma Cells

    OpenAIRE

    Yu, Jaehak; Roh, Sungwon; Lee, Jun-Seok; Yang, Byung-Hwan; Choi, Mi Ran; Chai, Young Gyu; Kim, Seok Hyeon

    2010-01-01

    Objective The present study aimed to determine the intracellular action of the antidepressant, venlafaxine, in C6 glioma cells using heat shock protein 70 (HSP70) immunocytochemistry and HSP70 Western blots; HSP70 is known to be associated with stress and depression. Methods The extent of HSP70 expression was measured after rat C6 glioma cells were treated with 1) dexamethasone only, 2) venlafaxine only, 3) simultaneous venlafaxine and dexamethasone, or 4) dexamethasone after venlafaxine pret...

  15. Study on HSP70 Gene Expression in Different Tissue of Cyprinus carpio%鲤鱼HSP70基因组织表达差异研究

    Institute of Scientific and Technical Information of China (English)

    林亚秋; 李瑞文

    2009-01-01

    [Objective] The aim of this study was to investigate whether HSP70 can be used as a stress monitoring indicator in Cyprinus carpio breeding. [Method] Based on HSP70 sequence of Cyprinus carpio (AY120894), one pair of primers was designed and synthesized, while the total RNA of liver tissues in Cyprinus carpio was extracted. Some cDNA fragments of Cyprinus carpio HSP70 were cloned by RT-PCR, and its differential expression in various tissues such as heart, intestine, mucus, gonad, swim bladder, gill and fin in Cyprinus carpio was also studied. [Result] The cDNA sequence of 480 bp was obtained from Cyprinus carpio HSP70 gene by RT-PCR amplification. Homology comparison between the deduced amino acid sequence after sequencing and that of other types of fish showed that the homology among Cyprinus carpio, Danio rerio, Ohcorhynehus mylciss, Paralichthys olivaceus, Xiphophoorus maculates and Carassius auratus was 96%, 98%, 98%, 96%, 96% and 96% respectively. The expression of HSP70 was detected in eight tissues of Cyprinus carpio. The expression was the highest in heart, followed by swim bladder and fin, but there was no significant difference between them (P>0.05). There was no significant difference among the expression in three tissues of intestine, mucus and fat (P>0.05), but their expression was significantly higher than those in gonad and gill (P<0.05). [Conclusion] HSP70 gene expression is a suitable criterion for monitoring the stress degree, stress capacity and healthy conditions in Cyprinus carpio breeding.

  16. Citrus Functional Genomics and Molecular Modeling in Relation to Citrus sinensis (Sweet Orange) Infection with Xylella fastidiosa (Citrus Variegated Chlorosis).

    Science.gov (United States)

    Dwivedi, Upendra N; Tiwari, Sameeksha; Prasanna, Pragya; Awasthi, Manika; Singh, Swati; Pandey, Veda P

    2016-08-01

    Citrus are among the economically most important fruit tree crops in the world. Citrus variegated chlorosis (CVC), caused by Xylella fastidiosa infection, is a serious disease limiting citrus production at a global scale. With availability of citrus genomic resources, it is now possible to compare citrus expressed sequence tag (EST) data sets and identify single-nucleotide polymorphisms (SNPs) within and among different citrus cultivars that can be exploited for citrus resistance to infections, citrus breeding, among others. We report here, for the first time, SNPs in the EST data sets of X. fastidiosa-infected Citrus sinensis (sweet orange) and their functional annotation that revealed the involvement of eight C. sinensis candidate genes in CVC pathogenesis. Among these genes were xyloglucan endotransglycosylase, myo-inositol-1-phosphate synthase, and peroxidase were found to be involved in plant cell wall metabolism. These have been further investigated by molecular modeling for their role in CVC infection and defense. Molecular docking analyses of the wild and the mutant (SNP containing) types of the selected three enzymes with their respective substrates revealed a significant decrease in the binding affinity of substrates for the mutant enzymes, thus suggesting a decrease in the catalytic efficiency of these enzymes during infection, thereby facilitating a favorable condition for infection by the pathogen. These findings offer novel agrigenomics insights in developing future molecular targets and strategies for citrus fruit cultivation in ways that are resistant to X. fastidiosa infection, and by extension, with greater harvesting efficiency and economic value. PMID:27447755

  17. 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.

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

    OpenAIRE

    Andreasen, Anne Sofie; Pedersen-Skovsgaard, Theis; Mortensen, Ole Hartvig; van Hall, Gerrit; Moseley, Pope Lloyd; Pedersen, Bente Klarlund

    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 during endotoxin challenge and measured plasma cytokines and HSP70 protein expression. Methods This crossover study, conducted in eight healthy young men, was double-blind, randomized and placebo-controll...

  19. Using Lymphocyte and Plasma Hsp70 as Biomarkers for Assessing Coke Oven Exposure among Steel Workers

    OpenAIRE

    Yang, Xiaobo; Zheng, Jinping; Bai, Yun; Tian, Fengjie; Yuan, Jing; Sun, Jianya; Liang, Huashan; Guo, Liang; Tan, Hao; Chen, Weihong; Tanguay, Robert M.; Wu, Tangchun

    2007-01-01

    Background Hsp70, an early-response protein induced when organisms are confronted with simple or complicated environmental stresses, can act as either a cellular protector or a danger signal. Objectives The goal of this study was to evaluate levels of lymphocyte and/or plasma Hsp70 as biomarkers for assessing exposure response to complex coke oven emissions (COEs). Methods We recruited 101 coke oven workers and determined levels of polycyclic aromatic hydrocarbon (PAH) exposure, urinary 1-hyd...

  20. HSP70 overexpression may play a protective role in the mouse embryos stimulated by CUMS

    OpenAIRE

    Li, Xiao-Hong; Pang, Hou-Qing; QIN, LANG; Jin, Song; Zeng, Xun; Bai, Yu; Li, Shang-Wei

    2015-01-01

    Background We evaluated whether heat shock protein HSP70 plays a protective role in the embryos of Kunming mice subjected to chronic unpredictable mild stress. Methods Female mice were stimulated for 4 weeks with nine stressors and then divided into mild, moderate and severe stress groups. Superovulation was induced with a gonadotropin preparation (PMSG/HCG) and HSP70 expression in 2-cell embryos and day 4 embryos was detected by immunofluorescence (IF) and real-time polymerase chain reaction...

  1. Research Progress of HSP70 and Its Effects in Exercises%热休克蛋白70及其在运动中作用研究进展

    Institute of Scientific and Technical Information of China (English)

    郑妩媚; 王福文

    2015-01-01

    Heat shock protein 70 (HSP70) is a molecular chaperone that is highly expressed in response to stress and closely related to exercises. HSP70 plays an important role in body movement, and is considered as a new endogenous protection approach for myocardium in recent years. The research progress in biological characteristics of HSP70 and its protective effect on myocardial damage induced by exercises is reviewed in this paper, to provide basis for reasonable training in exercises.%热休克蛋白70(heat shock protein 70,HSP70)是一种在应激条件下诱导高表达的伴侣蛋白,与运动关系密切,在机体运动过程中起重要作用。近年,HSP70已成为较受重视的心肌内源性保护途径之一。本文主要介绍了HSP70的生物学特性及其对运动性心肌损伤保护作用的研究进展,以冀为合理进行运动训练提供依据。

  2. 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.

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

    Science.gov (United States)

    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. PMID:27094592

  4. Effect of betaine on HSP70 expression and cell survival during adaptation to osmotic stress.

    Science.gov (United States)

    Petronini, P G; De Angelis, E M; Borghetti, A F; Wheeler, K P

    1993-07-15

    Induced expression of the HSP70 gene in 3T3 and SV-3T3 cells was monitored by measurements of the synthesis of HSP70 and of the cellular contents of both HSP70 and its mRNA. The presence of betaine (N-trimethylglycine) at concentrations of 2.5-25 mM decreased the induction of HSP70 gene expression caused by incubation of 3T3 and SV-3T3 cells in hypertonic (0.5 osM) medium. This effect was accompanied by an enhancement of SV-3T3 cell adaptation, assayed by colony formation, to the hyperosmotic conditions. In contrast, the presence of betaine did not affect HSP70 gene expression induced in these cells by heat shock. After 6 h incubation with 25 mM betaine under hypertonic (0.5 osM) conditions the intracellular concentration of betaine in SV-3T3 cells was about 195 mM, compared with about 70 mM under isotonic (0.3 osM) conditions. Hence, with this concentration of extracellular betaine, the marked increase in the accumulation of betaine within the cells presumably counteracts the imposed osmotic pressure and eliminates the signal that otherwise initiates increased expression of the HSP70 gene. PMID:8343134

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

    Science.gov (United States)

    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.

  6. Heat shock proteins in Trypanosoma cruzi: identification and localization of HSP70 and HSP60 proteins and structure of HSP60 genes (brief report).

    Science.gov (United States)

    de Marval, M G; Souto-Padron, T; Gottesdiener, K; Silva, R; van der Ploeg, L H; Rondinelli, E

    1993-01-01

    To identify the members of the HSP70 and HSP60 families of Trypanosoma cruzi, we analysed 35S methionine epimastigote cells by two dimensional Western blot. At 29 degrees C, an HSP70 monoclonal antibody (anti-D. melanogaster) recognized eight isotypes. At least five of these were heat-induced. Polyclonal antibody against the 65 KDa antigen (anti-M. tuberculosis) recognized three isotypes with identical molecular weights, but different microliters. Only one isoform was heat induced. The cellular distribution of HSP70 and HSP60 was studied by immunoelectron microscopy. Anti-HSP70 reactive protein was localized in the cytoplasm, mitochondria and nucleus, while anti-HSP60 protein was found in the mitochondrion and in close association with the kinetoplast. To characterize the HSP60 gene and its proteins, we isolated a genomic T. cruzi clone encoding the HSP60 gene. T. cruzi HSP60 genes could be shown to be organized in 2100 nt tandem arrays. RELP in the HSP60 genes revealed that at least three different types of HSP60 genes were encoded in the T cruzi genome. The predicted open reading frame measured exhibits about 50% identity to other HSP60 described. Expression of these HSP60 genes could not be induced by 2 hours heat shock at 37 degrees C. Post-transcriptional mechanisms may be responsible for HSP60 induction in T. cruzi. PMID:7670543

  7. Identification of an allosteric pocket on human hsp70 reveals a mode of inhibition of this therapeutically important protein.

    Science.gov (United States)

    Rodina, Anna; Patel, Pallav D; Kang, Yanlong; Patel, Yogita; Baaklini, Imad; Wong, Michael J H; Taldone, Tony; Yan, Pengrong; Yang, Chenghua; Maharaj, Ronnie; Gozman, Alexander; Patel, Maulik R; Patel, Hardik J; Chirico, William; Erdjument-Bromage, Hediye; Talele, Tanaji T; Young, Jason C; Chiosis, Gabriela

    2013-12-19

    Hsp70s are important cancer chaperones that act upstream of Hsp90 and exhibit independent anti-apoptotic activities. To develop chemical tools for the study of human Hsp70, we developed a homology model that unveils a previously unknown allosteric site located in the nucleotide binding domain of Hsp70. Combining structure-based design and phenotypic testing, we discovered a previously unknown inhibitor of this site, YK5. In cancer cells, this compound is a potent and selective binder of the cytosolic but not the organellar human Hsp70s and has biological activity partly by interfering with the formation of active oncogenic Hsp70/Hsp90/client protein complexes. YK5 is a small molecule inhibitor rationally designed to interact with an allosteric pocket of Hsp70 and represents a previously unknown chemical tool to investigate cellular mechanisms associated with Hsp70. PMID:24239008

  8. 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

  9. Induction of heat shock protein 70 (Hsp70 prevents neuregulin-induced demyelination by enhancing the proteasomal clearance of c-Jun

    Directory of Open Access Journals (Sweden)

    Rick T Dobrowsky

    2012-12-01

    Full Text Available Modulating molecular chaperones is emerging as an attractive approach to treat neurodegenerative diseases associated with protein aggregation, DPN (diabetic peripheral neuropathy and possibly, demyelinating neuropathies. KU-32 [N-(7-((2R,3R,4S,5R-3,4-dihydroxy-5-methoxy-6,6-dimethyl-tetrahydro-2H-pyran-2-yloxy-8-methyl-2-oxo-2H-chromen-3-ylacetamide] is a small molecule inhibitor of Hsp90 (heat shock protein 90 and reverses sensory deficits associated with myelinated fibre dysfunction in DPN. Additionally, KU-32 prevented the loss of myelinated internodes induced by treating myelinated SC (Schwann cell-DRG (dorsal root ganglia sensory neuron co-cultures with NRG1 (neuregulin-1 Type 1. Since KU-32 decreased NRG1-induced demyelination in an Hsp70-dependent manner, the goal of the current study was to clarify how Hsp70 may be mechanistically linked to preventing demyelination. The activation of p42/p44 MAPK (mitogen-activated protein kinase and induction of the transcription factor c-Jun serve as negative regulators of myelination. NRG1 activated MAPK, induced c-Jun expression and promoted a loss of myelin segments in DRG explants isolated from both WT (wild-type and Hsp70 KO (knockout mice. Although KU-32 did not block the activation of MAPK, it blocked c-Jun induction and protected against a loss of myelinated segments in WT mice. In contrast, KU-32 did not prevent the NRG1-dependent induction of c-Jun and loss of myelin segments in explants from Hsp70 KO mice. Overexpression of Hsp70 in myelinated DRG explants prepared from WT or Hsp70 KO mice was sufficient to block the induction of c-Jun and the loss of myelin segments induced by NRG1. Lastly, inhibiting the proteasome prevented KU-32 from decreasing c-Jun levels. Collectively, these data support that Hsp70 induction is sufficient to prevent NRG1-induced demyelination by enhancing the proteasomal degradation of c-Jun.

  10. Effects of aspartame on hsp70, bcl-2 and bax expression in immune organs of Wistar albino rats

    Science.gov (United States)

    Choudhary, Arbind Kumar; Devi, Rathinasamy Sheela

    2016-01-01

    Abstract Aspartame, a “first generation sweetener”, is widely used in a variety of foods, beverages, and medicine. The FDA has determined the acceptable daily intake (ADI) value of aspartame to be 50 mg/kg·day, while the JECFA (Joint FAO/WHO Expert Committee on Food Additives) has set this value at 40 mg/kg of body weight/day. Safety issues have been raised about aspartame due to its metabolites, specifically toxicity from methanol and/or its systemic metabolites formaldehyde and formic acid. The immune system is now recognized as a target organ for many xenobiotics, such as drugs and chemicals, which are able to trigger unwanted apoptosis or to alter the regulation of apoptosis. Our previous studies has shown that oral administration of aspartame [40 mg/(kg·day)] or its metabolites for 90 days increased oxidative stress in immune organs of Wistar albino rats. In this present study, we aimed to clarify whether aspartame consumption over a longer period (90-days) has any effect on the expression of hsp70, bcl-2 and bax at both mRNA transcript and protein expression levels in immune organs. We observed that oral administration of aspartame for 90 days did not cause any apparent DNA fragmentation in immune organs of aspartame treated animals; however, there was a significant increase in hsp70 expression, apart from significant alteration in bcl-2 and bax at both mRNA transcript and protein expression level in the immune organs of aspartame treated animals compared to controls. Hence, the results indicated that hsp70 levels increased in response to oxidative injury induced by aspartame metabolites; however, these metabolites did not induce apoptosis in the immune organs. Furthermore, detailed analyses are needed to elucidate the precise molecular mechanisms involved in these changes.

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

    Directory of Open Access Journals (Sweden)

    G. Tomasello

    2011-10-01

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

  12. A Surveillance Function of the HSPB8-BAG3-HSP70 Chaperone Complex Ensures Stress Granule Integrity and Dynamism.

    Science.gov (United States)

    Ganassi, Massimo; Mateju, Daniel; Bigi, Ilaria; Mediani, Laura; Poser, Ina; Lee, Hyun O; Seguin, Samuel J; Morelli, Federica F; Vinet, Jonathan; Leo, Giuseppina; Pansarasa, Orietta; Cereda, Cristina; Poletti, Angelo; Alberti, Simon; Carra, Serena

    2016-09-01

    Stress granules (SGs) are ribonucleoprotein complexes induced by stress. They sequester mRNAs and disassemble when the stress subsides, allowing translation restoration. In amyotrophic lateral sclerosis (ALS), aberrant SGs cannot disassemble and therefore accumulate and are degraded by autophagy. However, the molecular events causing aberrant SG formation and the molecular players regulating this transition are largely unknown. We report that defective ribosomal products (DRiPs) accumulate in SGs and promote a transition into an aberrant state that renders SGs resistant to RNase. We show that only a minor fraction of aberrant SGs is targeted by autophagy, whereas the majority disassembles in a process that requires assistance by the HSPB8-BAG3-HSP70 chaperone complex. We further demonstrate that HSPB8-BAG3-HSP70 ensures the functionality of SGs and restores proteostasis by targeting DRiPs for degradation. We propose a system of chaperone-mediated SG surveillance, or granulostasis, which regulates SG composition and dynamics and thus may play an important role in ALS. PMID:27570075

  13. Changes of levels of hs-CRP, Hsp70, and MPO in patients with preeclampsia

    Institute of Scientific and Technical Information of China (English)

    Xia Liu

    2016-01-01

    Objective:To observe the changes of the levels of hs-CRP, Hsp70, and MPO in patients with preeclampsia (PE) and their clinical significance. Methods:A total of 60 pregnant women with mild and severe PE who were admitted in our hospital from January, 2013 to March, 2014 were included in the study and divided into the mild group and the severe group with 30 cases in each group. The levels of serum hs-CRP, Hsp70, and MPO were detected, and their relations with PE were analyzed. The lipid metabolism related indicators were meanwhile detected. A total of 30 healthy later pregnant women at the same stage were served as the control group for comparative analysis. Results:The levels of serum hs-CRP, Hsp70, and MPO in the mild and severe PE group were significantly higher than those in the control group (P0.05). Conclusions: The abnormal lipid metabolism in different degrees exists in the pregnant women with PE, and hs-CRP, Hsp70, and MPO are together involved in the pathogenesis of PE. Detection of the levels of hs-CRP, Hsp70, and MPO in the pregnant women with PE can accurately reflect the condition of PE.

  14. Induction of hsp70 by the herbicide oxyfluorfen (Goal) in the Egyptian Nile fish, Oreochromis niloticus.

    Science.gov (United States)

    Hassanein, H M; Banhawy, M A; Soliman, F M; Abdel-Rehim, S A; Müller, W E; Schröder, H C

    1999-07-01

    This paper deals with the expression of the biomarker hsp70 in the liver and kidney of the freshwater fish Oreochromis niloticus following exposure to the herbicide oxyfluorfen (Goal). Fishes were exposed to three concentrations, the 96-h LC50 (3 mg/L), the 96-h (1/2)LC50 (1.5 mg/L), and the 96-h (1/4)LC50 (0.75 mg/L) of oxyfluorfen for 6, 15, and 24 days, respectively, and samples were taken at three different time periods for each concentration. The livers responded to the herbicide by an induction of the expression of both the constitutive (hsp75; Mr 75 kDa) and the inducible (hsp73; Mr 73 kDa) hsp70 proteins. In kidney, the herbicide induced a time-dependent increase in the expression of the constitutive hsp70 (hsp75) as well, but the inducible hsp70 (hsp73) required much longer incubation periods to reach maximal levels (15 and 24 days). Our results suggest that expression of hsp70 in fish is a sensitive indicator of cellular responses to herbicide exposure in the aquatic environment. PMID:10341045

  15. 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.

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

    International Nuclear Information System (INIS)

    DNA double strand breaks (DSBs) in living cells can be directly provoked by ionising radiation. DSBs can be visualized by immunostaining the phosphorylated histone γH2AX. Our concern was to test the feasibility of γH2AX 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 γH2AX 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 Si3N4 window showed a homogenous Hsp70 expression pattern

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

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    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.

  18. A Novel Protein Interaction between Nucleotide Binding Domain of Hsp70 and p53 Motif

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    Asita Elengoe

    2015-01-01

    Full Text Available Currently, protein interaction of Homo sapiens nucleotide binding domain (NBD of heat shock 70 kDa protein (PDB: 1HJO with p53 motif remains to be elucidated. The NBD-p53 motif complex enhances the p53 stabilization, thereby increasing the tumor suppression activity in cancer treatment. Therefore, we identified the interaction between NBD and p53 using STRING version 9.1 program. Then, we modeled the three-dimensional structure of p53 motif through homology modeling and determined the binding affinity and stability of NBD-p53 motif complex structure via molecular docking and dynamics (MD simulation. Human DNA binding domain of p53 motif (SCMGGMNR retrieved from UniProt (UniProtKB: P04637 was docked with the NBD protein, using the Autodock version 4.2 program. The binding energy and intermolecular energy for the NBD-p53 motif complex were −0.44 Kcal/mol and −9.90 Kcal/mol, respectively. Moreover, RMSD, RMSF, hydrogen bonds, salt bridge, and secondary structure analyses revealed that the NBD protein had a strong bond with p53 motif and the protein-ligand complex was stable. Thus, the current data would be highly encouraging for designing Hsp70 structure based drug in cancer therapy.

  19. Detection of constitutive and inducible HSP70 proteins in formalin fixed human brain tissue.

    Science.gov (United States)

    Preusse-Prange, A; Modrow, J-H; Schwark, T; von Wurmb-Schwark, N

    2014-02-01

    The investigation of formalin fixed and paraffin embedded tissue is a routine method in forensic histology. Since these samples are usually stored for decades they provide a unique tissue bank for different scientific issues. In the past, numerous studies were conducted using different kinds of paraffin embedded tissues. However, it is well known that formalin affects macromolecules and thus might hamper reliable and reproducible molecular experiments. The aim of this study was to find out if the treatment with formalin has a negative effect on different protein detection methods and additionally to define the dimension of those possible deleterious effects. We incubated brain tissue samples in formalin for up to three months. After incubation, the samples were analyzed using immunohistochemistry (IHC) and Western blotting to specifically detect and quantify members of the HSP70 superfamily (heat shock proteins). Our study shows that the Western blot analysis of formalin fixed tissues does not allow a reliable detection of proteins at all, while a reproducible detection by IHC was still possible after one month of incubation.

  20. Protective Effects of Overexpression TCR Vβ5.2-HSP70 and TCR Vβ8.2-HSP70 against Collagen-Induced Arthritis in Rats

    Institute of Scientific and Technical Information of China (English)

    Jing Xiao; Shentao Li; Wei Wang; Yun Li; Wenming Zhao

    2007-01-01

    Collagen-induced arthritis (CIA) is an animal model, which closely resembles human rheumatoid arthritis (RA) in pathogenesis and pathology. Evidence suggests that the inhibition of T lymphocytes or their functions can alleviate the progression of arthritis. So the administration of arthritogenic T cell receptor (TCR) variable region peptide or DNA vaccines encoding pathogenic TCR Vβ variable region may provide useful information for designing specific immunotherapies against autoimmune diseases. Heat shock proteins (HSPs) have the function of raising antigenic immunogenicity and HSP70 has a protective effect against arthritis. We previously demonstrated the presence of pathogenic predominant T cell receptor Vβ5.2 and Vβ8.2 clonotypes in the joints of CIA rats. In this study, we constructed the recombinant eukaryotic expression vectors pTARGET-TCR Vβ5.2/8.2-HSP70, and evaluated their protective effects on CIA rats. Protective effects were observed in CIA rats by injecting these recombinant DNA vaccines, which could alleviate arthritis index, decrease the levels of IFN-γ and anti-CⅡ antibody in serum, and increase the levels of IL-4. Pathological changes were not as serious as those observed in control CIA rats. The rat injected with two combined vaccines showed better protective effects than CIA rats administered with individual vaccine. These results showed that recombinant DNA vaccines pTARGET-TCR Vβ5.2-HSP70 and pTARGET-TCR Vβ8.2-HSP70 could significantly alleviate the arthritic symptoms of CIA rats, and better protective effects could be achieved if these two vaccines were used in combination.

  1. Functional analysis of Drosophila HSP70 promoter with different HSE numbers in human cells.

    Science.gov (United States)

    Kust, Nadezda; Rybalkina, Ekaterina; Mertsalov, Ilya; Savchenko, Ekaterina; Revishchin, Alexander; Pavlova, Gali

    2014-01-01

    The activation of genetic constructs including the Drosophila hsp70 promoter with four and eight HSE sequences in the regulatory region has been described in human cells. The promoter was shown to be induced at lower temperatures compared to the human hsp70 promoter. The promoter activity increased after a 60-min heat shock already at 38 °C in human cells. The promoter activation was observed 24 h after heat shock for the constructs with eight HSEs, while those with four HSEs required 48 h. After transplantation of in vitro heat-shocked transfected cells, the promoter activity could be maintained for 3 days with a gradual decline. The promoter activation was confirmed in vivo without preliminary heat shock in mouse ischemic brain foci. Controlled expression of the Gdnf gene under a Drosophila hsp70 promoter was demonstrated. This promoter with four and eight HSE sequences in the regulatory region can be proposed as a regulated promoter in genetic therapeutic systems.

  2. PP2A mediated AMPK inhibition promotes HSP70 expression in heat shock response.

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    Ting Wang

    Full Text Available BACKGROUND: Under stress, AMP-activated protein kinase (AMPK plays a central role in energy balance, and the heat shock response is a protective mechanism for cell survival. The relationship between AMPK activity and heat shock protein (HSP expression under stress is unclear. METHODOLOGY/PRINCIPAL FINDINGS: We found that heat stress induced dephosphorylation of AMPKα subunit (AMPKα in various cell types from human and rodent. In HepG2 cells, the dephosphorylation of AMPKα under heat stress in turn caused dephosphorylation of acetyl-CoA carboxylase and upregulation of phosphoenolpyruvate carboxykinase, two downstream targets of AMPK, confirming the inhibition of AMPK activity by heat stress. Treatment of HepG2 cells with phosphatase 2A (PP2A inhibitor okadaic acid or inhibition of PP2A expression by RNA interference efficiently reversed heat stress-induced AMPKα dephosphorylation, suggesting that heat stress inhibited AMPK through activation of PP2A. Heat stress- and other HSP inducer (CdCl(2, celastrol, MG132-induced HSP70 expression could be inhibited by AICAR, an AMPK specific activator. Inhibition of AMPKα expression by RNA interference reversed the inhibitory effect of AICAR on HSP70 expression under heat stress. These results indicate that AMPK inhibition under stress contribute to HSP70 expression. Mechanistic studies showed that activation of AMPK by AICAR had no effect on heat stress-induced HSF1 nuclear translocation, phosphorylation and binding with heat response element in the promoter region of HSP70 gene, but significantly decreased HSP70 mRNA stability. CONCLUSIONS/SIGNIFICANCE: These results demonstrate that during heat shock response, PP2A mediated AMPK inhibition upregulates HSP70 expression at least partially through stabilizing its mRNA, which suggests a novel mechanism for HSP induction under stress.

  3. Echinococcus granulosus: in vitro effects of ivermectin and praziquantel on hsp60 and hsp70 levels.

    Science.gov (United States)

    Martinez, J; Perez-Serrano, J; Bernadina, W E; Rodriguez-Caabeiro, F

    1999-11-01

    Martinez, J., Perez-Serrano, J., Bernadina, W. E., Rodriguez-Caabeiro, F. 1999 Echinococcus granulosus: In vitro effects of ivermectin and praziquantel on hsp60 and hsp70 levels. Experimental Parasitology93, 171-180. Organisms or cells exposed to injurious stresses such as heat shock or chemicals respond by increased (or altered) expression of heat-shock proteins (HSPs). Conversely, an earlier exposure to stress can prepare cells to cope with a subsequent more severe stress. In the present study, protoscolices of Echinococcus granulosus were subjected to several anthelmintic treatments, involving storage of the protoscolices for 18, 30, and 50 h with 0.1 mg/ml of ivermectin (IV), praziquantel (PZ), and a combination of each with albendazole (ALB). The organisms were analyzed for the effects of drug treatment on cell integrity and on levels of hsp60 and hsp70 production. Drug efficacy was evaluated by microscopy and by protein content measurement. Hsp60 and hsp70 were detected by Western blotting and incubation with anti-hsp60 and anti-hsp70 antibody, respectively, and quantitation of these proteins was obtained using image analysis. Incubation with IV alone produced the most damage to the protoscolices as indicated by viability loss, decreased protein content, and altered hsp60 and hsp70 levels; incubation with IV + ALB produced less damage as manifested by fewer changes in the aforementioned damage parameters but PZ and PZ + ALB, in this context, were poor anthelmintics. Exposure of protoscolices to thermal stress prior to anthelmintic treatment, in most cases, increased drug efficacy. It is concluded that in the E. granulosus model system drug efficacy is associated with decreased levels of hsp70 expression and increased levels of hsp60 expression.

  4. Peptide deformylase inhibitor actinonin reduces celastrol’s HSP70 induction while synergizing proliferation inhibition in tumor cells

    International Nuclear Information System (INIS)

    Celastrol is a promising anti-tumor agent, yet it also elevates heat shock proteins (HSPs), especially HSP70, this effect believed to reduce its anti-tumor effects. Concurrent use of siRNA to increase celastrol’s anti-tumor effects through HSP70 interference has been reported, but because siRNA technology is difficult to clinically apply, an alternative way to curb unwanted HSP70 elevation caused by celastrol treatment is worth exploring. In this work, we explore three alternative strategies to control HSP70 elevation: (1) Searching for cancer cell types that show no HSP70 elevation in the presence of celastrol (thus recommending themselves as suitable targets); (2) Modifying HSP70-inducing chemical groups, i.e.: the carboxyl group in celastrol; and (3) Using signaling molecule inhibitors to specifically block HSP70 elevation while protecting and/or enhancing anti-tumor effects. The first strategy was unsuccessful since celastrol treatment increased HSP70 in all 7 of the cancer cell types tested, this result related to HSF1 activation. The ubiquity of HSF1 expression in different cancer cells might explain why celastrol has no cell-type limitation for HSP70 induction. The second strategy revealed that modification of celastrol’s carboxyl group abolished its ability to elevate HSP70, but also abolished celastrol’s tumor inhibition effects. In the third strategy, 11 inhibitors for 10 signaling proteins reportedly related to celastrol action were tested, and five of these could reduce celastrol-caused HSP70 elevation. Among these, the peptide deformylase (PDF) inhibitor, actinonin, could synergize celastrol’s proliferation inhibition. Concurrent use of the chemical agent actinonin could reduce celastrol’s HSP70 elevation and also enhance proliferation inhibition by celastrol. This combination presents a novel alternative to siRNA technology and is worth further investigation for its potentially effective anti-tumor action

  5. Effects of HSP70 Antisense Oligonucleotide on the Proliferation and Apoptosis of Human Hepatocellular Carcinoma Cells

    Institute of Scientific and Technical Information of China (English)

    杨雪; 贺海斌; 杨威; 宋涛; 郭成; 郑鑫; 刘青光

    2010-01-01

    The study investigated the effects of heat shock protein 70(HSP70) antisense oligonucleotide(ASODN) on the proliferation and apoptosis of a human hepatocellular carcinoma cell line(SMMC-7721 cells) in vitro.HSP70 oligonucleotide was transfected into SMMC-7721 cells by the mediation of SofastTM transfection reagent.Inhibition rate of SMMC-7721 cells was determined by using MTT method.Apoptosis rate and cell cycle distribution were measured by flow cytometry.Immunocytochemistry staining was used to observe th...

  6. 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...

  7. 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

  8. Crystallization and X-ray data analysis of the 10 kDa C-terminal lid subdomain from Caenorhabditis elegans Hsp70

    Energy Technology Data Exchange (ETDEWEB)

    Worrall, Liam; Walkinshaw, Malcolm D., E-mail: m.walkinshaw@ed.ac.uk [School of Biological Sciences, University of Edinburgh, The King’s Buildings, Mayfield Road, Edinburgh EH9 3JR,Scotland (United Kingdom)

    2006-09-01

    Crystals of the C-terminal 10 kDa lid subdomain from the C. elegans chaperone Hsp70 have been obtained that diffract X-rays to ∼3.5 Å and belong to space group I2{sub 1}2{sub 1}2{sub 1}. Analysis of X-ray data and initial heavy-atom phasing reveals 24 monomers in the asymmetric unit related by 432 non-crystallographic symmetry. Hsp70 is an important molecular chaperone involved in the regulation of protein folding. Crystals of the C-terminal 10 kDa helical lid domain (residues 542–640) from a Caenorhabditis elegans Hsp70 homologue have been produced that diffract X-rays to ∼3.4 Å. Crystals belong to space group I2{sub 1}2{sub 1}2{sub 1}, with unit-cell parameters a = b = 197, c = 200 Å. The Matthews coefficient, self-rotation function and Patterson map indicate 24 monomers in the asymmetric unit, showing non-crystallographic 432 symmetry. Molecular-replacement studies using the corresponding domain from rat, the only eukaryotic homologue with a known structure, failed and a mercury derivative was obtained. Preliminary MAD phasing using SHELXD and SHARP for location and refinement of the heavy-atom substructure and SOLOMON for density modification produced interpretable maps with a clear protein–solvent boundary. Further density-modification, model-building and refinement are currently under way.

  9. 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

    2008-01-01

    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 CD3(+)/CD16(-)/5

  10. Expression and Localization of an Hsp70 Protein in the Microsporidian Encephalitozoon cuniculi

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    Carrie E. Jolly

    2010-01-01

    Full Text Available Microsporidia spore surface proteins are an important, under investigated aspect of spore/host cell attachment and infection. For comparison analysis of surface proteins, we required an antibody control specific for an intracellular protein. An endoplasmic reticulum-associated heat shock protein 70 family member (Hsp70; ECU02_0100; “C1” was chosen for further analysis. DNA encoding the C1 hsp70 was amplified, cloned and used to heterologously express the C1 Hsp70 protein, and specific antiserum was generated. Two-dimensional Western blotting analysis showed that the purified antibodies were monospecific. Immunoelectron microscopy of developing and mature E. cuniculi spores revealed that the protein localized to internal structures and not to the spore surface. In spore adherence inhibition assays, the anti-C1 antibodies did not inhibit spore adherence to host cell surfaces, whereas antibodies to a known surface adhesin (EnP1 did so. In future studies, the antibodies to the ‘C1’ Hsp70 will be used to delineate spore surface protein expression.

  11. 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 cluster (Fe-S) biogenesis, or protein synthesis. Here we demonstrate that mutations in HSPA9, a mitochondrial HSP70 homologue located in the 5q...

  12. 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

    1996-01-01

    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 constitutiv

  13. Association of eNOS and HSP70 gene polymorphisms with glaucoma in Pakistani cohorts

    NARCIS (Netherlands)

    Ayub, H.; Khan, M.I.; Micheal, S.; Akhtar, F.; Ajmal, M.; Shafique, S.; Ali, S.H.; Hollander, A.I. den; Ahmed, A.; Qamar, R.

    2010-01-01

    PURPOSE: To investigate the involvement of stress-regulating genes, endothelial nitric oxide synthase (eNOS) and heat shock protein 70 (HSP70) with primary open angle glaucoma (POAG) and primary closed angle glaucoma (PCAG). METHODS: POAG and PCAG patients recruited from different areas of Pakistan

  14. Alkannin, HSP70 inducer, protects against UVB-induced apoptosis in human keratinocytes.

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    Yoko Yoshihisa

    Full Text Available Alkannin is an active constituent from the root extract of Alkanna tinctoria of the Boraginaceae family and it may have utility as a heat shock protein 70 (HSP70 inducer in living organisms. Here, the effects of alkannin-induced HSP70 on ultraviolet (UV B (40 mJ/cm(2-induced apoptosis were investigated in human keratinocyte HaCaT cells. Pretreatment of cells with alkannin (1 µM caused significant inhibition of UVB-induced apoptosis and caspase-3 cleavage. On the other hand, the addition of KNK437 (HSP70 inhibitor reversed the action of alkannin increasing UVB-induced apoptosis in a dose-dependent manner. In addition, differences in gene expression associated with the suppression of UVB-induced apoptosis in the presence of alkannin were investigated using Gene Chip assay. Our results indicate that alkannin suppresses UVB-induced apoptosis through the induction of HSP70 in human keratinocytes, and therefore, we suggest the usefulness of using alkannin as an antiaging agent.

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

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    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. 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.

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

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    Jian Ye

    Full Text Available BACKGROUND: 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. OBJECTIVES: We hypothesized that PAHs might induce systemic inflammation and stress response, contributing to altered cardiac autonomic function. METHODS: 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. RESULTS: 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. CONCLUSIONS: 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.

  18. Radioresistance of human glioma spheroids and expression of HSP70, p53 and EGFr

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    Fedrigo Carlos A

    2011-11-01

    Full Text Available Abstract Background Radiation therapy is routinely prescribed for high-grade malignant gliomas. However, the efficacy of this therapeutic modality is often limited by the occurrence of radioresistance, reflected as a diminished susceptibility of the irradiated cells to undergo cell death. Thus, cells have evolved an elegant system in response to ionizing radiation induced DNA damage, where p53, Hsp70 and/or EGFr may play an important role in the process. In the present study, we investigated whether the content of p53, Hsp70 and EGFr are associated to glioblastoma (GBM cell radioresistance. Methods Spheroids from U-87MG and MO59J cell lines as well as spheroids derived from primary culture of tumor tissue of one GBM patient (UGBM1 were irradiated (5, 10 and 20 Gy, their relative radioresistance were established and the p53, Hsp70 and EGFr contents were immunohistochemically determined. Moreover, we investigated whether EGFr-phospho-Akt and EGFr-MEK-ERK pathways can induce GBM radioresistance using inhibitors of activation of ERK (PD098059 and Akt (wortmannin. Results At 5 Gy irradiation UGBM1 and U-87MG spheroids showed growth inhibition whereas the MO59J spheroid was relatively radioresistant. Overall, no significant changes in p53 and Hsp70 expression were found following 5 Gy irradiation treatment in all spheroids studied. The only difference observed in Hsp70 content was the periphery distribution in MO59J spheroids. However, 5 Gy treatment induced a significant increase on the EGFr levels in MO59J spheroids. Furthermore, treatment with inhibitors of activation of ERK (PD098059 and Akt (wortmannin leads to radiosensitization of MO59J spheroids. Conclusions These results indicate that the PI3K-Akt and MEK-ERK pathways triggered by EGFr confer GBM radioresistance.

  19. Distinctive properties of the 5'-untranslated region of human hsp70 mRNA.

    Science.gov (United States)

    Rubtsova, Maria P; Sizova, Daria V; Dmitriev, Sergei E; Ivanov, Dmitri S; Prassolov, Vladimir S; Shatsky, Ivan N

    2003-06-20

    A relaxed cap-dependence of translation of the mRNA-encoding mammalian heat shock protein Hsp70 may suggest that its 5'-untranslated region (UTR) possesses an internal ribosome entry site (IRES). In this study, this possibility has been tested in transfected cells using plasmids that express dicistronic mRNAs. Using a reporter gene construct, Renilla luciferase/Photinus pyralis luciferase, we show that the 216-nt long 5'-UTR of Hsp70 mRNA acts as an IRES that directs ribosomes to the downstream start codon by a cap-independent mechanism. The relative activity of this IRES (100-fold over the empty vector) is similar to that of the classical picornaviral IRESs. Additional controls indicate that this high expression of the downstream reporter is not due to readthrough from the upstream cistron, nor is it due to translation of cryptic monocistronic transcripts. The effect of small deletions within the 5'-UTR of Hsp70 mRNA on the IRES activity varies in dependence on their position within the 5'-UTR sequence. With the exception of deletion of nt 33-50, it is small for the 5'-terminal half of the 5'-UTR and rather strong for the 3'-terminal section. However, neither of these small deletions abolishes the IRES activity completely. Excision of larger sections (>50 nt) by truncation of the 5'-UTR from the 5'-end or by internal deleting results in a dramatic impairment of the IRES function. Taken together, these data suggest that the IRES activity of the 5'-UTR of Hsp70 mRNA requires integrity of almost the entire sequence of the 5'-UTR. The data are discussed in terms of a model that allows a three-dimensional rather than linear mode of selection of the initiation region surrounding the start codon of Hsp70 mRNA.

  20. Evaluation of toxic potential of captan: Induction of hsp70 and tissue damage in transgenic Drosophila melanogaster (hsp70-lacZ) Bg9.

    Science.gov (United States)

    Nazir, Aamir; Mukhopadhyay, Indranil; Saxena, D K; Siddiqui, M Saeed; Chowdhuri, D Kar

    2003-01-01

    The study investigated the working hypothesis that a widely used fungicide captan exerts toxic effects on nontarget organisms. Transgenic Drosophila melanogaster (hsp70-lacZ) was used as a model by assaying stress gene expression as an endpoint for cytotoxicity and also to evaluate whether stress gene expression is sufficient enough to protect and to prevent tissue damage against toxic insult of the chemical. The study was further extended to understand the effect of the pesticide on development, life cycle, and reproduction of the organism and finally to evaluate a concentration of the chemical to be nontoxic to the organism. The study showed that (i) captan causes cytotoxicity at and above 0.015 ppm; (ii) at 0.0015 ppm captan, absence of hsp70 expression in the exposed organism was evaluated as the concentration referred to as no observed adverse effect level (NOAEL) for Drosophila; (iii) emergence pattern of flies was affected only at the highest concentration of captan by 4 days, while hatching and survivorship were unaffected even at this concentration; (iv) reproductive performance was significantly affected only at 125.0 and 1250.0 ppm captan, while in the lower dietary concentrations no such deleterious effects were observed; (v) at 1250.0 ppm, hsp70 failed to protect the cells from toxicant assault after 48 h exposure, thus leading to tissue damage as revealed by Trypan Blue staining. The present study shows the cytotoxic potential of captan and further reveals the application of stress genes in determining NOAEL and its expression as bioindicator of exposure to environmental contaminants.

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

    Directory of Open Access Journals (Sweden)

    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.

  2. 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

  3. 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

    Directory of Open Access Journals (Sweden)

    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.

  4. Trypanosoma carassii hsp70 increases expression of inflammatory cytokines and chemokines in macrophages of the goldfish (Carassius auratus L.).

    Science.gov (United States)

    Oladiran, Ayoola; Belosevic, Miodrag

    2009-10-01

    We report on the cloning and characterization of Trypanosoma carassii 70 KDa heat shock protein (hsp70). T. carassii hsp70 was secreted/excreted into culture medium in vitro and was recognized by sera from infected fish. Recombinant hsp70 (rhsp70) activated goldfish macrophages and stimulated the production of pro-inflammatory cytokines including interferon gamma (IFNgamma), tumor necrosis factor (TNF)-alpha, interleukin (IL)-1beta, (IL)-12 and chemokines CCL-1 and CXCL-8 (IL-8). T. carassii hsp70-induced cytokine expression was abrogated by pronase treatment of macrophages confirming the existence of receptor(s) on goldfish macrophage surface that recognize parasite molecule. Parasite hsp70 also up-regulated the expression inducible nitric oxide synthase (iNOS) isoforms A and B and induced a strong nitric oxide response of goldfish macrophages. PMID:19527750

  5. Study of the role of Hsp70.1 in oridonin-treated hepatocarcinoma cells%Hsp70.1在冬凌草甲素处理的肝癌细胞中的作用研究

    Institute of Scientific and Technical Information of China (English)

    王辉; 叶燕; 黄树林; 禹志领

    2011-01-01

    目的 研究Hsp70.1在冬凌草甲素处理后的肝癌细胞HepG2中的表达情况及其与冬凌草甲素抗肿瘤活性的关系.方法 采用蛋白印迹法及实时荧光定量PCR技术观察Hsp70.1在冬凌草甲素处理的肝癌细胞HepG2的表达变化,并采用RNA干扰技术观察抑制Hsp70.1表达后对冬凌草甲素抗肝癌效果的影响.结果 冬凌草甲素作用12 h后,HepG2细胞的Hsp70.1蛋白及RNA表达均增加.Hsp70.1短发夹RNA(shRNA)转染抑制Hsp70.1表达后,与对照组相比,Hsp70.1 shRNA转染组、冬凌草甲素处理组或两者联合应用组的HepG2细胞的存活率均明显降低.结论 Hsp70.1可促进HepG2细胞的生存,对HepG2细胞和冬凌草甲素处理的HepG2细胞具有保护作用.%Objective To explore the expression of Hsp70. 1 in oridonin-treated hepatocarcinoma HepC2 cells and the correlation of Hsp70. 1 with the anticancer activity of oridonin. Methods By using Western blot and real-time PCR analysis,we tested the expression of Hsp70. 1 protein and mRNA in HepG2 cells treated with oridonin. After HepG2 cells were transfected with Hsp70.1 shRNA.the influence of Hsp70.1 on the anticancer activity of oridonin was investigated. Results The levels of Hsp70. 1 protein and mRNA increased in oridonin-treated HepG2 cells. Transient transfection of Hsp70. 1-specific shRNA suppressed Hsp70. 1 expression. Compared with control cells,Hsp70.1 shRNA-transfection,oridonin-treatment and combination of both-treatment groups significantly reduced cell viability. Conclusion Hsp70. 1 upregulation contributes to HepG2 cells survival,which suggests that Hsp70. 1 acts as a protective factor in both HepG2 cells and oridonin-treated HepG2 cells.

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

    Directory of Open Access Journals (Sweden)

    Rouhollahi E

    2015-10-01

    Full Text Available Elham Rouhollahi,1 Soheil Zorofchian Moghadamtousi,2 Fatemeh Hajiaghaalipour,3 Maryam Zahedifard,2 Faezeh Tayeby,2 Khalijah Awang,4 Mahmood Ameen Abdulla,3 Zahurin Mohamed1 1Pharmacogenomics Laboratory, Department of Pharmacology, Faculty of Medicine, 2Institute of Biological Sciences, Faculty of Science, 3Department of Biomedical Science, Faculty of Medicine, 4Department of Chemistry, Faculty of Science, University of Malaya, Kuala Lumpur, Malaysia 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

  7. Inducible Hsp70 in the Regulation of Cancer Cell Survival: Analysis of Chaperone Induction, Expression and Activity

    Directory of Open Access Journals (Sweden)

    Elisa Zorzi

    2011-10-01

    Full Text Available Understanding the mechanisms that control stress is central to realize how cells respond to environmental and physiological insults. All the more important is to reveal how tumour cells withstand their harsher growth conditions and cope with drug-induced apoptosis, since resistance to chemotherapy is the foremost complication when curing cancer. Intensive research on tumour biology over the past number of years has provided significant insights into the molecular events that occur during oncogenesis, and resistance to anti-cancer drugs has been shown to often rely on stress response and expression of inducible heat shock proteins (HSPs. However, with respect to the mechanisms guarding cancer cells against proteotoxic stresses and the modulatory effects that allow their survival, much remains to be defined. Heat shock proteins are molecules responsible for folding newly synthesized polypeptides under physiological conditions and misfolded proteins under stress, but their role in maintaining the transformed phenotype often goes beyond their conventional chaperone activity. Expression of inducible HSPs is known to correlate with limited sensitivity to apoptosis induced by diverse cytotoxic agents and dismal prognosis of several tumour types, however whether cancer cells survive because of the constitutive expression of heat shock proteins or the ability to induce them when adapting to the hostile microenvironment remains to be elucidated. Clear is that tumours appear nowadays more “addicted” to heat shock proteins than previously envisaged, and targeting HSPs represents a powerful approach and a future challenge for sensitizing tumours to therapy. This review will focus on the anti-apoptotic role of heat shock 70kDa protein (Hsp70, and how regulatory factors that control inducible Hsp70 synthesis, expression and activity may be relevant for response to stress and survival of cancer cells.

  8. Inducible Hsp70 in the Regulation of Cancer Cell Survival: Analysis of Chaperone Induction, Expression and Activity

    Energy Technology Data Exchange (ETDEWEB)

    Zorzi, Elisa [OncoHematology Clinic of Pediatrics, University-Hospital of Padova, 35100 Padova (Italy); Bonvini, Paolo, E-mail: paolo.bonvini@unipd.it [OncoHematology Clinic of Pediatrics, University-Hospital of Padova, 35100 Padova (Italy); Fondazione Città della Speranza, 36030 Monte di Malo, Vicenza (Italy)

    2011-10-21

    Understanding the mechanisms that control stress is central to realize how cells respond to environmental and physiological insults. All the more important is to reveal how tumour cells withstand their harsher growth conditions and cope with drug-induced apoptosis, since resistance to chemotherapy is the foremost complication when curing cancer. Intensive research on tumour biology over the past number of years has provided significant insights into the molecular events that occur during oncogenesis, and resistance to anti-cancer drugs has been shown to often rely on stress response and expression of inducible heat shock proteins (HSPs). However, with respect to the mechanisms guarding cancer cells against proteotoxic stresses and the modulatory effects that allow their survival, much remains to be defined. Heat shock proteins are molecules responsible for folding newly synthesized polypeptides under physiological conditions and misfolded proteins under stress, but their role in maintaining the transformed phenotype often goes beyond their conventional chaperone activity. Expression of inducible HSPs is known to correlate with limited sensitivity to apoptosis induced by diverse cytotoxic agents and dismal prognosis of several tumour types, however whether cancer cells survive because of the constitutive expression of heat shock proteins or the ability to induce them when adapting to the hostile microenvironment remains to be elucidated. Clear is that tumours appear nowadays more “addicted” to heat shock proteins than previously envisaged, and targeting HSPs represents a powerful approach and a future challenge for sensitizing tumours to therapy. This review will focus on the anti-apoptotic role of heat shock 70kDa protein (Hsp70), and how regulatory factors that control inducible Hsp70 synthesis, expression and activity may be relevant for response to stress and survival of cancer cells.

  9. 旋毛虫HSP70基因的原核表达及其免疫原性%Prokaryotic expression of HSP70 gene of Trichinella spiralis and immunogenicity analysis of the expression product

    Institute of Scientific and Technical Information of China (English)

    徐佳; 禹洋; 唐颖; 俞昭旸; 李晓云; 宋铭忻

    2012-01-01

    利用RT-PCR方法扩增旋毛虫HSP70基因,将其扩增产物克隆到pEASY-Blunt Simple载体中,经限制性酶切后与pET-28a(+)载体连接,进一步转化至感受态细胞Transetta(DE3)中,用IPTG诱导表达重组旋毛虫HSP70。以纯化的重组HSP70制备多克隆抗体,并对HSP70进行免疫印迹分析及免疫组织化学染色。结果表明,重组质粒的鉴定和DNA测序均正确;SDS-PAGE和Western-blot分析结果显示,在70ku处获得一目的条带;免疫组织化学染色结果显示,该HSP70具有很好的免疫原性。证实,成功构建了重组表达质粒pET28a-HSP70,为深入研究旋毛虫HSP70的功能奠定了基础。%HSP70 gene of Trichinella spiralis was amplified by RT-PCR and then cloned into pEASY-Blunt Simple vector.The recombinant was digested and linked with pET-28a(+) vector.The recombinant plasmid was confirmed by PCR and DNA sequencing.The positive plasmid was transformed into competent Escherichia coli Transetta(DE3) and induced by IPTG for expression.The recombinant Trichinella HSP70 was purified to prepare polyclonal antibody.Trichinella HSP70 was analysed by Western-blot and stained by immunohistochemistry.The recombinant protein was confirmed to be about 70 ku in size.The positive signals were localized in the Trichinella by immunohistochemistry.The results showed that the recombinant expression plasmid pET28a-HSP70 was successfully constructed and laid the foundation for further studies on the function of HSP70.

  10. Expression of class Ⅰ MHC molecule, HSP70 and TAP in human hepatocellular carcinoma

    Institute of Scientific and Technical Information of China (English)

    Xiao-Ling Deng; Wei Chen; Mei-Ying Cai; Da-Peng Wei

    2003-01-01

    AIM: To demonstrate whether class Ⅰ MHC molecule,transporter associated with antigen processing (TAP), and heat-shock proteion70 (HSP70) expressed in liver cancer cells before the design and construction of CTL vaccine against hepatocellular carcinoma (HCC).METHODS: We studied 30 HCC specimens by labeled streptavidin biotin (LSAB) method of immunohistochemistry.RESULTS: The results showed that the majority of HCC cells investigated naturally expressed class Ⅰ MHC and TAP,which were different from other tumor cells. Furthermore,we found that HSP70 expressed not only in cellular cytoplasm, but also on the cell surface in HCC.CONCLUSION: Our findings indicate that our understanding about immune escape mechanisms employed by HCC cells may be further improved. It is important to design and construct CTL vaccine against HCC.

  11. Characterisation of a pea hsp70 gene which is both developmentally and stress-regulated.

    Science.gov (United States)

    Dhankher, O P; Drew, J E; Gatehouse, J A

    1997-05-01

    A pea pod cDNA library was screened for sequences specific to lignifying tissue. A cDNA clone (pLP19) encoding the C-terminal region of a hsp70 heat shock protein hybridised only to pod mRNA from pea lines where pod lignification occurred. Expression of pLP19 was induced by heat shock in leaves, stems and roots of pea and chickpea plants. Four different poly(A) addition sites were observed in cDNAs derived from the same gene as pLP19. This gene was fully sequenced; unlike most hsp70 genes, it contains no introns. The 5'-flanking sequence contains heat shock elements and other potential regulatory sequences.

  12. Nuclear HSP90 and HSP70 in COPD patients treated with formoterol or formoterol and corticosteroids

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    Holownia A

    2009-12-01

    Full Text Available Abstract Objective Heat shock proteins assist cellular protein folding and are required for the normal activity of steroid receptors. In this study we assessed nuclear HSP90 and HSP70 proteins and mRNA levels in cells isolated from induced sputum of chronic obstructive pulmonary disease patients treated for 4 weeks with formoterol (F or formoterol+budesonide (F/ICS. Methods Nuclear heat shock protein levels were assessed by Western blot and specific mRNAs were quantified in cell lysates using qRT-PCR. Results Both HSP90 and HSP70 protein levels were higher in the F/ICS-treated patients in comparison with the F-treated group (by 31%, P Conclusions It is possible that increased nuclear heat shock proteins may play a role in the attenuation of the response to glucocorticoids in COPD patients.

  13. 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. PMID:27591626

  14. Overexpression of MuHSP70 gene from Macrotyloma uniflorum confers multiple abiotic stress tolerance in transgenic Arabidopsis thaliana.

    Science.gov (United States)

    Masand, Shikha; Yadav, Sudesh Kumar

    2016-02-01

    A 70-KD heat shock protein (HSP70) is one of the most conserved chaperones. It is involved in de novo protein folding and prevents the aggregation of unfolded proteins under lethal environmental factors. The purpose of this study is to characterise a MuHSP70 from horsegram (Macrotyloma uniflorum) and elucidating its role in stress tolerance of plants. A MuHSP70 was cloned and characterised from a natural drought stress tolerant HPK4 variety of horsegram (M. uniflorum). For functional characterization, MuHSP70 was overexpressed in transgenic Arabidopsis. Overexpression of MuHSP70 was found to provide tolerance to the transgenic Arabidopsis against various stresses such as heat, cold, drought, salinity and oxidative stress. MuHSP70 transgenics were observed to maintain the shoot biomass, root length, relative water content, and chlorophyll content during exposure to multi-stresses relative to non-transgenic control. Transgenic lines have further shown the reduced levels of MDA, H2O2, and proteolytic activity. Together, these findings suggest that overexpression of MuHSP70 plays an important role in improving abiotic stress tolerance and could be a crucial candidate gene for exploration in crop improvement program. PMID:26694324

  15. 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...... 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...

  16. Mutation detection in the human HSP70B′ gene by denaturing high-performance liquid chromatography

    OpenAIRE

    Hecker, Karl H.; Asea, Alexzander; Kobayashi, Kaoru; Green, Stacy; Tang, Dan; Calderwood, Stuart K

    2000-01-01

    Variances, particularly single nucleotide polymorphisms (SNP), in the genomic sequence of individuals are the primary key to understanding gene function as it relates to differences in the susceptibility to disease, environmental influences, and therapy. In this report, the HSP70B′ gene is the target sequence for mutation detection in biopsy samples from human prostate cancer patients undergoing combined hyperthermia and radiation therapy at the Dana-Farber Cancer Institute, using temperature...

  17. 草鱼hsp70和hsp90对温度急性变化的响应%Effect of rapid temperature change on expression of hsp70 and hsp90 in grass carp(Ctenopharyngodon idella)

    Institute of Scientific and Technical Information of China (English)

    周鑫; 董云伟; 王芳; 董双林

    2013-01-01

    Grass carp ( Ctenopharyngodon idella) is one of the most important cultured fishes in China. During transportation in summer,massive mortality often occurs. As molecular chaperones,Hsps assist cells in their recovery from stress and promote cytoprotection. Limited reports described the expression of Hsps in grass carp at high temperatures, and the relationship between oxygen consumption, expression of heat shock protein and mortality needs to be investigated based on the oxygen- and capability- limited thermal tolerance theory (OCLTT) to further elucidate the thermal adaptation of this species to high temperature in view of integrated biology. In the present study,the expressions of hsp70 and hsp90 after rapid temperature increase were studied. Grass carps were acclimated at 20 t and then exposed to designated temperatures(20,22,24, 26,28,30,32,34 ℃) for 3 h heat shock. After 2 h-recovery at 20℃, liver, muscle and gill of tested fish were sampled and hsp70 and hsp90 were determined using semi-quantitive real-time PCR. Expression of hsps increased with temperature increase until 32℃ in muscle and gill, and then decreased at 34 ℃. In liver, expression of hsps kept increasing in all temperature treatments. These results indicated that hsp70 and hsp90 were sensitive to temperature increase in grass carp. Analyzed heat shock proteins data with previous results of oxygen consumption and lethal temperatures of grass carp, the physiological adaptations of grass carp could be explained by oxygen- and capability- limited thermal tolerance theory (OCLTT), which pointed out that the imbalance between oxygen demand and tissue oxygen supply ability was the primary factor which limited organisms' thermal tolerance and the ability of aerobic metabolism greatly limited the survival of organism beyond critical temperatures. With temperature increase, the maximum value of oxygen consumption in grass carp occurred at 28 ℃, and then the oxygen consumption decreased when

  18. Hsp70 confines tumor progression of rat histiocytoma and impedes the cytotoxicity induced by natural killer cells and peritoneal macrophages

    Institute of Scientific and Technical Information of China (English)

    Amere Subbarao Sreedhar

    2010-01-01

    Objective:To study the role of inducible form of heat shock protein 70 (Hsp70) in the host tumor regression of rat tumor model.Methods: We examined the role of Hsp70 in host tumorigenicity andin vitro cellular cytotoxicity using a rat histocytoma. The differential tumor growth and regression kinetics were studied and correlated with the expression of Hsp70, activation of macrophages and natural killer (NK) cells, and circulating or tumor infiltrating immune molecules in the host system.Results: The sub cuteaneous (s.c.) tumor regression was correlated with increased serum cytokines such as IL-12, TNFα,IFNγ and Hsp70. Despite of similar increase of Hsp70 in intraperitoneal (i.p.) tumor implanted animals, animals succumb to tumor growth, further, evidently, no immune molecule activation was observed. The viral promoter driven Hsp70 over expression in these tumor cells restrained solid tumor growth, however, failed to inhibit ascites growth. The NK cells from s.c. immunized animals induces cytotoxicity in the presence of anti-tumor antibody, which necessitated CD40-L expression, conversely, NK cells from i.p. immunized animals failed to induce cytotoxicity. The NK cells from s.c. or i.p. implanted animals with Hsp70 positive tumor cells failed to induce such cytotoxicity. The peritoneal macrophages isolated from s.c. tumor implanted animals when co-cultured with parental BC-8 cells lyses tumor cells, nevertheless entail macrophage specific TNFα expression. On the contrary, Hsp70 expressing BC-8 tumor cells were resistant to peritoneal macrophage induced cytolysis.Conclusions:This study brings out that Hsp70 possibly involved in regulating the host tumor response and cellular cytotoxicity.

  19. HSF1-TPR interaction facilitates export of stress-induced HSP70 mRNA.

    Science.gov (United States)

    Skaggs, Hollie S; Xing, Hongyan; Wilkerson, Donald C; Murphy, Lynea A; Hong, Yiling; Mayhew, Christopher N; Sarge, Kevin D

    2007-11-23

    Stress conditions inhibit mRNA export, but mRNAs encoding heat shock proteins continue to be efficiently exported from the nucleus during stress. How HSP mRNAs bypass this stress-associated export inhibition was not known. Here, we show that HSF1, the transcription factor that binds HSP promoters after stress to induce their transcription, interacts with the nuclear pore-associating TPR protein in a stress-responsive manner. TPR is brought into proximity of the HSP70 promoter after stress and preferentially associates with mRNAs transcribed from this promoter. Disruption of the HSF1-TPR interaction inhibits the export of mRNAs expressed from the HSP70 promoter, both endogenous HSP70 mRNA and a luciferase reporter mRNA. These results suggest that HSP mRNA export escapes stress inhibition via HSF1-mediated recruitment of the nuclear pore-associating protein TPR to HSP genes, thereby functionally connecting the first and last nuclear steps of the gene expression pathway, transcription and mRNA export.

  20. Functional analysis of Drosophila HSP70 promoter with different HSE numbers in human cells.

    Directory of Open Access Journals (Sweden)

    Nadezda Kust

    Full Text Available The activation of genetic constructs including the Drosophila hsp70 promoter with four and eight HSE sequences in the regulatory region has been described in human cells. The promoter was shown to be induced at lower temperatures compared to the human hsp70 promoter. The promoter activity increased after a 60-min heat shock already at 38 °C in human cells. The promoter activation was observed 24 h after heat shock for the constructs with eight HSEs, while those with four HSEs required 48 h. After transplantation of in vitro heat-shocked transfected cells, the promoter activity could be maintained for 3 days with a gradual decline. The promoter activation was confirmed in vivo without preliminary heat shock in mouse ischemic brain foci. Controlled expression of the Gdnf gene under a Drosophila hsp70 promoter was demonstrated. This promoter with four and eight HSE sequences in the regulatory region can be proposed as a regulated promoter in genetic therapeutic systems.

  1. Isoforms of Hsp70-binding human LDL in adult Schistosoma mansoni worms.

    Science.gov (United States)

    Pereira, Adriana S A; Cavalcanti, Marília G S; Zingali, Russolina B; Lima-Filho, José L; Chaves, Maria E C

    2015-03-01

    Schistosoma mansoni is one of the most common parasites infecting humans. They are well adapted to the host, and this parasite's longevity is a consequence of effective escape from the host immune system. In the blood circulation, lipoproteins not only help to conceal the worm from attack by host antibodies but also act as a source of lipids for S. mansoni. Previous SEM studies showed that the low-density lipoprotein (LDL) particles present on the surface of adult S. mansoni worms decreased in size when the incubation time increased. In this study, immunocytochemical and proteomic analyses were used to locate and identify S. mansoni binding proteins to human plasma LDL. Ultrathin sections of adult worms were cut transversely from the anterior, medial and posterior regions of the parasite. Immunocytochemical experiments revealed particles of gold in the tegument, muscle region and spine in male worms and around vitelline cells in females. Immunoblotting and 2D-electrophoresis using incubations with human serum, anti-LDL antibodies and anti-chicken IgG peroxidase conjugate were performed to identify LDL-binding proteins in S. mansoni. Analysis of the binding proteins using LC-MS identified two isoforms of the Hsp70 chaperone in S. mansoni. Hsp70 is involved in the interaction with apoB in the cytoplasm and its transport to the endoplasmic reticulum. However, further studies are needed to clarify the functional role of Hsp70 in S. mansoni, mainly related to the interaction with human LDL.

  2. Polyglutamine expansion in Drosophila: thermal stress and Hsp70 as selective agents

    Indian Academy of Sciences (India)

    Brian R Bettencourt; Catherine C Hogan; Mario Nimali

    2007-04-01

    Repetitive DNA sequences that encode polyglutamine tracts are prone to expansion and cause highly deleterious phenotypes of neurodegeneration. Despite this tendency, polyglutamine tracts (``polyQs”) are conserved features of eukaryotic genomes. PolyQs are the most frequent protein-coding homotypic repeat in insect genomes, and are found predominantly in genes encoding transcription factors conserved from Drosophila through human. Although highly conserved across species, polyQ lengths vary widely within species. In D. melanogaster, polyQs in 25 genes have more alleles and higher heterozygosity than all other poly-amino acid tracts. The heat shock protein Hsp70 is a principal suppressor of polyQ expansions and may play a key role in modulating the phenotypes of the alleles that encode them. Hsp70 also promotes tolerance of natural thermal stress in Drosophila and diverse organisms, a role which may deplete the chaperone from buffering against polyQ toxicity. Thus in stressful environments, natural selection against long polyQ alleles more prone to expansion and deleterious phenotypes may be more effective. This hypothesis can be tested by measuring the phenotypic interactions between Hsp70 and polyQ transgenes in D. melanogaster undergoing natural thermal stress, an approach which integrates comparative genomics with experimental and ecological genetics.

  3. Identification of an Allosteric Pocket on Human Hsp70 Reveals a Mode of Inhibition of This Therapeutically Important Protein

    OpenAIRE

    Rodina, Anna; Patel, Pallav D.; Kang, Yanlong; Patel, Yogita; Baaklini, Imad; Wong, Michael J. H.; Taldone, Tony; Yan, Pengrong; Yang, Chenghua; Maharaj, Ronnie; Gozman, Alexander; Patel, Maulik R.; Patel, Hardik J.; Chirico, William; Erdjument-Bromage, Hediye

    2013-01-01

    Hsp70s are important cancer chaperones that act upstream of Hsp90 and exhibit independent anti-apoptotic activities. To develop chemical tools for the study of human Hsp70, we developed a homology model that unveils a previously unknown allosteric site located in the nucleotide binding domain of Hsp70. Combining structure-based design and phenotypic testing, we discovered a previously unknown inhibitor of this site, YK5. In cancer cells, this compound is a potent and selective binder of the c...

  4. Histological, ultrastructural and heat shock protein 70 (HSP70) responses to heat stress in the sea cucumber Apostichopus japonicus.

    Science.gov (United States)

    Xu, Dongxue; Sun, Lina; Liu, Shilin; Zhang, Libin; Yang, Hongsheng

    2015-08-01

    The aquaculture industry for Apostichopus japonicus has suffered severe economic and resource losses due to high temperature in recent summers. There is increasing concern about the effect of high temperature on this species. Histological, ultrastructural and HSP70 responses to heat stress were investigated in the intestine of A. japonicus. Tissue degradation was observed in muscular, submucosal and mucosal layers, with significant decrease in plicae circulares of the mucosal layer. Ultrastructural damage intensified with increasing stress time, and indicators of cell apoptosis were evident after 192 h heat stress. Immunostaining showed HSP70 mainly in mucosa and serosa, with faint staining in non-stressed individuals (the control group) and denser staining under stress (the 6, 48 and 192 h groups). Western blot detection confirmed ocurrence of HSP70 in all groups and significant up-regulation under stress. The rapid and persistent response of HSP70 implies its critical role in the heat shock response of A. japonicus. PMID:25917397

  5. Coagulansin-A has beneficial effects on the development of bovine embryos in vitro via HSP70 induction

    OpenAIRE

    Khan, Imran; Lee, Kyeong-Lim; Fakruzzaman, Md.; Song, Seok-Hwan; Ihsan-ul-Haq,; Mirza, Bushra; Yan, Chang Guo; Kong, Il-Keun

    2016-01-01

    Treatment with the steroidal lactone, coagulansin-A, improves bovine oocyte maturation and embryo development in vitro by inducing heat shock protein 70 (HSP70), which reduces the levels of reactive oxygen species (ROS), DNA damage and inflammation.

  6. An inducible HSP70 gene from the midge Chironomus dilutus: Characterization and transcription profile under environmental stress

    Science.gov (United States)

    Karouna-Renier, N. K.; Rao, K.R.

    2009-01-01

    In the present study, we identified and characterized an inducible heat shock protein 70 (HSP70) from the midge Chironomus dilutus and investigated the transcriptional profile of the gene under baseline and environmentally stressful conditions. Using real-time quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR), we observed increased expression of CD-HSP70-1 in response to both heat shock and copper stress. We also investigated the expression of this gene during midge development. All C. dilutus developmental stages expressed CD-HSP70-1 under normal conditions, although at extremely low levels. Phylogenetic analysis of the amino acid sequence demonstrated distinct clustering of this gene with inducible HSP70s from other insect species. ?? 2008 The Authors.

  7. Inhibition of cellular proliferation by the Wilms tumor suppressor WT1 requires association with the inducible chaperone Hsp70

    OpenAIRE

    Maheswaran, Shyamala; Englert, Christoph; Zheng, Gang; Lee, Sean Bong; Wong, Jenise; Harkin, D Paul; Bean, James; Ezzell, Robert; Garvin, A. Julian; McCluskey, Robert T.; DeCaprio, James A.; Haber, Daniel A.

    1998-01-01

    The Wilms tumor suppressor WT1 encodes a zinc finger transcription factor that is expressed in glomerular podocytes during a narrow window in kidney development. By immunoprecipitation and protein microsequencing analysis, we have identified a major cellular protein associated with endogenous WT1 to be the inducible chaperone Hsp70. WT1 and Hsp70 are physically associated in embryonic rat kidney cells, in primary Wilms tumor specimens and in cultured cells with inducible expression of WT1. Co...

  8. Elimination of Enhanced Thermal Resistance of Spheroid Culture Model of Prostate Carcinoma Cell Line by Inhibitors of Hsp70 Induction

    Directory of Open Access Journals (Sweden)

    Samideh Khoei

    2010-01-01

    Full Text Available AbstractObjective: The purpose of this study was to investigate the enhanced thermal resistancemechanism of the DU145 tumor spheroid cultures as compared to the prostate carcinomacell line's monolayer cultures.Materials and Methods: DU145 cells were cultured either as spheroids or monolayers.Cultures were treated with hyperthermia in a precision water bath (at 43°C for 60 minutesand/or quercetin (50 and 500 μM for monolayer and spheroid cultures respectively. Afterhyperthermic treatment, the cell viability colony forming ability, and the expression of heatshock protein 70 (Hsp70 were examined in both culture systems. Hsp70 expression wasstudied using the western blot method.Results: Our results showed that the DU145 monolayer and spheroid cell culture treatmentwith hyperthermia alone resulted in a marked survival inhibition. Furthermore, thespheroids showed a more significant resistance to hyperthermia compared to the monolayercultures (p = 0.01. They also produced more Hsp70 than the monolayer cultures.Treatment of cells with quercetin reduced the Hsp70 level in both culture systems. However,with the reduced Hsp70 levels, thermal resistance of the spheroids showed a greaterdecrease in relation to that of the monolayers.Conclusion: The results suggest that the enhanced hyperthermia resistance mechanismof the spheroid cultures compared to that of the monolayer cultures can be attributed tospheroids' Hsp70 production.

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

    Science.gov (United States)

    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. PMID:27334602

  10. Nuclear HSP90 and HSP70 in COPD patients treated with formoterol or formoterol and corticosteroids

    OpenAIRE

    Holownia A; Mroz RM; Kielek A; Chyczewska E; Braszko JJ

    2009-01-01

    Abstract Objective Heat shock proteins assist cellular protein folding and are required for the normal activity of steroid receptors. In this study we assessed nuclear HSP90 and HSP70 proteins and mRNA levels in cells isolated from induced sputum of chronic obstructive pulmonary disease patients treated for 4 weeks with formoterol (F) or formoterol+budesonide (F/ICS). Methods Nuclear heat shock protein levels were assessed by Western blot and specific mRNAs were quantified in cell lysates usi...

  11. Characterizing HSF1 Binding and Post-Translational Modifications of hsp70 Promoter in Cultured Cortical Neurons: Implications in the Heat-Shock Response.

    Directory of Open Access Journals (Sweden)

    Andrea V Gómez

    Full Text Available Causes of lower induction of Hsp70 in neurons during heat shock are still a matter of debate. To further inquire into the mechanisms regulating Hsp70 expression in neurons, we studied the activity of Heat Shock Factor 1 (HSF1 and histone posttranslational modifications (PTMs at the hsp70 promoter in rat cortical neurons. Heat shock induced a transient and efficient translocation of HSF1 to neuronal nuclei. However, no binding of HSF1 at the hsp70 promoter was detected while it bound to the hsp25 promoter in cortical neurons during heat shock. Histone PTMs analysis showed that the hsp70 promoter harbors lower levels of histone H3 and H4 acetylation in cortical neurons compared to PC12 cells under basal conditions. Transcriptomic profiling data analysis showed a predominant usage of cryptic transcriptional start sites at hsp70 gene in the rat cerebral cortex, compared with the whole brain. These data support a weaker activation of hsp70 canonical promoter. Heat shock increased H3Ac at the hsp70 promoter in PC12 cells, which correlated with increased Hsp70 expression while no modifications occurred at the hsp70 promoter in cortical neurons. Increased histone H3 acetylation by Trichostatin A led to hsp70 mRNA and protein induction in cortical neurons. In conclusion, we found that two independent mechanisms maintain a lower induction of Hsp70 in cortical neurons. First, HSF1 fails to bind specifically to the hsp70 promoter in cortical neurons during heat shock and, second, the hsp70 promoter is less accessible in neurons compared to non-neuronal cells due to histone deacetylases repression.

  12. hsp70 genes in the human genome: Conservation and differentiation patterns predict a wide array of overlapping and specialized functions

    Directory of Open Access Journals (Sweden)

    Macario Alberto JL

    2008-01-01

    Full Text Available Abstract Background Hsp70 chaperones are required for key cellular processes and response to environmental changes and survival but they have not been fully characterized yet. The human hsp70-gene family has an unknown number of members (eleven counted over ten years ago; some have been described but the information is incomplete and inconsistent. A coherent body of knowledge encompassing all family components that would facilitate their study individually and as a group is lacking. Nowadays, the study of chaperone genes benefits from the availability of genome sequences and a new protocol, chaperonomics, which we applied to elucidate the human hsp70 family. Results We identified 47 hsp70 sequences, 17 genes and 30 pseudogenes. The genes distributed into seven evolutionarily distinct groups with distinguishable subgroups according to phylogenetic and other data, such as exon-intron and protein features. The N-terminal ATP-binding domain (ABD was conserved at least partially in the majority of the proteins but the C-terminal substrate-binding domain (SBD was not. Nine proteins were typical Hsp70s (65–80 kDa with ABD and SBD, two were lighter lacking partly or totally the SBD, and six were heavier (>80 kDa with divergent C-terminal domains. We also analyzed exon-intron features, transcriptional variants and protein structure and isoforms, and modality and patterns of expression in various tissues and developmental stages. Evolutionary analyses, including human hsp70 genes and pseudogenes, and other eukaryotic hsp70 genes, showed that six human genes encoding cytosolic Hsp70s and 27 pseudogenes originated from retro-transposition of HSPA8, a gene highly expressed in most tissues and developmental stages. Conclusion The human hsp70-gene family is characterized by a remarkable evolutionary diversity that mainly resulted from multiple duplications and retrotranspositions of a highly expressed gene, HSPA8. Human Hsp70 proteins are clustered into

  13. Isolation and Identification of Mycoplasma ovipneumoniae and Genetic Diversity of HSP70 Gene%绵羊肺炎支原体的分离鉴定及其 HSP70基因遗传多样性

    Institute of Scientific and Technical Information of China (English)

    胡政香; 乔军; 孟庆玲; 张金生; 陈创夫

    2015-01-01

    The colonial morphology,biochemical indexes and molecular biology of the pathogens isolated from lung samples of 7 suspected M.ovipneumoniae infection sheep farms were analyzed to discuss genetic diversity of M.ovipneumoniaeep epidemic strain in Xinjiang.Results:5 isolated strains all are M. ovipneumoniae.The colony is papillary and thallus is polymorphism.15 isolated strains can hydrolyze glucose,but can not hydrolyze arginine and urea.15 isolated strains can fade and adsorb methylene blue milk,which results in erythrocytes hemolysis.The size of 16Sr RNA sequence is 361 bp.The nucleotide homology and amino acid sequence homology of HSP70 gene among 5 isolated strains are 94.1% and 93.2%respectively.No.593 locus of the isolated strains is inserted with Gln compared with the standard strain Y98.There is genetic diversity in the isolated strains in Xinjiang.%为探明新疆地区绵羊肺炎支原体(Mycoplasma ovipneumoniae ,MO)流行株是否存在遗传多样性,用 KM2培养基对塔城地区7个羊场疑似 MO 感染的肺脏病料进行病原分离,并进行菌落形态、生化试验和分子生物学鉴定。结果表明:15株分离均为 MO,菌落呈乳头状,菌体呈多形性;能水解葡萄糖但不能水解精氨酸、尿素,可使美兰牛乳褪色、吸附并引起红细胞溶血;16Sr RNA 序列大小为361 bp。其中,5株 MO HSP 70基因的核苷酸同源性为94.1%,氨基酸同源性为93.2%,与 MO 标准毒株 Y98相比,分离株在593位点均插入 Gln(谷氨酰胺)。MO 新疆分离株已发生遗传变异,存在遗传多样性。

  14. Co-expression of heat shock protein (HSP) 40 and HSP70 in Pinctada martensii response to thermal, low salinity and bacterial challenges.

    Science.gov (United States)

    Li, Jun; Zhang, Yuehuan; Liu, Ying; Zhang, Yang; Xiao, Shu; Yu, Ziniu

    2016-01-01

    Heat shock protein (HSP) 40 proteins are a family of molecular chaperones that bind to HSP70 through their J-domain and regulate the function of HSP70 by stimulating its adenosine triphosphatase activity. In the present study, a HSP40 homolog named PmHSP40 was cloned from the hemocytes of pearl oyster Pinctada martensii using EST and rapid amplification of cDNA ends (RACE) techniques. The full-length cDNA of PmHSP40 was 1251 bp in length, which included a 5' untranslated region (UTR) of 75 bp, an open reading frame (ORF) of a 663 bp, and a 3' UTR of 513 bp. The deduced amino acid sequence of PmHSP40 contains a J domain in the N-terminus. In response to thermal and low salinity stress challenges, the expression of PmHSP40 in hemocytes and the gill were inducible in a time-dependent manner. After bacterial challenge, PmHSP40 transcripts in hemocytes increased and peaked at 6 h post injection. In the gill, PmHSP40 expression increased, similar to expression in hemocytes; however, transcript expression of PmHSP40 was significantly up-regulated at 12 h post injection. Furthermore, the transcripts of PmHSP70 showed similar kinetics as that of PmHSP40, with highest induction during thermal, low salinity stress and bacterial challenges. Altogether these results demonstrate that PmHSP40 is an inducible protein under thermal, low salinity and bacterial challenges, suggesting its involvement in both environmental and biological stresses, and in the innate immunity of the pearl oyster. PMID:26679110

  15. Low sequence identity but high structural and functional conservation: The case of Hsp70/Hsp90 organizing protein (Hop/Sti1) of Leishmania braziliensis.

    Science.gov (United States)

    Batista, Fernanda A H; Seraphim, Thiago V; Santos, Clelton A; Gonzaga, Marisvanda R; Barbosa, Leandro R S; Ramos, Carlos H I; Borges, Júlio C

    2016-06-15

    Parasites belonging to the genus Leishmania are subjected to extensive environmental changes during their life cycle; molecular chaperones/co-chaperones act as protagonists in this scenario to maintain cellular homeostasis. Hop/Sti1 is a co-chaperone that connects the Hsp90 and Hsp70 systems, modulating their ATPase activities and affecting the fate of client proteins because it facilitates their transfer from the Hsp70 to the Hsp90 chaperone. Hop/Sti1 is one of the most prevalent co-chaperones, highlighting its importance despite the relatively low sequence identity among orthologue proteins. This multi-domain protein comprises three tetratricopeptides domains (TPR1, TPR2A and TPR2B) and two Asp/Pro-rich domains. Given the importance of Hop/Sti1 for the chaperone system and for Leishmania protozoa viability, the Leishmania braziliensis Hop (LbHop) and a truncated mutant (LbHop(TPR2AB)) were characterized. Structurally, both proteins are α-helix-rich and highly elongated monomeric proteins. Functionally, they inhibited the ATPase activity of Leishmania braziliensis Hsp90 (LbHsp90) to a similar extent, and the thermodynamic parameters of their interactions with LbHsp90 were similar, indicating that TPR2A-TPR2B forms the functional center for the LbHop interaction with LbHsp90. These results highlight the structural and functional similarity of Hop/Sti1 proteins, despite their low sequence conservation compared to the Hsp70 and Hsp90 systems, which are phylogenetic highly conserved.

  16. Low sequence identity but high structural and functional conservation: The case of Hsp70/Hsp90 organizing protein (Hop/Sti1) of Leishmania braziliensis.

    Science.gov (United States)

    Batista, Fernanda A H; Seraphim, Thiago V; Santos, Clelton A; Gonzaga, Marisvanda R; Barbosa, Leandro R S; Ramos, Carlos H I; Borges, Júlio C

    2016-06-15

    Parasites belonging to the genus Leishmania are subjected to extensive environmental changes during their life cycle; molecular chaperones/co-chaperones act as protagonists in this scenario to maintain cellular homeostasis. Hop/Sti1 is a co-chaperone that connects the Hsp90 and Hsp70 systems, modulating their ATPase activities and affecting the fate of client proteins because it facilitates their transfer from the Hsp70 to the Hsp90 chaperone. Hop/Sti1 is one of the most prevalent co-chaperones, highlighting its importance despite the relatively low sequence identity among orthologue proteins. This multi-domain protein comprises three tetratricopeptides domains (TPR1, TPR2A and TPR2B) and two Asp/Pro-rich domains. Given the importance of Hop/Sti1 for the chaperone system and for Leishmania protozoa viability, the Leishmania braziliensis Hop (LbHop) and a truncated mutant (LbHop(TPR2AB)) were characterized. Structurally, both proteins are α-helix-rich and highly elongated monomeric proteins. Functionally, they inhibited the ATPase activity of Leishmania braziliensis Hsp90 (LbHsp90) to a similar extent, and the thermodynamic parameters of their interactions with LbHsp90 were similar, indicating that TPR2A-TPR2B forms the functional center for the LbHop interaction with LbHsp90. These results highlight the structural and functional similarity of Hop/Sti1 proteins, despite their low sequence conservation compared to the Hsp70 and Hsp90 systems, which are phylogenetic highly conserved. PMID:27103305

  17. 赤拟谷盗HSP70基因克隆和竞争定量PCR检测%Cloning and quantitative competitive PCR assay of HSP70 gene in Tribolium castaneum ( Coleoptera: Tenebrionidae)

    Institute of Scientific and Technical Information of China (English)

    苏丽娟; 董晓慧; 尹新明

    2011-01-01

    To study the expression changes of heat shock protein 70 (a highly conserved protein) in Tribolium castaneum after exposure to heat stress, a fragment of 681 bp encoding for HSP70 was amplified and sequenced from T.castaneum.The fragment encoded 227 amino acid residues with the GenBank accession no.HM345948.The result of homology analysis showed that this fragment shared 97% identity with hsp70 from L.decemlineata (AF322911.1 ).Comparison of the deduced amino acid sequences of HSP70 in T.castaneum with those in Leptinotarsa decemlineata, Mamestra brassicae, Drosophila melanogaster and Liriomyza sativae indicated that they shared more than 94% identity.The internal competitor used in quantitative competitive PCR was obtained by RT-PCR.The detection system of hsp70 was constructed by PCR amplification using the same quantity of target cDNA and a series of diluted internal competitor as template.The linear equation of standard curve was Y= 1.032X- 1.618 (r2 =0.975).This study provides a very convenient method for the quantitation of the hsp70 expression changes in T.castaneum and offers the basic data for the prevention and control of pests using thermal control technology.%为研究赤拟谷盗Tribolium castaneum受到热胁迫后高度保守的热激蛋白70(heat shock protein 70,HSP70)基因的表达变化,本研究扩增了681 bp的赤拟谷盗hsp70片段,编码227个氨基酸残基,GenBank登录号为HM345948.同源性分析表明:赤拟谷盗hsp70核苷酸序列与马铃薯甲虫Leptinotarsa decemlineata的hsp70(GenBank登录号:AF322911.1)同源性最高,为97%;其推测的蛋白序列与马铃薯甲虫、甘蓝夜蛾Mamestra brassicae、黑腹果蝇Drosophila melanogaster和美洲斑潜蝇Liriomyza,sativae的HSP70蛋白均有94%以上的同源性.利用RT-PCR技术得到与赤拟谷盗hsp70进行竞争定量的内部竞争物,以等量的目标cDNA和一系列稀释的竞争模板进行竞争PCR扩增,构建了hsp70的竞争定量PCR检测体系,该

  18. HPV16E7-HSP70 Hybrid DNA Vaccine Induces E7-Specific Cytotoxic T Cells and Antitumor Immunity

    Institute of Scientific and Technical Information of China (English)

    ZHU Liqin; LI Hui; XIONG Jinhu; WANG Tongxiang; OU Xuan; WEI Yun; WU Xinxing

    2006-01-01

    Using human papillomavirus type 16 (HPV16) E7 as an antigen and Heat Shock Protein 70 as adjuvant, we constructed a DNA vaccine by linking HSP70 gene to E7C91G gene. Mice, after being immunized with E7C91G-HSP70, E7C91G/HSP70, E7C91G, and wild E7 DNA vaccines respectively, produced E7 specific CD8+ T-cell precursor frequencies oF280. 33±2.52, 144.34±4. 04, 164.34±5.13 and 82.33± 3.51 respectively within every 1 × 105 mouse splenocytes. This proves that E7C91G-HSP70 fusion vaccine can significantly enhance the E7 specific cellular immunity within the mice body(p<0.01). After being immunized with E7C91G-HSP70 fusion vaccine, tumor-bearing mice of the group being treated have significantly longer latency and survival periods, comparing with other three categories of E7 vaccines. Experiment shows that this vaccine has a significant effect on enhancing E7 positive tumor-treatment within mice body. After being immunized with E7C91G-HSP70 vaccine, there were no pathological changes found in livers, kidneys and spleens of the mice, which proves that the vaccine is quite safe. After all,E7C91G-HSP70 fusion vaccine has a much stronger tumor- treatment effect than that of wild type E7 DNA vaccine.

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

    Directory of Open Access Journals (Sweden)

    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.

  20. Developmental expression of Hsp90, Hsp70 and HSF during morphogenesis in the vetigastropod Haliotis asinina.

    Science.gov (United States)

    Gunter, Helen M; Degnan, Bernard M

    2007-08-01

    Heat shock proteins (Hsps) have dual functions, participating in both the stress response and a broad range of developmental processes. At physiological temperatures, it has been demonstrated in deuterostomes (vertebrates) and ecdysozoans (insects) that Hsps are expressed in tissues that are undergoing differentiation and morphogenesis. Here we investigate the developmental expression of Hsp70, Hsp90 and their regulatory transcription factor heat shock transcription factor (HSF) in the marine gastropod Haliotis asinina, a representative of the 3rd major lineage of bilaterian animals, the Lophotrochozoa. HasHsp70, HasHsp90 and HasHSF are maternally expressed in H. asinina and are progressively restricted to the micromere lineage during cleavage. During larval morphogenesis, they are expressed in unique and overlapping patterns in the prototroch, foot, and mantle. Hsp expression peaked in these tissues during periods of cell differentiation and morphogenesis, returning to lower levels after morphogenesis was complete. These patterns of Hsp and HSF expression in H. asinina are akin to those observed in ecdysozoans and deuterostomes, with Hsps being activated in cells and tissues undergoing morphogenesis.

  1. Priming the immune system of Penaeid shrimp by bacterial HSP70 (DnaK).

    Science.gov (United States)

    Phuoc, L H; Hu, B; Wille, M; Hien, N T; Phuong, V H; Tinh, N T N; Loc, N H; Sorgeloos, P; Bossier, P

    2016-05-01

    This study was conducted to test the effect of DnaK on priming immune responses in Penaeid shrimp. Juvenile-specific pathogen-free (SPF) P. vannamei shrimp were injected with 0.05 μg recombinant DnaK. One hour post-DnaK priming, a non-lethal dose of Vibrio campbellii (10(5) CFU shrimp(-1)) was injected. Other treatments include only DnaK or V. campbellii injection or control with blank inocula. The haemolymph of three shrimp from each treatment was collected at 1.5, 6, 9 and 12 h post-DnaK priming (hpp). It was verified that injection with DnaK and V. campbellii challenge affected the transcription of 3 immune genes, transglutaminase-1 (TGase-1), prophenoloxidase-2 (proPO-2) and endogenous HSP70 (lvHSP70). In P. monodon, shrimp were first injected with DnaK at a dose of 10 μg shrimp(-1) and one hour later with 10(6) CFU of V. harveyi (BB120) shrimp(-1). Shrimp injected with DnaK showed a significant increase in proPO expression compared to the control (P < 0.05). Yet a double injection (DnaK and Vibrio) seemed to cause an antagonistic response at the level of expression, which was not equalled at the level of PO activity. Those results suggest that DnaK is able to modulate immune responses in P. vannamei and P. monodon.

  2. 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)

  3. Expression, Purification and Immunologic Identification of the Recombinant HSP70 Gene From Echinococcus Granulosus%细粒棘球蚴重组HSP70基因的表达、纯化及免疫学鉴定

    Institute of Scientific and Technical Information of China (English)

    于晶晶; 于辛酉; 王娅娜; 师志云; 马锐; 卜阳; 罗永云; 赵巍

    2008-01-01

    目的 构建细粒棘球蚴(Echinococcus granulosus,Eg)热休克蛋白70(Heat shock protein,HSP70)基因的重组质粒并原核表达、纯化及对其免疫特性进行鉴定.方法 从重组质粒HSP70/pGEM-T中酶切HSP70目的片段,亚克隆入表达载体pET-28a,转化入大肠杆菌E.coli BL21(DE3)plysS进行融合表达,经His-band树脂纯化试剂盒小量纯化,SDS-PAGE 和Western blot方法鉴定表达产物.结果 成功构建Eg.HSP70/ pET-28a/ E.coli BL21基因工程菌株,诱导表达重组蛋白和纯化分离得到14kDa Eg.HSP70均能被细粒棘球蚴天然抗原免疫的兔多克隆抗血清识别.结论 初步证实该重组蛋白具有较好的抗原性.

  4. Hsp70 - a biomarker for tumor detection and monitoring of outcome of radiation therapy in patients with squamous cell carcinoma of the head and neck

    International Nuclear Information System (INIS)

    Tumor but not normal cells frequently overexpress heat shock protein 70 (Hsp70) and present it on their cell surface (mHsp70) from where it can be actively released. Therefore, membrane (mHsp70) and soluble Hsp70 (sHsp70) were investigated as potential tumor biomarkers and for monitoring the outcome of radiation therapy. Biopsies and blood were collected from patients with squamous cell carcinoma of the head and neck (SCCHN) at different time points (before, during therapy and in the follow-up period). Hsp70 membrane expression was determined on single cell suspensions of tumor biopsies and reference tissues by flow cytometry, sHsp70 protein and antibody levels were determined in the serum of patients and healthy donors by ELISA and NK cell markers that are related to the presence of sHsp70 were analyzed in the patient’s peripheral blood lymphocytes (PBL). Tumor biopsies exhibited significantly increased mHsp70 expression levels compared to the reference tissue. Soluble Hsp70 levels were significantly higher in SCCHN patients compared to healthy human volunteers and high mHsp70 expression levels on tumor cells were associated with high sHsp70 levels in the serum of patients. Following surgery and radiotherapy sHsp70 levels in patients dropped in patients without tumor relapse in the follow-up period. In contrast to sHsp70 protein, anti-Hsp70 antibody levels remained nearly unaltered in the serum of SCCHN patients before and after therapy. Furthermore, sHsp70 protein but not anti-Hsp70 antibody levels were found to be associated with the tumor volume in SCCHN patients before start of therapy. The expression densities of the activatory NK cell markers CD56, CD94, NKG2D, NKp30, Nkp44, and NKp46 differed in patients following therapeutic intervention. A significant increase in the density of NKG2D was observed in SCCHN patients in the follow-up period after surgery and radiotherapy. We suggest sHsp70 as a potential biomarker for detecting tumors and for monitoring

  5. 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

  6. Cytological and molecular characterization of three gametoclones of Citrus clementina

    OpenAIRE

    GERMANA, MARIA ANTONIETTA; Aleza, Pablo; Carrera, Esther; Chen, Chunxian; Chiancone, Benedetta; Costantino, Gilles; Dambier, Dominique; Deng, Xiuxin; Federici, Claire T.; Froelicher, Yann; Guo, Wenwu; Ibanez, Victoria; Juarez, Jose; Kwok, Kevin; Luro, François

    2013-01-01

    Abstract Background Three gametoclonal plants of Citrus clementina Hort. ex Tan., cv. Nules, designated ESP, FRA, and ITA (derived from three labs in Spain, France, and Italy, respectively), were selected for cytological and molecular characterization in order to elucidate genomic rearrangements provoked by haploidization. The study included comparisons of their ploidy, homozygosity, genome integrity, and gene dosage, using chromosome counting, flow c...

  7. Role of HSP70 in motoneuron survival after excitotoxic stress in a rat spinal cord injury model in vitro.

    Science.gov (United States)

    Shabbir, Ayisha; Bianchetti, Elena; Cargonja, Renato; Petrovic, Antonela; Mladinic, Miranda; Pilipović, Kristina; Nistri, Andrea

    2015-12-01

    The outcome for gait recovery from paralysis due to spinal lesion remains uncertain even when damage is limited. One critical factor is the survival of motoneurons, which are very vulnerable cells. To clarify the early pathophysiological mechanisms of spinal damage, an in vitro injury model of the rat spinal cord caused by moderate excitotoxicity was used. With this preparation we investigated whether motoneuron survival was dependent on the expression of the neuroprotective protein HSP70. In the present study excitotoxicity evoked by kainate induced delayed (24 h) loss (35%) of motoneurons, which became pyknotic with translocation of the cell death biomarker apoptosis-inducing factor (AIF) to the nucleus. This process was concomitant with suppression of locomotor network electrical activity. Surviving cells showed strong expression of HSP70 without nuclear AIF. The HSP70 inhibitor VER155008 per se induced neurotoxicity similar to that of kainate, while the HSP90 inhibitor geldanamycin did not damage spinal tissue. Electrophysiological recording following kainate or VER155008 indicated depression of motoneuron field potentials, with decreased excitability and impaired synaptic transmission. When these two drugs were applied together, more intense neurotoxicity emerged. Our data indicate that HSP70 was one important contributor to motoneuron survival and suggest that enhancing HSP70 activity is a potential future strategy for neuroprotecting these cells. PMID:26490753

  8. Genome-wide identification of Hsp70 genes in channel catfish and their regulated expression after bacterial infection.

    Science.gov (United States)

    Song, Lin; Li, Chao; Xie, Yangjie; Liu, Shikai; Zhang, Jiaren; Yao, Jun; Jiang, Chen; Li, Yun; Liu, Zhanjiang

    2016-02-01

    Heat shock proteins 70/110 (Hsp70/110) are a family of conserved ubiquitously expressed heat shock proteins which are produced by cells in response to exposure to stressful conditions. Besides the chaperone and housekeeping functions, they are also known to be involved in immune response during infection. In this study, we identified 16 Hsp70/110 geness in channel catfish (Ictalurus punctatus) through in silico analysis using RNA-Seq and genome databases. Among them 12 members of Hsp70 (Hspa) family and 4 members of Hsp110 (Hsph) family were identified. Phylogenetic and syntenic analyses provided strong evidence in supporting the orthologies of these HSPs. In addition, we also determined the expression patterns of Hsp70/110 genes after Flavobacterium columnare and Edwardsiella ictaluri infections by meta-analyses, for the first time in channel catfish. Ten out of sixteen genes were significantly up/down-regulated after bacterial challenges. Specifically, nine genes were found significantly expressed in gill after F. columnare infection. Two genes were found significantly expressed in intestine after E. ictaluri infection. Pathogen-specific pattern and tissue-specific pattern were found in the two infections. The significantly regulated expressions of catfish Hsp70 genes after bacterial infections suggested their involvement in immune response in catfish. PMID:26693666

  9. Erianthus arundinaceus HSP70 (EaHSP70) Acts as a Key Regulator in the Formation of Anisotropic Interdigitation in Sugarcane (Saccharum spp. hybrid) in Response to Drought Stress.

    Science.gov (United States)

    Augustine, Sruthy Maria; Cherian, Anoop V; Syamaladevi, Divya P; Subramonian, N

    2015-12-01

    Plant growth during abiotic stress is a long sought-after trait especially in crop plants in the context of global warming and climate change. Previous studies on leaf epidermal cells have revealed that during normal growth and development, adjacent cells interdigitate anisotropically to form cell morphological patterns known as interlocking marginal lobes (IMLs), involving the cell wall-cell membrane-cortical actin continuum. IMLs are growth-associated cell morphological changes in which auxin-binding protein (ABP), Rho GTPases and actin are known to play important roles. In the present study, we investigated the formation of IMLs under drought stress and found that Erianthus arundinaceus, a drought-tolerant wild relative of sugarcane, develops such growth-related cell morphological patterns under drought stress. Using confocal microscopy, we showed an increasing trend in cortical F-actin intensity in drought-tolerant plants with increasing soil moisture stress. In order to check the role of drought tolerance-related genes in IML formation under soil moisture stress, we adopted a structural data mining strategy and identified indirect connections between the ABPs and heat shock proteins (HSPs). Initial experimental evidence for this connection comes from the high transcript levels of HSP70 observed in drought-stressed Erianthus, which developed anisotropic interdigitation, i.e. IMLs. Subsequently, by overexpressing the E. arundinaceus HSP70 gene (EaHSP70) in sugarcane (Saccharum spp. hybrid), we confirm the role of HSP70 in the formation of anisotropic interdigitation under drought stress. Taken together, our results suggest that EaHSP70 acts as a key regulator in the formation of anisotropic interdigitation in drought-tolerant plants (Erianthus and HSP70 transgenic sugarcane) under moisture stress in an actin-mediated pathway. The possible biological significance of the formation of drought-associated interlocking marginal lobes (DaIMLs) in sugarcane plants upon

  10. 猪带绦虫HSP70-4的真核表达及其抗血清的制备%Eukaryotic expression and anti-serum preparation of HSP70-4 from Taenia solium

    Institute of Scientific and Technical Information of China (English)

    殷静; 王帅; 刘光学; 何伟; 才学鹏

    2016-01-01

    为了研究猪带绦虫(Taenia solium)热休克蛋白70-4 (TsHSP70-4)基因序列的特征及其真核表达蛋白的抗原性,参考GeneDB中猪带绦虫基因组注释信息,设计特异性引物,RT-PCR扩增TsHSP70-4ORF序列.根据毕赤酵母密码子偏好性,对TsHSP70-4基因密码子进行优化,连接载体pPIC9K,在毕赤酵母中进行诱导表达,通过Western-blot及质谱测序进行蛋白鉴定.将纯化后的TsHSP70-4表达蛋白免疫新西兰大白兔,制备抗血清.结果显示,成功克隆出大小为1 953 bp的TsHSP70-4ORF序列.在毕赤酵母中进行了高效表达,获得大小约为95 ku的重组蛋白.Western-blot检测和质谱鉴定表明,获得的重组蛋白即为TsHSP70-4.免疫新西兰大白兔,制备出效价高达1∶409 600的抗血清.该研究为猪囊尾蚴病新疫苗的研制提供了依据.

  11. HSP70 expression of workers exposure to power frequency electromagnetic fields in 500 kV substation. 'WEN%500kV变电站工人HSP70表达水平研究

    Institute of Scientific and Technical Information of China (English)

    温翠菊; 闫雪华; 李丽; 黎丽春; 张丹英; 邹剑明

    2011-01-01

    Objective To explore the HSP70 expression of workers exposure to power frequency electromagnetic fields in 500 kV substation. Methods 263 workers and 252 administrative staff were selected as exposed and control group, respectively. The contents of HSP70 protein in sera were determined by ELISA assay. Results The differcnce of power frequency electric field and power frequency magnetic field between exposed and control group was statistically significant (P=0.000), (P=0.000), respectively. HSP70 protein contents were (17.71±13.63) and (16.65±9.60), respectively. The difference was not statistically significant (P=0.430). Conclusion The contents of HSP70 protein of workers exposure to power frequency electromagnetic fields in 500 kV substation are no different from the control group.%目的 探讨工频电磁场对500 kV变电站工人外周血HSP70表达水平的影响,寻找可实际应用早期效应标记物.方法 选择广东省内500kV变电站263名巡检人员作为暴露人群及抽取252名各变电站行政管理人员作为对照人群,分别抽取外周血5 ml,用HSP70 ELISA试剂盒进行测定.结果 500 kV变电站工人与对照组的工频电场(t=12.455,P=0.000)、工频磁场(t=18.942,P=0.000)暴露比较差异有统计学意义.经校正后,变电站工人与对照人群外周HSP70蛋白表达水平比较差异无统计学意义(F=0.624,P=0.430).结论 长期接触工频电磁场没有引起变电站工人HSP70表达水平增加.

  12. Evaluation of Antibody Responses Elicited by Immunization of Mice with a Pneumococcal Antigen Genetically Fused to Murine HSP70 and Murine Interleukin-4

    Institute of Scientific and Technical Information of China (English)

    Dennis O. GOR; Salamatu S. MAMBULA

    2006-01-01

    The heat shock (stress) protein HSP70 has been shown to be a potent stimulator of cellular immune responses. In order to determine whether HSP70 has the ability to stimulate antibody responses, we constructed and expressed fusion proteins consisting of murine HSP70 or murine interleukin (IL)-4 covalently linked to a pneumococcal cell wall-associated protein antigen designated PpmA. Immunization of mice with the PpmA-HSP70 fusion protein (PpmA-70) failed to elicit an increased PpmA-specific serum antibody response. In contrast, mice immunized with PpmA fused to IL-4 (PpmA-IL4), or PpmA fused to both IL-4 and HSP70 (PpmA-IL4-70) fusion proteins elicited high levels of PpmA-specific antibody responses.These data suggest that HSP70 has a limited capacity to stimulate immune responses to heterologous antigens in vivo.

  13. Inhibition of HSP70 Gene Expression by Modified Antisense and Its Effects on Embryonic Sensitivity to Heat Shock

    Institute of Scientific and Technical Information of China (English)

    TIAN Wen-ru; DU Li-yin; HE Jian-bin; LI Shou-jun

    2004-01-01

    Experiments were performed to evaluate the efficiency of inhibition of HSP70 gene expression by antisense oligonucleotides complementary to the mRNA of HSP70 and to test the effects of inhibition of HSP70 gene expression on subsequent embryonic sensitivity to heat shock. The results showed that transfection of pre-implantation embryos at 4-cell stage with 5 μM antisense oligo had no effect on in vitro blastocyst development. However, transfection with 10 to 40 μM antisense oligo had reduced in vitro blastocyst development to 15, 10% and 0; For the embryos which exposed to 40 μM As arrested at the 16-cell stage, there was no blastocyst formation within the heat shock groups. In contrast, transfection had no effect on embryonic sensitivity to heat shock, above 25% of embryos developed to blastocyst stage in control groups.

  14. [SWI/SNF Protein Complexes Participate in the Initiation and Elongation Stages of Drosophila hsp70 Gene Transcription].

    Science.gov (United States)

    Mazina, M Yu; Nikolenko, Yu V; Krasnov, A N; Vorobyeva, N E

    2016-02-01

    The participation of the SWI/SNF chromatin remodeling complex in the stimulation of the RNA polymerase II binding to gene promotors was demonstrated in all model eukaryotic organisms. It was shown eight years ago that the SWI/SNF complex influence on transcription is not limited to its role in initiation but also includes participation in elongation and alternative splicing. In the current work, we describe the subunit composition of the SWI/SNF complexes participating in initiation, preparing for the elongation and elongation of hsp70 gene transcription in Drosophila melanogaster. The data reveal the high mobility of the SWI/SNF complex composition during the hsp 70 gene transcription process. We suggest a model describing the process of sequential SWI/SNF complex formation during heat-shock induced transcription of the hsp 70 gene.

  15. HSP70 Over Expression Protected Neurocyte in the Area of Dentate Gyrus(DG)of Rat's Hippocampus From Hypoxia-Induced Apoptosis%HSP70高表达保护低氧诱导的大鼠海马DG区神经细胞凋亡

    Institute of Scientific and Technical Information of China (English)

    罗芳; 闫文; 杨银书; 卢娟; 徐强; 董大海; 徐文; 高琨; 高丽莉; 侯顺利; 左晶; 刘红

    2012-01-01

    Objective:To investigate the effect of the over expression of HSP70 on neurocyte in the area of dentate gyrus(DG)of rat's hippocampus in hypoxia-induced apoptosis. Methods: Cells were either heated in 41℃ water for 2 hours or affected with 10μmol/L sodium arsenite for 18h to induced the over expression of HSP70. While the control cells are added HSP70 antisense oligomer which reduced the HSP70 expression or not induced before the cells heated. Then the cells were incubated in hypoxia. Fragmentation of DNA was quantities, and HSP70 was analyzed at various times by the Western blot. Results: Both heat and sodium arsenite can induce the over expression of HSP70, which can significantly decreased hypoxia-induced apoptosis. Introduction of the HSP70 antisense oligomer prior to heating reserved the protective effect of HSP70. Conclusions: The over expression of HSP70 can protect the pre-blood cells from radiation-induced apoptosis.%目的:研究诱导HSP70高表达对低氧引起的大鼠海马DG区神经细胞凋亡的保护作用.方法:大鼠海马DG区神经细胞分别在41℃温浴2h和加入砷酸钠诱导HSP70高表达.对照低氧而不预热,并用HSP70反义寡核苷酸链抑制HSP70合成,观察HSP70与低氧大鼠海马DG区神经细胞凋亡的关系.DNA碎片法检测细胞凋亡,Western Blotting检测HSP70.结果:预热和砷酸钠都可以诱导细胞HSP70高表达;HSP70高表达可以明显减少低氧诱导的细胞凋亡.在预热前导入HSP70反义核酸,可以降低HSP70抑制细胞凋亡的作用.结论:HSP70高表达可以保护细胞由于低氧引起的细胞凋亡.

  16. Sperm sorting procedure induces a redistribution of Hsp70 but not Hsp60 and Hsp90 in boar spermatozoa.

    Science.gov (United States)

    Spinaci, Marcella; Volpe, Sara; Bernardini, Chiara; de Ambrogi, Marco; Tamanini, Carlo; Seren, Eraldo; Galeati, Giovanna

    2006-01-01

    Heat shock proteins, besides their protective function against stresses, have been recently indicated as key factors for sperm fertilizing ability. Since sexing sperm by high-speed flow-cytometry subjects them to different physical, mechanical, and chemical stresses, the present study was designed to verify, by immunofluorescence and Western blot, whether the sorting procedure induces any modification in the amount and cellular distribution of heat shock proteins 60, 70, and 90 (Hsp60, Hsp70, Hsp90). Immunolocalization and Western blot quantification of both Hsp60 and Hsp90 did not reveal differences between unsorted and sorted semen. On the contrary, a redistribution of Hsp70 immunoreactivity from the equatorial subsegment toward the equator of sperm cells was recorded after sorting; this relocation suggests capacitation-like changes of sperm membrane. This modification seems to be caused mainly by incubation with Hoechst 33342, while both passage of sperm through flow cytometer and laser beam represent only minor stimuli. A further Hsp70 redistribution seems to be due to the final steps of sperm sorting, charging, and deflection of drops, and to the dilution during collection. On the other hand, staining procedure and mechanical stress seem to be the factors most injurious to sperm viability. Moreover, Hsp70 relocation was deeply influenced by the storage method. In fact, storing sexed spermatozoa, after centrifugation, in a small volume in presence of seminal plasma induced a reversion of Hsp70 redistribution, while storage in the diluted catch fluid of collection tubes caused Hsp70 relocation in most sorted spermatozoa. PMID:16870948

  17. Batroxobin Against Anoxic Damage of Rat Hippocampal Neurons in Culture: Morphological Changes and Hsp70 Expression

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    @@Batroxobin,the thrombin-like enzyme,is used for therapeutic defibrination. We have found that batroxobin has good therapeutic effect in ischemic reperfusion rats and clinical practices in vivo. But we have not studied the neuroprotective effect of batroxobin on anoxic hippocampal neurons in vitro. The purpose of this study was to obtain further information on the mechanism of the batroxobin-induced neuroprotection and examine the neuroprotective effect on neurons exposed to anoxia. The effect of batroxobin on anoxic damages in cultured hippocampal neurons of neonatal rats was investigated by using morphological changes and heat shock protein 70Kd (Hsp70) immunoreactive expression as indicators. The results indicate that batroxobin, besides its defibrination, may have a direct neuroprotective effect on anoxic damage of hippocampal neurons.

  18. HSP70 mediates survival in apoptotic cells-Boolean network prediction and experimental validation.

    Science.gov (United States)

    Vasaikar, Suhas V; Ghosh, Sourish; Narain, Priyam; Basu, Anirban; Gomes, James

    2015-01-01

    Neuronal stress or injury results in the activation of proteins, which regulate the balance between survival and apoptosis. However, the complex mechanism of cell signaling involving cell death and survival, activated in response to cellular stress is not yet completely understood. To bring more clarity about these mechanisms, a Boolean network was constructed that represented the apoptotic pathway in neuronal cells. FasL and neurotrophic growth factor (NGF) were considered as inputs in the absence and presence of heat shock proteins known to shift the balance toward survival by rescuing pro-apoptotic cells. The probabilities of survival, DNA repair and apoptosis as cellular fates, in the presence of either the growth factor or FasL, revealed a survival bias encoded in the network. Boolean predictions tested by measuring the mRNA level of caspase-3, caspase-8, and BAX in neuronal Neuro2a (N2a) cell line with NGF and FasL as external input, showed positive correlation with the observed experimental results for survival and apoptotic states. It was observed that HSP70 contributed more toward rescuing cells from apoptosis in comparison to HSP27, HSP40, and HSP90. Overexpression of HSP70 in N2a transfected cells showed reversal of cellular fate from FasL-induced apoptosis to survival. Further, the pro-survival role of the proteins BCL2, IAP, cFLIP, and NFκB determined by vertex perturbation analysis was experimentally validated through protein inhibition experiments using EM20-25, Embelin and Wedelolactone, which resulted in 1.27-, 1.26-, and 1.46-fold increase in apoptosis of N2a cells. The existence of a one-to-one correspondence between cellular fates and attractor states shows that Boolean networks may be employed with confidence in qualitative analytical studies of biological networks.

  19. Effect of vicanicin and protolichesterinic acid on human prostate cancer cells: role of Hsp70 protein.

    Science.gov (United States)

    Russo, A; Caggia, S; Piovano, M; Garbarino, J; Cardile, V

    2012-01-01

    With the aim of identifying novel agents with antigrowth and pro-apoptotic activity on prostate cancer cells, in the present study, we evaluated the effect of five lichen secondary metabolites the depsides atranorin (1), diffrattaic (2) and divaricatic (3) acids, the depsidone vicanicin (4) and the protolichesterinic acid (5) on cell growth in androgen-sensitive (LNCaP) and androgen-insensitive (DU-145) human prostate cancer cells. The cell viability was measured using MTT assay. LDH release, a marker of membrane breakdown, was also measured. For the detection of apoptosis, the evaluation of DNA fragmentation (COMET assay) and caspase-3 activity assay were employed. The expression of Bcl-2, Bax, TRAIL, COX-2, NOS2 and Hsp70 proteins was detected by western blot analysis. Generation of reactive oxygen species was measured by using a fluorescent probe. It was observed that atranorin (1), diffrattaic (2) and divaricatic (3) acids showed a lower activity inhibiting the prostate cancer cells only at more high concentrations (25 and 50μM). Whereas compounds vicanicin (4) and protolichesterinic acid (5) showed a dose-response relationship in the range of 6.25-50μM concentrations in DU-145 and LNCaP cells, activating an apoptotic process. The novel finding, in the present study, is that apoptosis induced by these compounds appears to be mediated, at least in part, via the inhibition of Hsp70 expression, that may be correlated with a modulation of redox-sensitive mechanisms. The combination of vicanicin (4) and protolichesterinic acid (5) with other anti-prostate cancer therapies could be considered a promising strategy that warrants further in vivo evaluation. PMID:22063921

  20. Potential Response to Selection of HSP70 as a Component of Innate Immunity in the Abalone Haliotis rufescens

    Science.gov (United States)

    Brokordt, Katherina B.; González, Roxana C.; Farías, William J.; Winkler, Federico M.

    2015-01-01

    Assessing components of the immune system may reflect disease resistance. In some invertebrates, heat shock proteins (HSPs) are immune effectors and have been described as potent activators of the innate immune response. Several diseases have become a threat to abalone farming worldwide; therefore, increasing disease resistance is considered to be a long-term goal for breeding programs. A trait will respond to selection only if it is determined partially by additive genetic variation. The aim of this study was to estimate the heritability (h2) and the additive genetic coefficient of variation (CVA) of HSP70 as a component of innate immunity of the abalone Haliotis rufescens, in order to assess its potential response to selection. These genetic components were estimated for the variations in the intracellular (in haemocytes) and extracellular (serum) protein levels of HSP70 in response to an immunostimulant agent in 60 full-sib families of H. rufescens. Levels of HSP70 were measured twice in the same individuals, first when they were young and again when they were pre-harvest adults, to estimate the repeatability (R), the h2 and the potential response to selection of these traits at these life stages. High HSP70 levels were observed in abalones subjected to immunostimulation in both the intracellular and extracellular haemolymph fractions. This is the first time that changes in serum levels of HSP70 have been reported in response to an immune challenge in molluscs. HSP70 levels in both fractions and at both ages showed low h2 and R, with values that were not significantly different from zero. However, HSP70 induced levels had a CVA of 13.3–16.2% in young adults and of 2.7–8.1% in pre-harvest adults. Thus, despite its low h2, HSP70 synthesis in response to an immune challenge in red abalone has the potential to evolve through selection because of its large phenotypic variation and the presence of additive genetic variance, especially in young animals. PMID

  1. Correlation of Hsp70 Serum Levels with Gross Tumor Volume and Composition of Lymphocyte Subpopulations in Patients with Squamous Cell and Adeno Non-Small Cell Lung Cancer

    OpenAIRE

    Gunther, Sophie; Ostheimer, Christian; Stangl, Stefan; Specht, Hanno M; Mozes, Petra; Jesinghaus, Moritz; Vordermark, Dirk; Combs, Stephanie E; Peltz, Friedhelm; Jung, Max P.; Multhoff, Gabriele

    2015-01-01

    Heat-shock protein 70 (Hsp70) is frequently found on the plasma membrane of a large number of malignant tumors including non-small cell lung cancer (NSCLC) and gets released into the blood circulation in lipid vesicles. On the one hand, a membrane (m)Hsp70-positive phenotype correlates with a high aggressiveness of the tumor; on the other hand, mHsp70 serves as a target for natural killer (NK) cells that had been pre-stimulated with Hsp70-peptide TKD plus low-dose interleukin-2 (TKD/IL-2). Fo...

  2. Cloning and procaryotic expression of HSP70 gene of bovine Theileria sergenti%牛瑟氏泰勒虫HSP70基因的克隆及原核表达

    Institute of Scientific and Technical Information of China (English)

    金超; 贾立军; 薛书江; 王娜; 钱年超; 张守发

    2010-01-01

    根据GenBank(D12692.1)上发表的牛瑟氏泰勒虫HSP70基因序列设计、合成1对特异性的引物,采用PCR方法扩增牛瑟氏泰勒虫HSP70基因片段,将扩增产物与pMD18-T simple载体连接,重组质粒经PCR、酶切鉴定后测序;用pGEX-4T-1表达栽体构建重组质粒pGEX-HSP70,经IPTG诱导表达后进行SDS-PAGE、Western-blot分析.结果表明:扩增的HSP70基因与D12692.1序列同源性为99.3%;表达的融合蛋白分子质量约为68 ku,而且可与牛瑟氏泰勒虫阳性血清发生特异性反应.

  3. Expression and significance of MGMT and HSP70 in colorectal carcinoma%MGMT和HSP70在大肠癌中的表达及临床意义

    Institute of Scientific and Technical Information of China (English)

    吉艳荣; 薄爱华; 吉彦丽; 张彦; 牛书雷; 冯丹

    2009-01-01

    目的:探讨热休克蛋白(HSP)70和DNA修复酶O6-甲基鸟嘌呤-DNA甲基转移酶(O6-methyguanine-DNA methytransferase,MGMT)在大肠癌中的表达及临床意义.方法:采用免疫组织化学SABC法检测61例大肠癌组织中MGMT与HSP70的表达情况.结果:MGMT与HSP70在大肠癌中表达的阳性率分别为50.82%和62.30%.其中,二者都与性别及分化程度相关性不明显(P>0.05);而与年龄和浸润深度有相关性(P<0.05),且二者呈正相关(P<0.05;r =0.32).结论:MGMT与HSP70在大肠癌的发生发展中可能起重要作用,检测它们对大肠癌的防治有重要的临床指导意义.

  4. 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. PMID:24950414

  5. 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...

  6. ELEVATED HSP70 STIMULATES CELL PROLIFERATION AND IS CYTOPROTECTIVE AGAINST HEAT AND VINCRISTINE TOXICITY IN MCF-7 CELLS.

    Science.gov (United States)

    Heat-shock proteins (HSPs) play important roles in regulating cell growth and protecting cells from adverse effects of heat and chemical stress. In many types of cancer, elevated HSP70 levels are associated with poor prognosis and resistance to chemotherapeutic agents. In the pre...

  7. Safrole oxide induced neuronal differentiation of rat bone-marrow mesenchymal stem cells by elevating Hsp70.

    Science.gov (United States)

    Zhao, YanChun; Xin, Jie; Sun, ChunHui; Zhao, BaoXiang; Zhao, Jing; Su, Le

    2012-11-01

    In a previous study, we found that at low concentrations, safrole oxide (SFO) could induce vascular endothelial cell (VEC) transdifferentiation into neuron-like cells; however, whether SFO could induce bone-marrow mesenchymal stem cell (BMSC) neural differentiation was unknown. Here, we found that SFO could effectively induce BMSC neural differentiation in the presence of serum and fibroblast growth factor 2 and did not affect cell viability at low concentrations. The levels of neuron-specific enolase and neurofilament-L were increased greatly, but that of glial fibrillary acidic protein was absent with SFO treatment for 48h. Furthermore, SFO could increase the level of heat shock protein 70 (Hsp70), an important factor in neuronal differentiation. Knockdown of Hsp70 by its small interfering RNA blocked SFO-induced BMSC differentiation. Thus, SFO is a novel inducer of BMSC differentiation to neuron-like cells and Hsp70 is implicated in the differentiation process. We provide a new tool for obtaining neuron-like cells from BMSCs and for further investigating the new effect of Hsp70 on BMSC neuronal differentiation.

  8. 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...

  9. A small molecule inhibitor for ATPase activity of Hsp70 and Hsc70 enhances the immune response to protein antigens

    Science.gov (United States)

    Baek, Kyung-Hwa; Zhang, Haiying; Lee, Bo Ryeong; Kwon, Young-Guen; Ha, Sang-Jun; Shin, Injae

    2015-12-01

    The ATPase activities of Hsp70 and Hsc70 are known to be responsible for regulation of various biological processes. However, little is known about the roles of Hsp70 and Hsc70 in modulation of immune responses to antigens. In the present study, we investigated the effect of apoptozole (Az), a small molecule inhibitor of Hsp70 and Hsc70, on immune responses to protein antigens. The results show that mice administered with both protein antigen and Az produce more antibodies than those treated with antigen alone, showing that Az enhances immune responses to administered antigens. Treatment of mice with Az elicits production of antibodies with a high IgG2c/IgG1 ratio and stimulates the release of Th1 and Th2-type cytokines, suggesting that Az activates the Th1 and Th2 immune responses. The observations made in the present study suggest that inhibition of Hsp70 and Hsc70 activities could be a novel strategy designing small molecule-based adjuvants in protein vaccines.

  10. hELP3 Subunit of the Elongator Complex Regulates the Transcription of HSP70 Gene in Human Cells

    Institute of Scientific and Technical Information of China (English)

    Qiuju HAN; Xiaozhe HOU; Dongmei SU; Lina PAN; Jizhou DUAN; Liguo CUI; Baiqu HUANG; Jun LU

    2007-01-01

    The human Elongator complex is remarkably similar to its yeast counterpart in several aspects.In a previous study, we analyzed the functions of the human elongation protein 3 (hELP3) subunit of the human Elongator by using an in vivo yeast complementation system. However, direct evidence for hELP3 functions in regulating gene expression in human cells was not obtained. In this study, we used hELP3 antisense oligonucleotide inhibitors to knock down hELP3 gene expression to investigate its function in human 293T cells. The results showed that specific reduction of hELP3 mRNA and protein caused a significant suppression of HSP70-2 gene expression, and this was accompanied by histone H3 hypoacetylation and decreased RNA polymerase Ⅱ density at the HSP70-2 gene. Moreover, the data also showed that hELP3 exerted the transcriptional regulatory function directly through its presence on the HSP70-2 gene. Data presented in this report provide further insight and direct evidence of the functions of hELP3 in HSP70-2 gene transcriptional elongation in human cells.

  11. BiPPred: Combined sequence- and structure-based prediction of peptide binding to the Hsp70 chaperone BiP.

    Science.gov (United States)

    Schneider, Markus; Rosam, Mathias; Glaser, Manuel; Patronov, Atanas; Shah, Harpreet; Back, Katrin Christiane; Daake, Marina Angelika; Buchner, Johannes; Antes, Iris

    2016-10-01

    Substrate binding to Hsp70 chaperones is involved in many biological processes, and the identification of potential substrates is important for a comprehensive understanding of these events. We present a multi-scale pipeline for an accurate, yet efficient prediction of peptides binding to the Hsp70 chaperone BiP by combining sequence-based prediction with molecular docking and MMPBSA calculations. First, we measured the binding of 15mer peptides from known substrate proteins of BiP by peptide array (PA) experiments and performed an accuracy assessment of the PA data by fluorescence anisotropy studies. Several sequence-based prediction models were fitted using this and other peptide binding data. A structure-based position-specific scoring matrix (SB-PSSM) derived solely from structural modeling data forms the core of all models. The matrix elements are based on a combination of binding energy estimations, molecular dynamics simulations, and analysis of the BiP binding site, which led to new insights into the peptide binding specificities of the chaperone. Using this SB-PSSM, peptide binders could be predicted with high selectivity even without training of the model on experimental data. Additional training further increased the prediction accuracies. Subsequent molecular docking (DynaDock) and MMGBSA/MMPBSA-based binding affinity estimations for predicted binders allowed the identification of the correct binding mode of the peptides as well as the calculation of nearly quantitative binding affinities. The general concept behind the developed multi-scale pipeline can readily be applied to other protein-peptide complexes with linearly bound peptides, for which sufficient experimental binding data for the training of classical sequence-based prediction models is not available. Proteins 2016; 84:1390-1407. © 2016 Wiley Periodicals, Inc.

  12. Adaptive capability as indicated by behavioral and physiological responses, plasma HSP70 level, and PBMC HSP70 mRNA expression in Osmanabadi goats subjected to combined (heat and nutritional) stressors

    Science.gov (United States)

    Shilja, Shaji; Sejian, V.; Bagath, M.; Mech, A.; David, C. G.; Kurien, E. K.; Varma, Girish; Bhatta, Raghavendra

    2016-09-01

    A study was conducted to assess the impact of heat and nutritional stress simultaneously on the adaptive capability as indicated by behavioral and physiological responses, plasma heat shock protein 70 (HSP70) level, and peripheral blood mononuclear cells (PBMC) HSP70 gene expression in goats. Twenty-four adult Osmanabadi bucks (average body weight (BW) 16.0 kg) were used in the present study. The bucks were divided into four groups viz., C ( n = 6; control), HS ( n = 6; heat stress), NS ( n = 6; nutritional stress), and CS ( n = 6; combined stress). The study was conducted for a period of 45 days. C and HS bucks had ad libitum access to their feed while NS and CS bucks were under restricted feed (30 % intake of C bucks) to induce nutritional stress. The HS and CS bucks were exposed to solar radiation for 6 h a day between 10:00 a.m. and 4:00 p.m. to induce heat stress. The data was analyzed using repeated measures analysis of variance. The standing time differed significantly ( P groups (C and HS) and restricted feeding groups (NS and CS). The highest ( P group while the lowest in the C and HS groups. The highest ( P group. Water intake recorded was significantly ( P groups. The highest respiration rate (RR), pulse rate (PR), and rectal temperature (RT) during the afternoon were also recorded in the CS group. Further, skin temperature of the head, flank, and scrotum during the afternoon was also higher ( P group. In addition, both plasma HSP70 concentration and PBMC HSP70 messenger RNA (mRNA) transcript expression were also significantly ( P group. It can be concluded from this study that when two stressors occur simultaneously, they may have severe impact on adaptive capabilities of Osmanabadi bucks as compared to that would occur individually. Further, the study indicated that lying time, drinking frequency, RR, RT, plasma HSP70, and PBMC HSP70 gene expression may act as ideal biological markers for assessing the impact of CS on adaptive capabilities in bucks.

  13. Adaptive capability as indicated by behavioral and physiological responses, plasma HSP70 level, and PBMC HSP70 mRNA expression in Osmanabadi goats subjected to combined (heat and nutritional) stressors

    Science.gov (United States)

    Shilja, Shaji; Sejian, V.; Bagath, M.; Mech, A.; David, C. G.; Kurien, E. K.; Varma, Girish; Bhatta, Raghavendra

    2015-12-01

    A study was conducted to assess the impact of heat and nutritional stress simultaneously on the adaptive capability as indicated by behavioral and physiological responses, plasma heat shock protein 70 (HSP70) level, and peripheral blood mononuclear cells (PBMC) HSP70 gene expression in goats. Twenty-four adult Osmanabadi bucks (average body weight (BW) 16.0 kg) were used in the present study. The bucks were divided into four groups viz., C (n = 6; control), HS (n = 6; heat stress), NS (n = 6; nutritional stress), and CS (n = 6; combined stress). The study was conducted for a period of 45 days. C and HS bucks had ad libitum access to their feed while NS and CS bucks were under restricted feed (30 % intake of C bucks) to induce nutritional stress. The HS and CS bucks were exposed to solar radiation for 6 h a day between 10:00 a.m. and 4:00 p.m. to induce heat stress. The data was analyzed using repeated measures analysis of variance. The standing time differed significantly (P groups (C and HS) and restricted feeding groups (NS and CS). The highest (P group while the lowest in the C and HS groups. The highest (P group. Water intake recorded was significantly (P groups. The highest respiration rate (RR), pulse rate (PR), and rectal temperature (RT) during the afternoon were also recorded in the CS group. Further, skin temperature of the head, flank, and scrotum during the afternoon was also higher (P group. In addition, both plasma HSP70 concentration and PBMC HSP70 messenger RNA (mRNA) transcript expression were also significantly (P group. It can be concluded from this study that when two stressors occur simultaneously, they may have severe impact on adaptive capabilities of Osmanabadi bucks as compared to that would occur individually. Further, the study indicated that lying time, drinking frequency, RR, RT, plasma HSP70, and PBMC HSP70 gene expression may act as ideal biological markers for assessing the impact of CS on adaptive capabilities in bucks.

  14. Hsp70-Hsp40 chaperone complex functions in controlling polarized growth by repressing Hsf1-driven heat stress-associated transcription.

    Directory of Open Access Journals (Sweden)

    Aleksandar Vjestica

    Full Text Available How the molecular mechanisms of stress response are integrated at the cellular level remains obscure. Here we show that the cellular polarity machinery in the fission yeast Schizosaccharomyces pombe undergoes dynamic adaptation to thermal stress resulting in a period of decreased Cdc42 activity and altered, monopolar growth. Cells where the heat stress-associated transcription was genetically upregulated exhibit similar growth patterning in the absence of temperature insults. We identify the Ssa2-Mas5/Hsp70-Hsp40 chaperone complex as repressor of the heat shock transcription factor Hsf1. Cells lacking this chaperone activity constitutively activate the heat-stress-associated transcriptional program. Interestingly, they also exhibit intermittent monopolar growth within a physiological temperature range and are unable to adapt to heat stress. We propose that by negatively regulating the heat stress-associated transcription, the Ssa2-Mas5 chaperone system could optimize cellular growth under different temperature regiments.

  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. 基于Hsp70基因的马梨形虫分类学定位分析%Taxonomic Status Assays of Theileria equi Based on Hsp70 Gene

    Institute of Scientific and Technical Information of China (English)

    罗金; 刘光远; 田占成; 谢俊仁; 张萍; 沈辉; 党根生

    2011-01-01

    [Objective] The objective of the study is to determine the taxonomic status of Theileria equi (T. equi). [Method] According to the Hsp70 gene sequence (AB248743.1) of Babesia equi (B.equi) in GenBank, the primers TeHsp70F and TeHsp70R were designed. And a fragment with the length of 1 920 bp was obtained by PCR. The gene sequence was compared with other 23 amino acids sequences. [Result] The fragment encods 639 amino acids, 208 hydrophobic amino acids and 167 polar amino acid. Identity analysis showed that T. equi has a closest relationship with B.equi (BAF0262S.1), and has a little relationship with T. parva (XP764717.1) and T. annulata (AAA30130.1), while T. equi has a distant relationship with B. caballi (BAF02619.1), which infects equine. [ Conclusion ] It was a correct taxonomic status for B. equi to Theileria species. And the taxonomic status was error before, for T. equi to Babesia species. In this paper, a proof was provided for taxonomic status of Theileria equi.%[目的]为了进一步确定马泰勒虫的分类学定位.[方法]根据GenBank上登录的马巴贝斯虫Hsp70基因序列(AB248743.1),设计引物TeHsp70F,TeHsp70R,以马泰勒虫基因组DNA为模板进行扩增,获得全长为1920 bp的核酸片段.将该基因序列与GenBank中23种已知虫种的相应序列进行分析比较.[结果]该片段编码639个氨基酸,其疏水性氨基酸达到208个,极性氨基酸167个.同一性分析显示,该基因片段与报道的马巴贝斯虫Hsp70基因(B.equi BAF02625.1)亲缘关系最近,其次是小泰勒虫(T.parva XP764717.1)和环形泰勒虫(T.annulataAAA30130.1),而与感染马属动物的另一个虫种驽巴贝斯虫(B.caballi BAF02619.1)亲缘关系较远.[结论]马泰勒虫Hsp70基因的克隆及系统发育分析显示,之前被定名为马巴贝斯虫(B.equi)的虫种隶属于泰勒虫虫种.本试验为马巴贝斯虫正确更名为马泰勒虫(T.equi)的分类地位提供了又一佐证.

  17. HSP70在鼠背轴型皮管IPC后的保护作用的实验研究%Study of HSP70 protective effect on ischemic preconditioning axial flap in rats

    Institute of Scientific and Technical Information of China (English)

    陈劲松; 夏双印; 杨大平

    2001-01-01

    Objective This study was to reveal the continuous changes of HSP70 and its regularity in the delayed protection of ischemic preconditioning axial flap in rats,then speculate the best time- piont of protection. Methods Animals were divided into three groups randomly(control、amputate after isehemia pretreatment and amputate delay after ischemia pretreatment). A left dorsal skin flap was crated in Wistar rat to ischemia precondition model to examine the survival rate of the skin flaps, electronic microscopic appearanee, the contents of HSP70 at various times after they were given different treatment. Results The survival rate of amputate after ischamia pretreatment and amPutate delay 48 h after ischemia pretreatment are 1.26 and 1.57 times as high as that of control. The celluar biosynthesis at 48 h after IPC is vigorous when there is a lot of proteins which were synthesized and stored. The contents of HSP70 at 48 h after IPC or at 72 h after IPC is much more than that of control, especially at the 48 h. Conclusions IPC can improve the survival rate of ischemia flap and amputate delay after IPC has more effect than IPC does. 48 h may be the best time - point of IPC protection. It is suggest that the mechanism of IPC protection can be related to the function of HSP70.%目的揭示IPC后皮瓣中延迟相HSP70表达的动态变化,提出其规律及保护作用的最佳时间。方法采用大鼠背部轴型皮管,制成活体原位IPC模型,检测皮管存活率,组织细胞超微结构,HSP70测定及统计学分析。结果 IPC组和IPC后延迟48h断蒂组与对照组相比分别提高了皮瓣存活面积1.26倍和1.57倍;IPC后48h组细胞合成功能旺盛,有大量的蛋白质合成并贮存;IPC后48、72 h组HSP70均较对照组升高,并且其表达以48h较为明显。结论 IPC对鼠背轴型皮管具有保护作用,而对延迟断蒂组保护作用更佳,以48h为最佳时间点。HSP70可能发挥着重要的作用。

  18. The Chaperone Balance Hypothesis: The Importance of the Extracellular to Intracellular HSP70 Ratio to Inflammation-Driven Type 2 Diabetes, the Effect of Exercise, and the Implications for Clinical Management

    Directory of Open Access Journals (Sweden)

    Mauricio Krause

    2015-01-01

    Full Text Available Recent evidence shows divergence between the concentrations of extracellular 70 kDa heat shock protein [eHSP70] and its intracellular concentrations [iHSP70] in people with type 2 diabetes (T2DM. A vital aspect regarding HSP70 physiology is its versatility to induce antagonistic actions, depending on the location of the protein. For example, iHSP70 exerts a powerful anti-inflammatory effect, while eHSP70 activates proinflammatory pathways. Increased eHSP70 is associated with inflammatory and oxidative stress conditions, whereas decreased iHSP70 levels are related to insulin resistance in skeletal muscle. Serum eHSP70 concentrations are positively correlated with markers of inflammation, such as C-reactive protein, monocyte count, and TNF-α, while strategies to enhance iHSP70 (e.g., heat treatment, chemical HSP70 inducers or coinducers, and physical exercise are capable of reducing the inflammatory profile and the insulin resistance state. Here, we present recent findings suggesting that imbalances in the HSP70 status, described by the [eHSP70]/[iHSP70] ratio, may be determinant to trigger a chronic proinflammatory state that leads to insulin resistance and T2DM development. This led us to hypothesize that changes in this ratio value could be used as a biomarker for the management of the inflammatory response in insulin resistance and diabetes.

  19. Correlation of Hsp70 serum levels with gross tumor volume and composition of lymphocyte subpopulations in patients with squamous cell and adeno non-small cell lung cancer

    Directory of Open Access Journals (Sweden)

    Sophie eGunther

    2015-11-01

    Full Text Available Heat shock protein 70 (Hsp70 is frequently found on the plasma membrane of a large number of malignant tumors including non-small cell lung cancer (NSCLC and gets released into the blood circulation in lipid vesicles. On the one hand a membrane (mHsp70-positive phenotype correlates with a high aggressiveness of the tumor, on the other hand, mHsp70 serves as a target for natural killer (NK cells that had been pre-stimulated with Hsp70-peptide TKD plus low dose interleukin-2 (TKD/IL-2. Following activation, NK cells show an up-regulated expression of activatory C-type lectin receptors, such as CD94/NKG2C, NKG2D and natural cytotoxicity receptors (NCRs; NKp44, NKp46, NKp30 and thereby gain the capacity to kill mHsp70-positive tumor cells. With respect to these results, the efficacy of ex vivo TKD/IL-2 stimulated, autologous NK cells is currently tested in a proof-of-concept phase II clinical trial in patients with squamous cell NSCLC after RCT at the TUM. Inclusion criteria are histological proven, non-resectable NSCLC in stage IIIA/IIIB, clinical responses to RCT and a mHsp70-positive tumor phenotype. The mHsp70 status is determined in the serum of patients using the lipHsp70 ELISA test which enables the quantification of liposomal and free Hsp70. Squamous cell and adeno NSCLC patients had significantly higher serum Hsp70 levels than healthy controls. A significant correlation of serum Hsp70 levels with the gross tumor volume (GTV was shown for adeno and squamous cell NSCLC. However, significantly elevated ratios of activated CD69+/CD94+ NK cells that are associated with low serum Hsp70 levels were observed only in patients with squamous cell lung cancer. These data might provide a first hint that the innate immune system might get activated by soluble Hsp70 in squamous cell but not adeno NSCLC.

  20. Ha-ras(val12) induces HSP70b transcription via the HSE/HSF1 system, but HSP70b expression is suppressed in Ha-ras(val12)-transformed cells.

    Science.gov (United States)

    Stanhill, A; Levin, V; Hendel, A; Shachar, I; Kazanov, D; Arber, N; Kaminski, N; Engelberg, D

    2006-03-01

    Heat shock proteins (Hsps) are overexpressed in many tumors, but are downregulated in some tumors. To check for a direct effect of Ha-Ras(val12) on HSP70 transcription, we transiently expressed the oncoprotein in Rat1 fibroblasts and monitored its effect on HSP70b promoter-driven reporter gene. We show that expression of Ha-Ras(val12) induced this promoter. Promoter analysis via systematic deletions and point mutations revealed that Ha-Ras(val12) induces HSP70b transcription via heat shock elements (HSEs). Also, Ha-Ras(val12) induction of HSE-mediated transcription was dramatically reduced in HSF1-/- cells. Yet, residual effect of Ha-Ras(val12) that was still measured in HSF1-/- cells suggests that some of the Ha-Ras(val12) effect is Hsf1-independent. When HSF1-/- cells, stably expressing Ha-Ras(val12), were grown on soft agar only small colonies were formed suggesting a role for heat shock factor 1 (Hsf1) in Ha-Ras(val12)-mediated transformation. Although Ha-ras(Val12) seems to be an inducer of HSP70's expression, we found that in Ha-ras(Val12-)transformed fibroblasts expression of this gene is suppressed. This suppression is correlated with higher sensitivity of Ha-ras(val12)-transformed cells to heat shock. We suggest that Ha-ras(Val12) is involved in Hsf1 activation, thereby inducing the cellular protective response. Cells that repress this response are perhaps those that acquire the capability to further proliferate and become transformed clones.

  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

    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 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 and...

  2. Modelling the role of the Hsp70/Hsp90 system in the maintenance of protein homeostasis.

    Directory of Open Access Journals (Sweden)

    Carole J Proctor

    Full Text Available Neurodegeneration is an age-related disorder which is characterised by the accumulation of aggregated protein and neuronal cell death. There are many different neurodegenerative diseases which are classified according to the specific proteins involved and the regions of the brain which are affected. Despite individual differences, there are common mechanisms at the sub-cellular level leading to loss of protein homeostasis. The two central systems in protein homeostasis are the chaperone system, which promotes correct protein folding, and the cellular proteolytic system, which degrades misfolded or damaged proteins. Since these systems and their interactions are very complex, we use mathematical modelling to aid understanding of the processes involved. The model developed in this study focuses on the role of Hsp70 (IPR00103 and Hsp90 (IPR001404 chaperones in preventing both protein aggregation and cell death. Simulations were performed under three different conditions: no stress; transient stress due to an increase in reactive oxygen species; and high stress due to sustained increases in reactive oxygen species. The model predicts that protein homeostasis can be maintained during short periods of stress. However, under long periods of stress, the chaperone system becomes overwhelmed and the probability of cell death pathways being activated increases. Simulations were also run in which cell death mediated by the JNK (P45983 and p38 (Q16539 pathways was inhibited. The model predicts that inhibiting either or both of these pathways may delay cell death but does not stop the aggregation process and that eventually cells die due to aggregated protein inhibiting proteasomal function. This problem can be overcome if the sequestration of aggregated protein into inclusion bodies is enhanced. This model predicts responses to reactive oxygen species-mediated stress that are consistent with currently available experimental data. The model can be used to

  3. 汉坦病毒诱导人脐静脉内皮细胞HSP70的表达及意义

    Institute of Scientific and Technical Information of China (English)

    余璐; 马恒; 刘彦仿; 杨守京

    2005-01-01

    目的:研究人脐静脉内皮细胞(HUVEC)感染汉坦病毒(Hantavirus,HTV)后应激反应的规律及意义.方法:采用免疫细胞化学染色法和核酸分子原位杂交,检测热休克蛋白70(HSP70)的表达;用RT-PCR观察HSP70 mRNA水平变化的规律.结果:HTV感染HUVEC后,免疫细胞化学染色法可检出HSP70呈高表达,原位杂交发现细胞质内有HSP70 mRNA的阳性信号.感染后不同时间点RT-PCR的结果表明,与对照组相比较,HSP70 mRNA的水平在感染后迅速升高并持续高表达(P<0.05).结论:HTV可诱导HUVEC高表达HSP70HSP70可能具有抑制病毒复制和保护内皮细胞的作用.

  4. Transcription factor TFIID recognizes DNA sequences downstream of the TATA element in the Hsp70 heat shock gene.

    OpenAIRE

    Emanuel, P A; Gilmour, D S

    1993-01-01

    The interaction between the Hsp70 heat shock gene promoter and a Drosophila protein complex which contains the TATA-binding protein depends on sequence-specific interactions located in the region downstream of the transcription start site. Immunopurification of the complex through the use of antibodies against the TATA-binding protein reveals that the complex is transcription factor TFIID. Binding assays with the immunopurified TFIID confirm that sequence-specific contacts are made in the reg...

  5. Comparative Study on the Immunogenicity between Hsp70 DNA Vaccine and Hsp65 DNA Vaccine in Human Mycobacterium Tuberculosis

    Institute of Scientific and Technical Information of China (English)

    DAI; Wuxing; HUANG; Hailang; YUAN; Ye; HU; Jiajie; HUANGFU; Yongmu

    2001-01-01

    The BALB/c mice were immunized with Hsp70 DNA and Hsp65 DNA vaccines in human Mycobacterium tuberculosis. Eight weeks after immunization, the eyeballs were removed, blood and spleen taken, and intraperitoneal macrophages were harvested. The lymphocytic stimulating index(SI) was used to measure the cellular proliferating ability and NO release to measure the phagocytic activity of the macrophages. With ELISA kit, the levels of interleukin-2 (IL-2) and interferon-γ(IFN-γ) in serum and the splenic lymphocytic cultured supernatant were detected. The results showed that after the mice were immunized with 100 μg/mouse of Hsp70 DNA vaccine intramuscularly, the splenic lymphocytic proliferating ability in the mice was significantly increased as compared with that in the control group, vector group and Hsp65 DNA vaccine group (P<0. 01); The contents of NO in the intraperitoneal macrophages of the mice were significantly lower than in the control group and Hsp65 DNA vaccine group (P<0. 01); The levels of serum IL-2 in the mice were significantly higher than in the control group, but there was no statistical difference between Hsp65 DNA group and vector group (P>0. 05); The contents of serum IFN-γ in the mice were significantly higher than in the control group, but significantly lower than in the Hsp65 DNA vaccine group (P<0. 05). It was indicated that immunization with Hsp70 DNA vaccine could obviously enhance the immune response, but its intensity seemed inferior to Hsp65 DNA vaccine. The anti-infection mechanisms and clinical use in the future of the vaccines of Hsp70 DNA and Hsp65 DNA are worth further studying.

  6. Association of HSP70 and genotoxic damage in lymphocytes of workers exposed to coke-oven emission

    OpenAIRE

    Xiao, Chengfeng; Chen, Sheng; Li, Jizhao; Hai, Tao; Lu, Qiaofa; Sun, Enling; Wang, Ruibo; Tanguay, Robert M.; Wu, Tangchun

    2002-01-01

    Heat shock proteins (Hsps) have been reported to protect cells, tissues, and organisms against damage from a wide variety of stressful stimuli. Whether they protect against deoxyribonucleic acid (DNA) damage in individuals exposed to environmental stresses and chemical carcinogens is unknown. In the study, we investigated the association between Hsp70 levels (the most abundant mammalian Hsp) and genotoxic damage in lymphocytes of workers exposed to coke-oven emission using Western dot blot an...

  7. Fusion of Hsp70 to Mage-a1 enhances the potency of vaccine-specific immune responses

    OpenAIRE

    Jiang, Juhong; Xie, Dan; Zhang, WenMin; Xiao, Gang; Wen, Jianming

    2013-01-01

    Background Heat shock proteins (HSPs) are capable of promoting antigen presentation of chaperoned peptides through interactions with receptors on antigen presenting cells. This property of HSPs suggests a potential function as an adjuvant-free carrier to stimulate immune responses against a covalently linked fusion partner. MAGE-A1 is a likely candidate for tumor immunotherapy due to its abundant immunogenic epitopes and strict tumor specificity. To analyze the influence of HSP70 conjugation ...

  8. 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

    Full Text Available BACKGROUND: 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. METHODOLOGY/PRINCIPAL FINDINGS: 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. CONCLUSION/SIGNIFICANCE: 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.

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

    Directory of Open Access Journals (Sweden)

    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.

  10. 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. PMID:27188431

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

    Directory of Open Access Journals (Sweden)

    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.

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

    Directory of Open Access Journals (Sweden)

    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.

  13. Survival of human epidermal keratinocytes after short-duration high temperature: synthesis of HSP70 and IL-8.

    Science.gov (United States)

    Bowman, P D; Schuschereba, S T; Lawlor, D F; Gilligan, G R; Mata, J R; DeBaere, D R

    1997-06-01

    Thermal injury by short pulses (1-30 s) of relatively high temperature (50-68 degrees C) was investigated in normal human epidermal keratinocytes (NHEK). NHEK were cultured on plastic cover-slips and dipped in medium held at various temperatures. Survival assessed by methylthiazol tetrazolium reduction assay at 6 days postheating demonstrated an inverse time-temperature relationship that indicated that most cells could survive after a 1-s, 60 degrees C exposure or a 30-s, 55 degrees C exposure. Arrhenius plots of the data indicated major transition points for cell injury at 50 and 60 degrees C. Heat shock protein 70 (HSP70) and interleukin-8 (IL-8) were both induced by elevation of temperature between 50 and 60 degrees C for as short a time as 1 s. HSP70 synthesis stimulated by short, high pulses of heat appeared to induce thermotolerance. These results demonstrate that brief exposure to relatively high temperature can induce HSP70 and IL-8 synthesis in keratinocytes. PMID:9227428

  14. 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.

  15. Effect of L-Arginine on Kidney Morphology and HSP70 Expression in Piglets%L-精氨酸对肾脏组织形态和HSP70表达的影响

    Institute of Scientific and Technical Information of China (English)

    张翠; 阮征; 印遇龙; 邓泽元; 吴信

    2012-01-01

    Objective: To investigate the effect of L-arginine(L-Arg) on the expression of heat shock protein 70 mRNA and protein in kidney cells and its renal protective effect and mechanism.Methods: Twelve weaned piglets were randomly divided into two groups(6 piglets per group): one was control group and the other was L-Arg group,both of which were consistent with each other in terms of sex,body weight and consanguinity.A daily basal diet was fed to both groups and in addition,L-Arg was supplemented to the basal diet for the L-Arg group.After 7 d of feeding,all the piglets were killed,and plasma was collected for determination of the contents of L-Arg,ornithine(Orn) and citrulline(Cit) and kidney was also collected to determine the mRNA expression levels of HSP70 and inducible nitric oxide synthase(iNOS) by PCR and the protein expression levels of HSP70 and iNOS by Western blotting and to observe morphological changes.Results: L-Arg supplementation significantly increased plasma levels of L-Arg and Orn,mRNA expression levels of HSP70 and iNOS and HSP70 protein expression level and reduced the number of erythrocytes in kidney tubules.Conclusion: L-Arg supplementation to the diet of piglets can elevate HSP70 protein and mRNA expression and iNOS mRNA expression in the kidney and protect the kidney through maintaining the conformational stability of proteins,inhibiting the activation of apoptotic pathways of cells and alleviating kidney damage induced by weaning stress.%目的:探讨L-精氨酸(L-Arg)对肾脏应激蛋白70(heat shock protein70,HSP70)表达量的影响,以及L-Arg对肾功能的保护作用机制。方法:以仔猪为模型,按照性别、体质量以及血缘一致原则随机分成2组:对照组、L-Arg组(基础日粮+0.6%L-Arg)。仔猪断奶后饲养7d,于第8天取血浆和肾脏组织,氨基酸分析仪测定血浆中的L-Arg、鸟氨酸(Orn)和瓜氨酸(Cit)。同时,荧光实时定量PCR测定肾脏中HSP70和诱导型一

  16. Correlation of Hsp70 Serum Levels with Gross Tumor Volume and Composition of Lymphocyte Subpopulations in Patients with Squamous Cell and Adeno Non-Small Cell Lung Cancer.

    Science.gov (United States)

    Gunther, Sophie; Ostheimer, Christian; Stangl, Stefan; Specht, Hanno M; Mozes, Petra; Jesinghaus, Moritz; Vordermark, Dirk; Combs, Stephanie E; Peltz, Friedhelm; Jung, Max P; Multhoff, Gabriele

    2015-01-01

    Heat-shock protein 70 (Hsp70) is frequently found on the plasma membrane of a large number of malignant tumors including non-small cell lung cancer (NSCLC) and gets released into the blood circulation in lipid vesicles. On the one hand, a membrane (m)Hsp70-positive phenotype correlates with a high aggressiveness of the tumor; on the other hand, mHsp70 serves as a target for natural killer (NK) cells that had been pre-stimulated with Hsp70-peptide TKD plus low-dose interleukin-2 (TKD/IL-2). Following activation, NK cells show an up-regulated expression of activatory C-type lectin receptors, such as CD94/NKG2C, NKG2D, and natural cytotoxicity receptors (NCRs; NKp44, NKp46, and NKp30) and thereby gain the capacity to kill mHsp70-positive tumor cells. With respect to these results, the efficacy of ex vivo TKD/IL-2 stimulated, autologous NK cells is currently tested in a proof-of-concept phase II clinical trial in patients with squamous cell NSCLC after radiochemotherapy (RCT) at the TUM. Inclusion criteria are histological proven, non-resectable NSCLC in stage IIIA/IIIB, clinical responses to RCT and a mHsp70-positive tumor phenotype. The mHsp70 status is determined in the serum of patients using the lipHsp70 ELISA test, which enables the quantification of liposomal and free Hsp70. Squamous cell and adeno NSCLC patients had significantly higher serum Hsp70 levels than healthy controls. A significant correlation of serum Hsp70 levels with the gross tumor volume was shown for adeno and squamous cell NSCLC. However, significantly elevated ratios of activated CD69(+)/CD94(+) NK cells that are associated with low serum Hsp70 levels were observed only in patients with squamous cell lung cancer. These data might provide a first hint that squamous cell NSCLC is more immunogenic than adeno NSCLC. PMID:26579130

  17. Conserved Distal Loop Residues in the Hsp104 and ClpB Middle Domain Contact Nucleotide-binding Domain 2 and Enable Hsp70-dependent Protein Disaggregation*

    OpenAIRE

    DeSantis, Morgan E.; Sweeny, Elizabeth A.; Snead, David; Leung, Eunice H.; Go, Michelle S.; Gupta, Kushol; Wendler, Petra; Shorter, James

    2013-01-01

    The homologous hexameric AAA+ proteins, Hsp104 from yeast and ClpB from bacteria, collaborate with Hsp70 to dissolve disordered protein aggregates but employ distinct mechanisms of intersubunit collaboration. How Hsp104 and ClpB coordinate polypeptide handover with Hsp70 is not understood. Here, we define conserved distal loop residues between middle domain (MD) helix 1 and 2 that are unexpectedly critical for Hsp104 and ClpB collaboration with Hsp70. Surprisingly, the Hsp104 and ClpB MD dist...

  18. Morphological, molecular and virulence characterization of three Lencanicillium species infecting Asian citrus psyllids in Huangyan citrus groves.

    Science.gov (United States)

    Lu, Lianming; Cheng, Baoping; Du, Danchao; Hu, Xiurong; Peng, Aitian; Pu, Zhanxu; Zhang, Xiaoya; Huang, Zhendong; Chen, Guoqing

    2015-02-01

    Citrus greening or Huanglongbing (HLB) is caused by the infection of Candidatus Liberibacter spp. in citrus plants. Since Asian citrus psyllid is the primary vector of this bacterial pathogen, the spread of HLB can be mitigated by suppressing Asian citrus psyllid populations in citrus groves using entomopathogens. To expand the current data on entomopathogens infecting Asian citrus psyllids, we isolated and characterized three different entomopathogens. Strains ZJLSP07, ZJLA08, and ZJLP09 infected the Asian citrus psyllid, Diaphorina citri Kuwayama, in Huangyan citrus groves. Based on molecular and morphological analyses, two were identified as Lecanicillium attenuatum and Lecanicillium psalliotae, and the third was recognized as an unidentified species of the genus, Lecanicillium. The corrected mortalities caused by strains ZJLSP07, ZJLA08 were 100% at 7days post-inoculation, while by ZJLP09 complete mortality occurred at 6days after inoculation, with 1.0×10(8)conidia/ml at 25°C and a relative humidity of 90% in the laboratory. Under the same condition, the corrected mortalities caused by strains ZJLSP07, ZJLA08 and ZJLP09 were 100%, 92.55% and 100%, respectively at 9days post-inoculation in the greenhouse. Our findings also revealed that these fungal strains infected D. citri using hyphae that penetrated deep into the insect tissues. Further, all three strains secreted the enzymes proteinases, chitinases and lipases with a potential to destroy insect tissues. Interestingly, strain ZJLP09 had an earlier invasion time and the highest levels of enzyme activities when compared to the other two strains. These findings have expanded the existing pool of entomopathogenic fungi that infect D. citri and can be potentially used for the management of D. citri populations. PMID:25593036

  19. Morphological, molecular and virulence characterization of three Lencanicillium species infecting Asian citrus psyllids in Huangyan citrus groves.

    Science.gov (United States)

    Lu, Lianming; Cheng, Baoping; Du, Danchao; Hu, Xiurong; Peng, Aitian; Pu, Zhanxu; Zhang, Xiaoya; Huang, Zhendong; Chen, Guoqing

    2015-02-01

    Citrus greening or Huanglongbing (HLB) is caused by the infection of Candidatus Liberibacter spp. in citrus plants. Since Asian citrus psyllid is the primary vector of this bacterial pathogen, the spread of HLB can be mitigated by suppressing Asian citrus psyllid populations in citrus groves using entomopathogens. To expand the current data on entomopathogens infecting Asian citrus psyllids, we isolated and characterized three different entomopathogens. Strains ZJLSP07, ZJLA08, and ZJLP09 infected the Asian citrus psyllid, Diaphorina citri Kuwayama, in Huangyan citrus groves. Based on molecular and morphological analyses, two were identified as Lecanicillium attenuatum and Lecanicillium psalliotae, and the third was recognized as an unidentified species of the genus, Lecanicillium. The corrected mortalities caused by strains ZJLSP07, ZJLA08 were 100% at 7days post-inoculation, while by ZJLP09 complete mortality occurred at 6days after inoculation, with 1.0×10(8)conidia/ml at 25°C and a relative humidity of 90% in the laboratory. Under the same condition, the corrected mortalities caused by strains ZJLSP07, ZJLA08 and ZJLP09 were 100%, 92.55% and 100%, respectively at 9days post-inoculation in the greenhouse. Our findings also revealed that these fungal strains infected D. citri using hyphae that penetrated deep into the insect tissues. Further, all three strains secreted the enzymes proteinases, chitinases and lipases with a potential to destroy insect tissues. Interestingly, strain ZJLP09 had an earlier invasion time and the highest levels of enzyme activities when compared to the other two strains. These findings have expanded the existing pool of entomopathogenic fungi that infect D. citri and can be potentially used for the management of D. citri populations.

  20. Multifunctional selenium nanoparticles as carriers of HSP70 siRNA to induce apoptosis of HepG2 cells

    Directory of Open Access Journals (Sweden)

    Li Y

    2016-07-01

    Full Text Available Yinghua Li,1 Zhengfang Lin,1 Mingqi Zhao,1 Tiantian Xu,1 Changbing Wang,1 Huimin Xia,1,* Hanzhong Wang,2,* Bing Zhu1,* 1Guangzhou Women and Children’s Medical Center, Guangzhou, Guangdong, 2State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, People’s Republic of China *These authors contributed equally to this work Abstract: Small interfering RNA (siRNA as a new therapeutic modality holds promise for cancer treatment, but it is unable to cross cell membrane. To overcome this limitation, nanotechnology has been proposed for mediation of siRNA transfection. Selenium (Se is a vital dietary trace element for mammalian life and plays an essential role in the growth and functioning of humans. As a novel Se species, Se nanoparticles have attracted more and more attention for their higher anticancer efficacy. In the present study, siRNAs with polyethylenimine (PEI-modified Se nanoparticles (Se@PEI@siRNA have been demonstrated to enhance the apoptosis of HepG2 cells. Heat shock protein (HSP-70 is overexpressed in many types of human cancer and plays a significant role in several biological processes including the regulation of apoptosis. The objective of this study was to silence inducible HSP70 and promote the apoptosis of Se-induced HepG2 cells. Se@PEI@siRNA were successfully prepared and characterized by various microscopic methods. Se@PEI@siRNA showed satisfactory size distribution, high stability, and selectivity between cancer and normal cells. The cytotoxicity of Se@PEI@siRNA was lower for normal cells than tumor cells, indicating that these compounds may have fewer side effects. The gene-silencing efficiency of Se@PEI@siRNA was significantly much higher than Lipofectamine 2000@siRNA and resulted in a significantly reduced HSP70 mRNA and protein expression in cancer cells. When the expression of HSP70 was diminished, the function of cell protection was also removed and cancer cells became more

  1. 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...

  2. 内源性大麻素2-AG促进HSP70基因的热应激表达%Endocannabinoid 2-AG enhances HSP70 gene expression in heat stress response in vitro

    Institute of Scientific and Technical Information of China (English)

    吴娟; 屈明玥; 张伟; 王登高; 王颖莹

    2009-01-01

    Objectives To investigate the regulation role of endocannabinoid in heat stress response and its possible receptor signaling pathway. Methods Rat glioma C6 cells were pretreated with different concentra-tions (10-7, 10-6 and 10-5mol/L) of endocannabinoid 2-Arachidonylglycerol (2-AG) for 1 h, then heat stress (42 ℃ water bath) was performed for 1 h. Reverse transcription-PCR and Western blotting were used to detect the expression levels of HSP70 mRNA and protein. The blockade effects of selective CB1 antagonist AM251 (10-4mol/L) or CB2 antagonist AM630 (10-4mol/L) were also observed by adding them to the culture medi-um when 2-AG was pretreated. Results The expression levels of HSP70 mRNA and protein in heat stressed C6 cells were significantly increased after 2-AG treatment. This enhancement was mainly through CB1 receptor signaling pathway. Conclusion Certain level of 2-AG significantly promotes the expression of HSP70 gene post heat stress, suggesting that 2-AG is involved in the regulation of heat stress response. The promotion effect of 2-AG in C6 cells is mainly mediated through CB1 receptor signaling pathway, but not CB2.%目的 探讨内源性大麻素对热应激反应是否存在调控作用,及其可能的受体调控机制.方法 以不同浓度(10-7、10-6、10-5mol/L)内源性大麻素2-AG(2-Arachidonylglycerol)预处理大鼠胶质瘤C6细胞1 h,热刺激(42℃水浴)1 h,通过RT-PCR及Western blot检测热休克蛋白70(heat shock protein 70,HSP70)的mRNA及蛋白表达水平;同时观察特异性受体拮抗剂AM251或AM630的拮抗效应,探讨信号通路机制.结果 内源性大麻素2-AG可显著促进热刺激处理的C6细胞HSP70基因及蛋白的表达,该调控效应由CB1介导.结论 一定浓度的内源性大麻素2-AG对热刺激处理的C6细胞内HSP70基因的表达有明显促进效应,表明2-AG对细胞热应激反应可能具有一定的调控能力;同时,在C6细胞内,该调控效应主要由CB1介导.

  3. The expressions of HSP70 and αB-crystallin in myocarditis associated with foot-and-mouth disease virus in lambs

    OpenAIRE

    Gulbahar, Mustafa Yavuz; Kabak, Yonca Betil; Karayigit, Mehmet Onder; Yarim, Murat; Guvenc, Tolga; Parlak, Unal

    2011-01-01

    This study describes the expression of heat shock protein70 (HSP70) and alpha-basic-crystallin (α-BC) and their association with apoptosis and some related adaptor proteins in the pathogenesis of foot-and-mouth disease virus (FMDV)-induced myocarditis in lambs. HSP70 was generally overexpressed in the myocardial tissues and inflammatory cells of FMDV-induced myocarditis with differential accumulation and localization in same hearts when compared to non-foot-and-mouth disease control hearts. α...

  4. 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.

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

    Science.gov (United States)

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

    2009-10-12

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

  6. Evaluation of immune effect of recombinant fusion protein targeting the prostate stem cell antigen based on PSCA and HSP70

    Directory of Open Access Journals (Sweden)

    Lei DONG

    2014-10-01

    Full Text Available Objective To explore the immune effect and antitumor activity of recombinant prostate stem cell protein (PSCA and heat shock protein 70 (HSP70 in a murine model of prostate cancer. Methods Twenty-five healthy male C57BL/6 mice were randomly divided into 5 groups (5 each: PSCA, HSP, PSCA+HSP, PSCA-HSP and control group. Mice in the first 4 groups were vaccinated with the corresponding proteins, and those in control group were faked with injection of phosphate buffer saline (PBS. After immunization with recombinant proteins, the PSCA-specific cellular immune responses were monitored with ELISPOT, intracellular cytokine staining assay, and flow cytometry, and ELISA assay was used to detect humoral immune responses. The tumor growth and survival of vaccined mice were observed. Results ELISPOT revealed that the mice in PSCA-HSP group generated much more IFN-γ spot-forming cells than those in other groups (P<0.05, and they could generate strong anti-PSCA antibody response. Results of flow cytometry showed that the number of CD8+/IFN-γ+ T cells was significantly higher in PSCAHSP group than that in other groups (P<0.05. ELISA results revealed that all the mice in PSCA, PSCA+HSP and PSCA-HSP group were induced to generate the PSCA-specific humoral immune response, and no statistical difference was found on the antibody levels among the three groups. Animal experiment showed that PSCA-HSP could inhibit the growth of PSCA-expressing tumors and prolong the survival time of vaccinated mice. Conclusion HSP70 is a chaperone with significant effect for protein vaccines, and the recombinant fusion protein PSCA-HSP70 could be of potential value for prostate cancer treatment. DOI: 10.11855/j.issn.0577-7402.2014.09.08

  7. 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

  8. Expression of Hsp27 and Hsp70 in Lymphocytes and Plasma in Healthy Workers and Coal Miners with Lung Cancer

    Institute of Scientific and Technical Information of China (English)

    王海椒; 邢景才; 王峰; 韩文慧; 任侯卯; 邬堂春; 陈卫红

    2010-01-01

    In coal mines, main occupational hazard is coal-mine dust, which can cause health problem including coal workers' pneumoconiosis and lung cancer. Some heat shock proteins (Hsps) have been reported as an acute response to a wide variety of stressful stimuli. Whether Hsps protect against chronic environmental coal-mine dust over years is unknown. It is also interesting to know that whether the expression of Hsp27 and Hsp70 proteins as a marker for exposure is associated risk of lung cancer among coal miners. ...

  9. Heat Stress Induces Extended Plateau of Hsp70 Accumulation--A Possible Cytoprotection Mechanism in Hepatic Cells.

    Science.gov (United States)

    Miova, Biljana; Dinevska-Kjovkarovska, Suzana; Esplugues, Juan V; Apostolova, Nadezda

    2015-10-01

    The relevance of heat preconditioning resides in its ability to protect cells from different kinds of injury by induction of heat shock proteins, a process in which the intensity of heat stress (HS) and duration of subsequent recovery are vital. This study evaluates the effects of moderate HS (45 min/43°C) and the time-dependent changes during recovery period of HSP70, Bcl-2 and p53 gene and protein expression in HepG2 cells. We also evaluated the effects of 0.4 mM aspirin (ASA) as a potential pharmacological co-inducer of HSP, both alone and in a combination with HS (ASA + HS). HS alone and ASA + HS caused a major up-regulation of HSP70 mRNA in the first 2 h, while HSP70 protein increased gradually and was especially abundant from 2 h to 24 h. Regarding Bcl-2, all treatments rendered similar results: gene expression was down-regulated in the first 2 h, after which there was protein elevation (12-48 h after HS). mRNA expression of p53 in HS- and (ASA + HS)-cells was down-regulated in the first 12 h. The immediate decrease of p53 protein after HS was followed by a biphasic increase. In conclusion, 0.4 mM ASA + HS does not act as a co-inducer of HSP70 in HepG2 cells, but promotes Bcl-2 protein expression during prolonged treatment. Our suggestion is that hepatic cells are most vulnerable in the first 2-6 h, but may have a high capacity for combating stress 12-24 h after HS. Finally, short-term exposure HS might be a "physiological conditioner" for liver cells to accumulate HSP and Bcl-2 proteins and thus obtain cytoprotection against an additional stress.

  10. Comparative evaluation of recombinant HSP70 (N & C-terminal) fragments in the detection of equine trypanosomosis.

    Science.gov (United States)

    Kumar, Jaideep; Chaudhury, A; Yadav, S C

    2016-06-15

    Trypanosomosis (Surra) is an economically important disease caused by Trypanosoma evansi which is an extracellular parasite present in the plasma, tissues and other body fluids of a wide range of hosts including domesticated animals. Currently, serological reports are based on detection of antibodies by ELISA using whole cell lysate (WCL) antigen, which has a limitation of persistence of anti-trypanosomal antibodies after successful treatment of the disease. Moreover, it has some ethical issues also like requirement of mice for in vivo maintenance of parasite for preparing the antigen. Therefore, in the present study, an attempt was made to evaluate the in vitro production of recombinant heat shock protein 70 (HSP70) for detection of antibodies in experimentally infected ponies. The amino acid sequence analysis of HSP70 revealed that N-terminal region of the protein was highly conserved while the C-terminal region was most divergent. The four different regions of HSP70 protein viz. HSP-1, HSP-2, HSP-3 and HSP-4 were cloned and expressed, among which HSP-1 (N-terminal region) & HSP-2 (C-terminal region) were truncated while HSP-3 & HSP-4 were complete C-terminal proteins. The recombinant fragments were probed with sequentially pooled experimental serum samples where antibodies were detected in these fragments from 10(th) day post infection till the termination of the experiment. Further, these recombinant fragments were also comparatively evaluated with WCL antigen in ELISA using experimental as well as field serum samples. It was observed that after successful treatment of infected ponies, there was a sharp fall in antibodies (within 90 days) when tested with recombinant HSP's fragments, while antibodies persisted even after 469 days when tested against WCL antigen. The sensitivity and specificity of all HSP70 fragments were also estimated from field serum samples with reference to WCL antigen ELISA. The HSP-1 showed minimum sensitivity (41.03%) among all the

  11. Modulation of Akt and ERK1/2 pathways by resveratrol in chronic myelogenous leukemia (CML cells results in the downregulation of Hsp70.

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    Soumyajit Banerjee Mustafi

    Full Text Available BACKGROUND: Resveratrol is known to downregulate the high endogenous level of Heat shock protein 70 (Hsp70 in Chronic Myelogenous Leukemia (CML K562 cells and induce apoptosis. Since Heat Shock Factor 1 (HSF1 controls transcription of Hsp70, we wanted to probe the signaling pathways responsible for transcriptional activation of HSF1. METHODOLOGY/PRINCIPAL FINDINGS: Cells exposed to 40microM Resveratrol rapidly abolished serine473 phosphorylation of Akt and significantly reduced its kinase activity. Inactivation of Akt pathway by Resveratrol subsequently blocked serine9 phosphorylation of Gsk3beta. Active non-phosphorylated Gsk3beta rendered HSF1 transcriptionally inactive and reduced Hsp70 production. Blocking PI3K/Akt activity also demonstrated similar effects on Hsp70 comparable to Resveratrol. Inactivation of Gsk3beta activity by inhibitors SB261763 or LiCl upregulated Hsp70. Resveratrol significantly modulated ERK1/2 activity as evident from hyper phosphorylation at T302/Y304 residues and simultaneous upregulation in kinase activity. Blocking ERK1/2 activation resulted in induction of Hsp70. Therefore, increase in ERK1/2 activity by Resveratrol provided another negative influence on Hsp70 levels through negative regulation of HSF1 activity. 17-allylamino-17-demethoxygeldanamycin (17AAG, a drug that inhibits Hsp90 chaperone and degrades its client protein Akt concomitantly elevated Hsp70 levels by promoting nuclear translocation of HSF1 from the cytosol. This effect is predominantly due to inhibition of both Akt and ERK1/2 activation by 17AAG. Simultaneously treating K562 with Resveratrol and 17AAG maintained phosho-ERK1/2 levels close to untreated controls demonstrating their opposite effects on ERK1/2 pathway. Resveratrol was found not to interfere with Bcr-Abl activation in K562 cells. CONCLUSION/SIGNIFICANCE: Thus our study comprehensively illustrates that Resveratrol acts downstream of Bcr-Abl and inhibits Akt activity but stimulates ERK

  12. In vitro Study on Role of Hsp70 Expression in DNA Damage of Human Embryonic Lung Cells Exposed to Benzo[a]pyrene

    Institute of Scientific and Technical Information of China (English)

    YA-JUAN GAO; CHENG-FENG XIAO; SHENG CHEN; RUI-BO WANG; HAN-ZHEN HE; ROBERT M TANGUAY; TANG-CHUN WU

    2004-01-01

    Objective Benzo[a]pyrene (B[a]P), a ubiquitous environmental pollutant, is a potent procarcinogen and mutagen that can elicit tumors, leading to malignancy. Heat shock proteins (Hsp) have been shown to protect cells against damages caused by various stresses including exposure to numerous chemicals. Whether Hsps, or more specifically Hsp70, are involved in repair of B[a]P-induced DNA damage is currently unknown. Methods We assessed the potential role of the inducible form of Hsp70 in B[a]P-induced DNA damage of human embryonic lung (HEL) cells using immunoblot and the comet assay (i.e., the single cell gel electrophoresis assay). Results Exposure to B[a]P induced a dose-dependent decrease in the level of Hsp70, but a dose-dependent increase in DNA damage both in untreated (control) HEL cells and in cells preconditioned by a heat treatment. Heat preconditioning prior to B[a]P exposure potentiated the effect of B[a]P at a low dose (10 (mol/L), but appeared to be protective at higher doses. There was a negative correlation between Hsp70 level and DNA damage in the non-preheated as well as in the preconditioned cells. Conclusion These data suggest that exposure of HEL cells to B[a]P may induce a dose-dependent reduction in the levels of the inducible Hsp70. The detailed mechanisms for the reduction of Hsp70 levels by B[a]P and the role of Hsp70 in DNA damage under different concentrations of B[a]P remains to be determined.

  13. Mild heat stress at a young age in Drosophila melanogaster leads to increased Hsp70 synthesis after stress exposure later in life

    Indian Academy of Sciences (India)

    Torsten Nygaard Kristensen; Jesper Givskov Sørensen; Volker Loeschcke

    2003-12-01

    In a number of animal species it has been shown that exposure to low levels of stress at a young age has a positive effect on stress resistance later in life, and on longevity. The positive effects have been attributed to the activation of defence/cleaning systems (heat shock proteins (Hsps), antioxidases, DNA repair) or to effects of a changed metabolic rate, or both. We investigated the effect of mild stress exposures early in life on Hsp70 synthesis after a harder stress exposure later in life in five isofemale lines of Drosophila melanogaster. Female flies were either exposed to repeated bouts of mild heat stress (3 h at 34°C) at a young age (days 2, 4 and 6 post-eclosion) or held under standard laboratory conditions. At 16 and 32 days of adult age, respectively, flies were exposed to a high temperature treatment known to induce Hsp70 in the investigated species (1 h at 37°ºC). Thereafter, the inducible Hsp70 levels were measured. Our data show a tendency towards increased Hsp70 synthesis with increased age for both ‘mild stress’ and ‘no stress’ flies. Moreover, the results show that flies exposed to mild stress at a young age synthesized more Hsp70 upon induction, compared to control flies, and that this difference was accentuated at 32 days compared to 16 days of age. Thus, bouts of mild heat stress at a young age impact on the physiological stress response system later in life. This may be caused by an increased ability to react to future stresses. Alternatively, the mild stress exposure at a young age may actually have caused cellular damages increasing the need for Hsp70 levels after stress exposure later in life. The importance of an Hsp70 upregulation (throughout life) in explaining the phenomenon of hormesis is discussed, together with alternative hypotheses, and suggestions for further studies.

  14. Functional evaluation of Heat Shock Proteins 70 (HSP70/HSC70) on Rhodnius prolixus (Hemiptera, Reduviidae) physiological responses associated with feeding and starvation.

    Science.gov (United States)

    Paim, Rafaela M M; Araujo, Ricardo N; Leis, Miguel; Sant'anna, Mauricio R V; Gontijo, Nelder F; Lazzari, Claudio R; Pereira, Marcos H

    2016-10-01

    Blood-sucking vectors must overcome thermal stress caused by intake of proportionally large amounts of warm blood from their hosts. In response to this, Heat Shock Proteins (HSPs) such as the widely studied HSP70 family (the inducible HSP70 and the cognate form HSC70, known for their role in preserving essential cellular functions) are rapidly up-regulated in their tissues. The triatomine Rhodnius prolixus is an important vector of Trypanosoma cruzi, the causative pathogen of Chagas' disease, and is also a model organism for studying insect biology and physiology. In this work, we observed that the expression of Rhodnius prolixus HSP70 was rapidly up-regulated in response to thermal shocks (0 °C and 40 °C) and also during the first hours after feeding on blood. HSP70/HSC70 RNAi knockdown elicited important alterations in R. prolixus physiological responses triggered by blood meal and starvation. HSP70/HSC70 knockdown insects showed lower resistance to prolonged starvation in comparison to appropriate controls, dying between 32 and 40 days after dsRNA injection. After blood feeding, the physiological effects of HSP70/HSC70 knockdown were more prominent and the insects died even earlier, within 14-20 days after feeding (21-27 days after dsRNA injection). These bugs showed impaired blood processing and digestion, reduced energetic metabolism and the midgut immune responses were compromised. Our findings suggest that HSP70/HSC70 depletion affected R. prolixus in starvation or fed conditions. After feeding, the arrival of blood in the digestive tract of knockdown insects fails to activate essential signaling pathways involved in blood processing, producing several alterations in their physiological processes enough to generate a premature death. PMID:27491440

  15. 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.

  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. 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.

  18. Protective Effects of Focal Ischemic Preconditioning and HSP70 Expression on Middle Cerebral Artery Occlusion in Rats

    Institute of Scientific and Technical Information of China (English)

    ZHAO Jianhua; SUN Senggang; CHEN Xiaowu

    2006-01-01

    To systematically evaluate the importance of protein synthesis in ischemic preconditioning (PC)-induced ischemic tolerance (IT), temporary middle cerebral artery occlusion (MCAO)by Longa (20 min) was used for PC (ischemic precondioning). Twenty-four hours of reperfusion was allowed after PC and before permanent MCAO to establish ischemic tolerance (IT) to compare with non-PC (sham-operated) rats (n=5 for each group). Infarct size and neurological deficits were measured 24 h after PMCAO. Samples of brain were taken for the determination of HSP70 expression by Western blot analysis. The effects of the protein synthesis inhibitor cycloheximide administered just before PC or administered long after PC but just before PMCAO on IT were also determined (n=5 for each group). Our results showed that hemispheric infarct was significantly reduced (P<0.01) only if PC was performed after 24 h, and PC significantly (P<0.05) reduced neurological deficits (similar to reductions in infarct size). Cycloheximide eliminated ischemic PC-induced IT effects on both brain injury and neurological deficits if administered before PC but not if administered long after PC but before PMCAO. PC produced no brain injury but did increase HSP70 protein 24 h after PC. Cycloheximide eliminated that effect. The results suggest that PC is a powerful inducer of ischemic brain tolerance as reflected by the preservation of brain tissue and motor function. PC induces IT that is dependent on de novo protein synthesis.

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

    Directory of Open Access Journals (Sweden)

    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.

  20. [Hsp70 Fused with the Envelope Glycoprotein E0 of Classical Swine Fever Virus Enhances Immune Responses in Balb/c Mice].

    Science.gov (United States)

    Xu, Qianqian; Zhang, Xiaomin; Jing, Jiao; Shi, Baojun; Wang, Shiqi; Zhou, Bin; Chen, Puyan

    2015-07-01

    Heat-shock protein (Hsp) 70 potentiates specific immune responses to some antigenic peptides fused to it. Here, the prokaryotic plasmids harboring the envelope glycoprotein E0 gene of classical swine fever virus (CSFV) and/or the Hsp70 gene of Haemophilus parasuis were constructed and expressed in Escherichia coli Rosseta 2(R2). The fusion proteins were then purified. Groups of Balb/c mice were immunized with these fusion proteins, respectively, and sera collected 7 days after the third immunization. Immune effects were determined via an enzyme-linked immunosorbent assay and flow cytometric analyses. E0-Hsp70 fusion protein and E0+Hsp70 mixture significantly improved the titer of E-specific antibody, levels of CD4+ and CD8+ T cells, and release of interferon-γ. These findings suggested that Hsp70 can significantly enhance the immune effects of the envelope glycoprotein E0 of CSFV, thereby laying the foundation of further application in pigs. PMID:26524908

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

    Directory of Open Access Journals (Sweden)

    Masako Nakamura

    2012-02-01

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

  2. Effect of Calmodulin (CaM) on Expression of Inducible Heat Shock Protein (HSP70) in Mice(Mus musculus) Preimplantation Embryo%钙调蛋白(CaM)对附植前小鼠胚胎热休克蛋白70(HSP70)表达的影响

    Institute of Scientific and Technical Information of China (English)

    索佳佳; 张保珍; 孙春玲; 王占美; 张双; 曹荣峰; 高善颂; 田文儒

    2013-01-01

    Heat shock protein(HSP)70 is synthesized in the preimplantation embryos of cow(Bos tarurs),ewe (Ovis aries) and mice(Mus musculus) under the conditions of heat shock in vitro,which enhances their thermotolerance to protect them from further heat damage.And Calmodulin(CaM)is the one of most important multifunctional receptor of Ca2 + and plays a key role in regulating the important gene expression in cell activities.To clarity whether the CaM participating in the expression of HSP70 in the mice embryos with heat shock and verify its mechanism,mice embryos were cultured in vitro to the blastocyst stage and then were randomly divided into control group(37℃),group treated with W7 without heat shock(37℃ +W7),heat shock group(39℃)and heat shock group treated with W7.The method of RT-PCR was used to detect the gene expression of CaM,HSP70 and heat shock factorl(HSF1),and the method of Western blot was used to detect the expression of CaM,HSP70 and HSF1.The combinations of both HSP70-CaM and HSP70-HSF1 were detected by using co-immunoprecipitation(Co-IP).The results showed that the CaM and HSP70 mRNA expression significantly increased in the mice embryos treated with 39℃ for 1 h (P<0.05).However,the CaM and HSP70 mRNA expression significantly decreased in both groups of embryo cultured at 37℃ (P<0.05) and 39℃ (P<0.01) treated with W7; W7 had no effect on the HSF1 mRNA expression in any groups of embryo.The synthesis of CaM,HSP70 and HSF1 significantly increased in the embryos treated at 39℃ for 1 h (P<0.05),and the CaM synthesis significantly decreased in the embryos cultured both at 37℃ and 39℃ and treated with W7 (P<0.05).W7 had no effect on neither HSP70 nor HSF1 synthesis in the embryos cultured at 37℃ (P>0.05) while it inhibited both HSP70 and HSF1 synthesis in the embryos cultured at 39℃ (P<0.05).The study also found that CaM combined with HSP70 and formed CaM-HSP70 complex in the mice embryos.The identification of interactions

  3. 运动对心肌、骨骼肌HSP70表达影响的研究进展%Progress of the Research on the Effect of Exercises on HSP70 Expression in Cardiac and Skeletal Muscles

    Institute of Scientific and Technical Information of China (English)

    曹桂霞; 白俊伟

    2010-01-01

    热休克蛋白的分子伴侣、抗氧化、抗损伤、协同免疫及抗细胞凋亡等多种功能,不仅生物界对它越来越关注,而且运动医学界的学者也对它产生了浓厚的兴趣.HSP70作为细胞应激蛋白,对不同强度与形式的运动,其反应和适应有很大差异,这为我们进一步认识运动规律,从而进行科学的体育锻炼和运动训练提供了新的思路和方法.

  4. 香蕉果皮2个HSP70基因片段的克隆及其在热激和冷藏过程中的表达%Cloning and expression analysis of two fragments of HSP70 cDNA in the peel of banana during storage at high and low temperature

    Institute of Scientific and Technical Information of China (English)

    贺立红; 陈建业; 王海蓝; 邝健飞; 陆旺金

    2012-01-01

    温度逆境胁迫可以诱导热激蛋白的表达,而热激处理能够减轻果实冷害。为了探讨温度逆境对香蕉热激蛋白基因表达的调控、辨析香蕉热激蛋白基因的表达与果实冷害的关系,采用同源序列法分离了香蕉果皮HStVO基因序列一根据已经报道的HStVO基因的氨基酸保守序列设计引物、以香蕉果皮总RNA为模板,用RT—PCR方法克隆香蕉的HSP70cDNA,Northern杂交分析该基因在不同贮藏温度下的表达特征。结果得到2个序列不同的HStVO基因片段,分别命名为Ma—HSP70-1和Ma—HSP70—2,Ma—HSP70-1和肘n—HSP70—2之间的碱基同源性为86.9%,氨基酸同源性为97.1%。Northern杂交的结果表明,38℃短期热激处理诱导了Ma-HSP70—2基因的表达,低温冷藏能使2个基因的表达增强。并初步认为Ma-HSP70可能与香蕉果实耐冷性有关。%Many evidences have indicated that temperature stress can induce heat shock protein expression and heat treatment can alleviate chilling injury of fruits. In the present study, two HSP70 fragments were cloned from banana peel using homology-based method in order to investigate their expressions regulated by temperature stress and the relationship between their expressions and chilling injury. Using PCR de- generate primers designed with reference to the conserved amino acid sequences of known HSP70, total RNA extracted from banana peel was used as templates and HSPTO eDNA fragments were amplified by RT-PCR in order to study their expression characteristics under different environments by Northern hy- bridization. Two different eDNA fragments, named Ma-HSP70-1 and Ma-HSP70-2, were cloned. The homology between two HSP70 is 86.9% at nueleotide acid sequence level and 97.1% at amino acid se- quence level. Using Ma-HSP70-1 and Ma-HSP70-2 as probes, Northern hybridization showed that short-time heat pretreatment (38 ℃ )only induced the expression of Ma-HSP70

  5. Acetylcholinesterase (AChE) and heat shock proteins (Hsp70) of gypsy moth (Lymantria dispar L.) larvae in response to long-term fluoranthene exposure.

    Science.gov (United States)

    Mrdaković, Marija; Ilijin, Larisa; Vlahović, Milena; Matić, Dragana; Gavrilović, Anja; Mrkonja, Aleksandra; Perić-Mataruga, Vesna

    2016-09-01

    Polycyclic aromatic hydrocarbons (PAHs) may affect biochemical and physiological processes in living organisms, thus impairing fitness related traits and influencing their populations. This imposes the need for providing early-warning signals of pollution. Our study aimed to examine changes in the activity of acetylcholinesterase (AChE) and the concentration of heat shock proteins (Hsp70) in homogenates of brain tissues of fifth instar gypsy moth (Lymantria dispar L.) larvae, exposed to the ubiquitous PAH, fluoranthene, supplemented to the rearing diet. Significantly increased activity of AChE in larvae fed on the diets with high fluoranthene concentrations suggests the necessity for elucidation of the role of AChE in these insects when exposed to PAH pollution. Significant induction of Hsp70 in gypsy moth larvae reared on the diets containing low fluoranthene concentrations, indicate that changes in the level of Hsp70 might be useful as an indicator of pollution in this widespread forest species. PMID:27343862

  6. Stress-induced phosphorylation of c-Jun-N-terminal kinases and nuclear translocation of Hsp70 in the Wistar rat hippocampus

    Directory of Open Access Journals (Sweden)

    Adžić M.

    2009-01-01

    Full Text Available Glucocorticoids are key regulators of the neuroendocrine stress response in the hippocampus. Their action is partly mediated through the subfamily of MAPKs termed c-Jun-N-terminal kinases (JNKs,whose activation correlates with neurodegeneration. The stress response also involves activation of cell protective mechanisms through various heat shock proteins (HSPs that mediate neuroprotection. We followed both JNKs and Hsp70 signals in the cytoplasmic and nuclear compartments of the hippocampus of Wistar male rats exposed to acute, chronic, and combined stress. The activity of JNK1 was decreased in both compartments by all three types of stress, while the activity of cytoplasmic JNK2/3 was elevated in acute and unaltered or lowered in chronic and combined stress. Under all stress conditions, Hsp70 translocation to the nucleus was markedly increased. The results suggest that neurodegenerative signaling of JNKs may be counteracted by increase of nuclear Hsp70,especially under chronic stress.

  7. Coagulansin-A has beneficial effects on the development of bovine embryos in vitro via HSP70 induction

    Science.gov (United States)

    Khan, Imran; Lee, Kyeong-Lim; Fakruzzaman, Md.; Song, Seok-Hwan; Ihsan-ul-Haq; Mirza, Bushra; Yan, Chang Guo; Kong, Il-Keun

    2016-01-01

    Coagulansin-A (withanolide) is the steroidal lactone obtained from Withania coagulans which belong to Solanaceae family. The present study investigated the effects of coagulansin-A on bovine oocyte maturation and embryo development in vitro. All these oocytes were aspirated from the ovaries obtained from Korean Hanwoo cows at a local abattoir. To determine whether coagulansin-A has beneficial effects on bovine oocyte maturation in vitro, 355 oocytes per group (control and treated) in seven replicates were subjected with different concentrations (1, 2.5, 5, 7.5 and 10 μM) of coagulansin-A. The coagulansin-A was added in the in vitro maturation (IVM) media followed by in vitro fertilization (IVF) and then in vitro culture (IVC). Only treatment with 5 μM coagulansin-A remarkably (P<0.05) improved embryos development (Day 8 blastocyst) having 27.30 and 40.01% for control and coagulansin-A treated groups respectively. Treatment with 5 μM coagulansin-A significantly induced activation of heat shock protein 70 (HSP70) (P<0.05). Immunofluorescence analysis revealed that 5 μM coagulansin-A treatment also significantly inhibited oxidative stress and inflammation during bovine embryo development in vitro by decreasing 8-oxoguanosine (8-OxoG) (P<0.05) and nuclear factor-κB (NF-κB) (P<0.05). The expressions of HSP70 and NF-κB were also conformed through real-time PCR (RT-PCR). Additionally, the terminal deoxynucleotidyl transferase dUTP nick-end labelling (TUNEL) assay confirmed that coagulansin-A treatment significantly improved the embryo quality and reduced bovine embryo DNA damage (P<0.05). The present study provides new information regarding the mechanisms by which coagulansin-A promotes bovine embryo development in vitro. PMID:26831738

  8. Novel Improved Synthesis of HSP70 Inhibitor, Pifithrin-μ. In Vitro Synergy Quantification of Pifithrin-μ Combined with Pt Drugs in Prostate and Colorectal Cancer Cells.

    Science.gov (United States)

    McKeon, Aoife M; Egan, Alan; Chandanshive, Jay; McMahon, Helena; Griffith, Darren M

    2016-01-01

    We describe a novel improved approach to the synthesis of the important and well-known heat shock protein 70 inhibitor (HSP70), pifithrin-μ, with corresponding and previously unreported characterisation. The first example of a combination study comprising HSP70 inhibitor pifithrin-μ and cisplatin or oxaliplatin is reported. We have determined, using the Chou-Talalay method, (i) moderate synergistic and synergistic effects in co-treating PC-3 prostate cancer cells with pifithrin-μ and cisplatin and (ii) significant synergistic effects including strong synergism in cotreating HT29 colorectal cancer cells with oxaliplatin and pifithrin-μ. PMID:27455212

  9. Novel Improved Synthesis of HSP70 Inhibitor, Pifithrin-μ. In Vitro Synergy Quantification of Pifithrin-μ Combined with Pt Drugs in Prostate and Colorectal Cancer Cells

    Directory of Open Access Journals (Sweden)

    Aoife M. McKeon

    2016-07-01

    Full Text Available We describe a novel improved approach to the synthesis of the important and well-known heat shock protein 70 inhibitor (HSP70, pifithrin-μ, with corresponding and previously unreported characterisation. The first example of a combination study comprising HSP70 inhibitor pifithrin-μ and cisplatin or oxaliplatin is reported. We have determined, using the Chou-Talalay method, (i moderate synergistic and synergistic effects in co-treating PC-3 prostate cancer cells with pifithrin-μ and cisplatin and (ii significant synergistic effects including strong synergism in cotreating HT29 colorectal cancer cells with oxaliplatin and pifithrin-μ.

  10. 2-phenylethynesulfonamide Prevents Induction of Pro-inflammatory Factors and Attenuates LPS-induced Liver Injury by Targeting NHE1-Hsp70 Complex in Mice.

    Directory of Open Access Journals (Sweden)

    Chao Huang

    Full Text Available The endotoxin-mediated production of pro-inflammatory cytokines plays an important role in the pathogenesis of liver disorders. Heat shock protein (Hsp70 overexpression has established functions in lipopolysaccharide (LPS-mediated inflammatory response. However, little is known about the role of Hsp70 activity in LPS signaling. We hypothesized that inhibition of Hsp70 substrate binding activity can ameliorate LPS-induced liver injury by decreasing induction of pro-inflammatory factors. In this study, C57/BL6 mice were injected intraperitoneally with LPS and 2-phenylethynesulfonamide (PES, an inhibitor of Hsp70 substrate binding activity. We found that i. PES prevented LPS-induced increase in serum alanine aminotransferase (ALT and aspartate aminotransferase (AST activity, infiltration of inflammatory cells, and liver cell apoptosis; ii. PES reduced inducible nitric oxide synthase (iNOS protein expression as well as serum nitric oxide (NO, tumor necrosis factor-α (TNF-α, and interleukin-6 (IL-6 content in LPS-stimulated mice; iii. PES reduced the mRNA level of iNOS, TNF-α, and IL-6 in LPS-stimulated liver. iiii. PES attenuated the degradation of inhibitor of κB-α (IκB-α as well as the phosphorylation and nuclear translocation of nuclear factor-κB (NF-κB in LPS-stimulated liver. Similar changes in the protein expression of inflammatory markers, IκB-α degradation, and NF-κB phosphorylation and nuclear translocation were observed in RAW 264.7 cells. Further mechanistic studies revealed that PES remarkably reduced the elevation of [Ca(2+]i and intracellular pH value (pHi in LPS-stimulated RAW 264.7 cells. Furthermore, PES significantly reduced the increase in Na(+/H(+ exchanger 1 (NHE1 association to Hsp70 in LPS-stimulated macrophages and liver, suggesting that NHE1-Hsp70 interaction is required for the involvement of NHE1 in the inflammation response. In conclusion, inhibition of Hsp70 substrate binding activity in vivo reduces the

  11. 旋毛虫重组HSP70对小鼠巨噬细胞TLR2/4表达的影响%Effect of 70 ku recombinant HSP70 from Trichinella spiralis on expres-sion of TLR2/4 in murine macrophage

    Institute of Scientific and Technical Information of China (English)

    李成; 禹洋; 徐佳; 刘畅; 周兴东; 李晓云

    2014-01-01

    观察旋毛虫重组热休克蛋白HSP70(rTs-HSP70)对体外培养小鼠巨噬细胞Toll样受体TLR2/4表达的影响。体外常规培养小鼠巨噬细胞系RAW264.7,分别加入不同浓度rTs-HSP70刺激培养24 h,半定量PCR方法检测培养细胞TLR2/4 mRNA表达,流式细胞仪检测表面标志分子CD80和CD86表达。平行设立不加刺激物阴性对照组,细菌脂多糖LPS阳性对照组;同时试验比较5μg·mL-1浓度的HSP70组和HSP70+LPS组(HSP70预刺激后再加入LPS刺激培养12 h)RAW264.7细胞TLR2/4 mRNA表达差异。结果表明,低浓度rTs-HSP70与巨噬细胞活化呈正相关,浓度为5μg·mL-1时,TLR2/4表达水平达到峰值(P5μg·mL-1)则对巨噬细胞RAW264.7表现为抑制作用;当对RAW264.7进行旋毛虫HSP70预刺激,可引起免疫抑制效应,抑制LPS对巨噬细胞TLR2/4的活化反应。试验可为旋毛虫相关分子免疫调节机制研究及旋毛虫病防治提供参考。%To observe the effect of 70 kDa recombinant heat shock protein from Trichinella spiralis (rTs-HSP70) on expression of TLR2/4 (toll-like receptors) in murine macrophage in vitro. Murine macrophage RAW264.7 cell lines were incubated in twelve-well plates and different concentrations of rTs-HSP70 were added, co-culturing for 24 hours. Then the cells’TLR2/4 expression were tested by semi-quantitative PCR, those of surface markers CD80 and CD86 by flow cytometry. Cells receiving no stimulation were taken as negative control, bacterial lipopolysaccharide (LPS) as positive control. At the same time, comparative studies on incubation with rTs-HSP70 alone, rTs-HSP70 in combination with LPS, were conducted at concentration 5 µg·mL-1 of HSP70. The results indicated that lower concentra-tions of rTs-HSP70 had a positive correlation with macrophages activation, TLR2/4 expression peaked at concentration of 5 µg·mL-1 (P5 µg·mL-1), a inhibitory effect on macrophage could be observed. Pre-stimulation of the

  12. Expression of the C-terminal Polypeptide of Planarian Heat Shock Protein 70 and Preparation of Its Polyclonal Antibody%日本三角涡虫热休克蛋白70(DjHSP70)C-端多肽表达及其抗血清制备

    Institute of Scientific and Technical Information of China (English)

    马克学; 娄昊; 陈广文; 刘德增

    2011-01-01

    首先运用在线生物学软件对日本三角涡虫(Degusia japonica)热休克蛋白70(DjHSP70)氨基酸序列进行亲水区分析,发现该蛋白C-端含有较多亲水性氨基酸,然后以该段多肽序列为基础构建原核表达载体.采用PCR方法扩增450 bp cDNA片段,编码DjHSP70 C-端150个氨基酸多肽.将双酶切的cDNA片段与pET-28a载体连接后导入BL21受体菌,在IPTG诱导下表达出21 ku融合蛋白,分子量与预期相符.该融合蛋白采用Ni2+ -NTA agarose树脂进行纯化,纯化结果电泳检测后经灰度扫描分析显示纯度在95%以上.融合蛋白免疫新西兰大白兔获得高效价的抗血清,Western blot检测显示该抗血清不仅具有很强的特异性,而且还识别小鼠HSP70.此项工作为进一步研究淡水涡虫抗逆性奠定了基础.%In this paper, hydrophilic domain of DjHSP70 was predicted by using intemet biosoftware. The DjHSP70 C-terminal contains numerous hydrophilic amino acids. Based on this hydrophilic domain of DjHSP70, the prokaryotic expression vector was constructed. PCR method was used to amplify 450 bp cDNA fragment encoding DjHSP70 C-terminal 150 amino acid polypeptides. After digested by Hind Ⅲ/Xho Ⅰ , this eDNA fragment was ligated to pET-28a expression vector. Recombinant plasmid was transformed into E. coii BL21 and a 21 ku fusion protein was expressed after the induction with IPTG, which was in agreement with the expected molecular weight. This fusion protein was purified using Ni2+ -NTA agarose and detected by SDSPAGE electrophoresis. Grayscale scanning showed that the purity of the purified protein was over 95%. The fusion protein was used as an antigen to immunize New Zealand rabbits to prepare the polyclonal antibody. The results showed that this anti-serum was not only very specific to DjHSP70, but also recognized mouse HSP70. This work has laid the foundation for further investigating stress responses in freshwater planarians.

  13. 热应激大鼠心脏损伤与血淋巴细胞热休克蛋白70的关系%Relationship between heat stress-induced cardiomyocyte injury and HSP70 in lymphocytes in rats

    Institute of Scientific and Technical Information of China (English)

    井然; 钱捷; 弓景波; 梅竹松; 王新兴; 钱令嘉

    2012-01-01

    group,significant increase of rectal temperature in rats was observed in heat stress groups(36, 37, 38, 39 ℃)(P<0.05). Q-T interval prolongation and R wave amplitude descend were observed in electrocardiogram in heat stress groups (37, 38, 39 ℃), and the rate of abnormal electrocardiogram in heat stress groups (36, 37, 38, 39 ℃) also increased as the ambient temperature increased(P<0.05). When environmental temperature was over 38 ℃, the activity of LDH and CK-MB in serum in heat stress groups increased significantly (P<0.05). In 36 ℃. and 37 ℃ heat stress groups, HSP70 level increased remarkably (P<0.05) as ambient temperature increased. But when ambient temperature was over 38 ℃ , HSP70 expression decreased with increased temperature,while still significantly higher than the normal temperature group. Conclusion Heat stress may induce the myocardium damage and change of HSP70 expression in blood lymphocytes in rats, which suggests that the HSP70 in blood lymphocytes maybe is a molecular marker and HSP70 plays a role in myocardium injury induced by heat exposure.

  14. Plasmodium falciparum Hop (PfHop Interacts with the Hsp70 Chaperone in a Nucleotide-Dependent Fashion and Exhibits Ligand Selectivity.

    Directory of Open Access Journals (Sweden)

    Tawanda Zininga

    Full Text Available Heat shock proteins (Hsps play an important role in the development and pathogenicity of malaria parasites. One of the most prominent functions of Hsps is to facilitate the folding of other proteins. Hsps are thought to play a crucial role when malaria parasites invade their host cells and during their subsequent development in hepatocytes and red blood cells. It is thought that Hsps maintain proteostasis under the unfavourable conditions that malaria parasites encounter in the host environment. Although heat shock protein 70 (Hsp70 is capable of independent folding of some proteins, its functional cooperation with heat shock protein 90 (Hsp90 facilitates folding of some proteins such as kinases and steroid hormone receptors into their fully functional forms. The cooperation of Hsp70 and Hsp90 occurs through an adaptor protein called Hsp70-Hsp90 organising protein (Hop. We previously characterised the Hop protein from Plasmodium falciparum (PfHop. We observed that the protein co-localised with the cytosol-localised chaperones, PfHsp70-1 and PfHsp90 at the blood stages of the malaria parasite. In the current study, we demonstrated that PfHop is a stress-inducible protein. We further explored the direct interaction between PfHop and PfHsp70-1 using far Western and surface plasmon resonance (SPR analyses. The interaction of the two proteins was further validated by co-immunoprecipitation studies. We observed that PfHop and PfHsp70-1 associate in the absence and presence of either ATP or ADP. However, ADP appears to promote the association of the two proteins better than ATP. In addition, we investigated the specific interaction between PfHop TPR subdomains and PfHsp70-1/ PfHsp90, using a split-GFP approach. This method allowed us to observe that TPR1 and TPR2B subdomains of PfHop bind preferentially to the C-terminus of PfHsp70-1 compared to PfHsp90. Conversely, the TPR2A motif preferentially interacted with the C-terminus of PfHsp90. Finally, we

  15. HSF1/HSP70通路抑制c-Jun氨基末端激酶的活化保护UVA诱导的HaCaT细胞凋亡%Protection of HSF1/HSP70 pathway on UVA-induced HaCaT cells apoptosis via inhibiting the activation of c-Jun N-terminal kinase

    Institute of Scientific and Technical Information of China (English)

    王晓雯; 王春波; 李丙华; 韩彦弢

    2012-01-01

    目的 观察热休克转录因子1( HSF1)与热休克蛋白70( HSP70)对紫外线A(UVA)诱导HaCaT细胞凋亡的保护作用及其机制.方法 建立8mJ/cm2 UVA辐射损伤HaCaT细胞的病理模型.将细胞随机分为对照组、8mJ/cm2 UVA照射组、HSP70转录抑制剂组(50 μmol/L槲皮素).Honechst 33258荧光染色观察细胞凋亡;蛋白质印迹法检测UVA辐射HaCaT细胞后p-HSF1和HSP70蛋白的经时变化及UVA辐射后孵育6h JNK(c-Jun氨基末端激酶)、p-JNK的蛋白表达;Real-Time PCR检测HSP70 mRNA的表达.结果 UVA辐射后HaCaT细胞内p-HSF1、HSP70蛋白表达量均出现先增加后减少的时间依赖性趋势,其中p-HSF1于lh开始增加,3h达高峰,HSP70于6h达高峰,24h基本恢复原始水平;UVA辐射前预先加入HSP70转录抑制剂槲皮素能显著抑制HSP70 mRNA的表达,增加p-JNK的表达量,同时Honechst 33258荧光染色观察其与UVA辐射组比较凋亡率明显升高.结论 8mJ/cm2 UVA辐射HaCaT细胞在一定时间内可使HSF1活化致HSP70表达增加.HSFl/HSP70通路对UVA诱导的HaCaT细胞凋亡具有保护作用,其机制与HSP70大量表达后抑制JNK的活化有关.%Objective To investigate the protective effect of heat shock factorl ( HSF1) and heat shock protein70 ( HSP70) on ultraviolet A ( UVA ) -induced HaCaT cells apoptosis and its mechanism. Methods The apoptotic HaCaT cell model was induced by UVA irradiation (8mJ/cm ). The cells were randomly divided into three groups, including a control group, a model group (8mJ/cm UVA) and a HSP70 transcription inhibitor group (50 μmol/L quercetin). The morphologic alteration of apoptotic cells was investigated by using Hoechst 33258 fluorescent staining. Western blotting was used to investigate protein expression levels of phosphorylated HSF1 and HSP70 at different time points, as well as c-Jun N-terminal kinase ( JNK ) andphosphorylated JNK were investigated after incubating for 6 hours following UVA irradiation. HSP70 mRNA was

  16. HSP70在喉乳头状瘤和声带白斑组织中的表达及其临床意义%Expressions of HSP70 in laryngeal papilloma and vocal leukoplakia tissues and their clinical significances

    Institute of Scientific and Technical Information of China (English)

    杨田田; 傅仲鹰; 王苹; 杨立新

    2012-01-01

    目的:探讨热休克蛋白70 (HSP70)及其上游调控因子HSF1在喉乳头状瘤和声带白斑中的表达,阐明其在喉癌发生中的作用.方法:应用免疫组织化学法检测6例声带白斑(声带白斑组)、6例喉乳头状瘤(喉乳头状瘤组)和7例声带息肉(对照组)的组织标本中HSP70和HSF1蛋白的表达.结果:HSP70和HSF1蛋白在喉乳头状瘤、声带白斑中的表达高于声带息肉,差异有统计学意义(P<0.05或P<0.01);声带白斑在所有组织中表达最高.对声带息肉、喉乳头状瘤、声带白斑中HSP70蛋白表达量与HSF1表达的关系进行直线回归分析,结果显示,HSP70蛋白表达量与HSF1的变化呈现正的直线相关(r=0.867,P<0.01).结论:HSP70和HSF1在声带白斑中的高表达可能与喉癌的发生密切相关,且HSP70和HSF1可能成为喉癌早期诊断的指标.

  17. Effects of covering behavior and exposure to a predatory crab Charybdis japonica on survival and HSP70 expression of juvenile sea urchins Strongylocentrotus intermedius.

    Science.gov (United States)

    Zhao, Chong; Ji, Nanjing; Zhang, Binglong; Sun, Ping; Feng, Wenping; Wei, Jing; Chang, Yaqing

    2014-01-01

    Predation is a complex process among predator, prey and environment. Juvenile sea urchins are more susceptible to predators than adults, which affects community structure. Behavior is involved in anti-predator responses by changes in the expression of anti-predator responsive genes. Here, we investigated the effects of exposure to a predatory crab Charybdis japonica and covering behavior on survival and HSP70 expression of juvenile sea urchins Strongylocentrotus intermedius. C. japonica consumed large numbers of juvenile S. intermedius in 12 hours with a mortality of 34.17±11.43%. Covering behavior did not significantly reduce predation. Exposure to C. japonica did not significantly upregulate HSP70 expression of juvenile S. intermedius in 12 hours. Covering behavior showed no significant regulative effect on the gene expression of HSP70 of juvenile S. intermedius exposed to C. japonica for 12 hours. The results indicate that the anti-predator function of covering behavior is limited and that HSP70 expression does not appear to play an important role in the anti-predator process of S. intermedius.

  18. HDJC9, a novel human type C DnaJ/HSP40 member interacts with and cochaperones HSP70 through the J domain

    International Nuclear Information System (INIS)

    HSP40s are a subfamily of heat shock proteins (HSPs) and play important roles in regulation of cell proliferation, survival and apoptosis by serving as chaperones for HSP70s. Up to date hundreds of HSP40 proteins derived from various species ranging from Escherichia coli to homo sapiens have been identified. Here we report the cloning and characterization of a novel human type C DnaJ homologue, HDJC9, containing a typical N-terminal J domain. HDJC9 is upregulated at both mRNA and protein levels upon various stress and mitogenic stimulations. HDJC9 is mainly localized in cell nuclei under normal culture conditions while it is transported into cytoplasm and plasma membrane upon heat shock stress through a non-classical and lipid-dependent pathway. HDJC9 can interact with HSP70s and activate the ATPase activity of HSP70s, both of which are dependent on the J domain. Our data suggest that HDJC9 is a novel cochaperone for HSP70s

  19. Expressions of HSP 70 and NF-κB in the peripheral blood lymphocyte of chronic gastritis patients of different syndrome patterns

    Institute of Scientific and Technical Information of China (English)

    胡玲

    2012-01-01

    Objective To study the expressions of heat shock protein 70 (HSP 70) and nuclear factor-kappa B(NF-κB) in the peripheral blood lymphocyte of chronic gastritis (CG) patients of Pi-Wei hygropyrexia syndrome (PWHS) and Pi-qi deficiency syndrome(PQDS),and to explore their correlation with Helicobacter pylori(Hp) infection. Methods

  20. The Changes of HSP70 in Skeletal Muscle after an Exhaustive Eccentric Exercise and the Effects of Acupuncture%一次力竭离心运动后大鼠骨骼肌HSP70的变化及针刺对其影响

    Institute of Scientific and Technical Information of China (English)

    李俊平; 马延超; 张炜

    2011-01-01

    Purpose:The purpose of this study was to observe the changes of HSP70 expression in rat gastrocnemius muscle after an exhaustive eccentric exercise,and to explore the effects of acupuncture on HSP70 expression in skeletal muscle after exercise.Method:Male SD rats were randomly divided into four groups: control group,non-acupuncture group and acupuncture group after an exhaustive eccentric exercise.The rats of non-acupuncture and acupuncture group were carried out an exhaustive eccentric exercise.After the exercise,the rats of acupuncture group were handled acupuncture on the gastrocnemius vertically from the further side(the angle of the needle was 30 degree) and through the belly of gastrocnemius where the needle stayed for 5 minutes.Then the gastrocnemius was obtained from rats of different groups.The expression of HSP70 of gastrocnemius was detected by western blot and immunofluorescence histochemical method.Results:1)The expression of HSP70 of gastrocnemius in non-acupuncture group were increased markedly immediately,12 hours and 24 hours after exhaustive eccentric exercise(P0.05).2)The expression of HSP70 of gastrocnemius in acupuncture group were increased markedly only 12 hours after exhaustive eccentric exercise(P0.05).3)The expression of HSP70 of gastrocnemius in acupuncture group was significantly lower than non-acupuncture group immediately and 24 hours after exhaustive eccentric exercise(P0.05).4)The expression of HSP70 of gastrocnemius in non-acupuncture group mainly located in cytoplasm.The expression of HSP70 of gastrocnemius in acupuncture group mainly located in nucleus immediately after exhaustive eccentric exercise,and then transferred to cytoplasm gradually.Conclusions:1)Immediately after an exhaustive eccentric exercise,HSP70 expression of rat gastrocnemius were induced markedly,and then decreased gradually.2) HSP70 expression of rat gastrocnemius induced by an exhaustive eccentric exercise might be

  1. A comparative study of the timecourse of the expression of the thermo‑inducible HSP70 gene in clinical and environmental isolates of Aspergillus fumigatus.

    Science.gov (United States)

    Sharafi, Golnaz; Khosravi, Ali Reza; Vahedi, Ghasem; Yahyaraeyat, Ramak; Abbasi, Teimur

    2016-05-01

    The internal environment within animals or humans provides different conditions to invading saprophytic fungal pathogens, requiring the differential regulation of genes in comparison to environmental conditions. Understanding the mechanisms by which pathogens regulate genes within the host may be key in determining pathogen behavior within the host and may additionally facilitate further investigation into novel therapeutic agents. The heat shock protein (HSP)70 gene and its associated proteins have been frequently reported to be among the most highly expressed and dominant proteins present within various locations at physiological temperatures. The present study examined relative gene expression levels of the HSP70 gene in Aspergillus fumigatus isolates from both clinical and environmental origins, at a range of temperature points (20, 30, 37 and 42˚C) over five days, using reverse transcription‑quantitative polymerase chain reaction, comparing with a standard A. fumigatus strain incubated at 25˚C. The results indicated a differential gene expression pattern for the environmental and clinical isolates. During the five days, the HSP70 expression levels in the clinical samples were higher than in the environmental samples. However, the difference in the expression levels between the two groups at 42˚C was reduced. The mean HSP70 expression level over the five incubation days demonstrated a gradual and continual increasing trend by temperature elevation in both groups at 30, 37 and 42˚C, however, at 20˚C both groups demonstrated reduced expression. The temperature shift from 20 to 42˚C resulted in HSP70 induction and up to a 10‑ and 8.6‑fold change in HSP70 expression levels on the fifth day of incubation in the clinical and environmental groups, respectively. In conclusion, incubation at 37 and 42˚C resulted in the highest expression levels in both experimental groups, with these temperature points important for the induction of HSP70

  2. A hypoxia-induced decrease of either MICA/B or Hsp70 on the membrane of tumor cells mediates immune escape from NK cells.

    Science.gov (United States)

    Schilling, Daniela; Tetzlaff, Fabian; Konrad, Sarah; Li, Wei; Multhoff, Gabriele

    2015-01-01

    Recent findings suggest that hypoxia of the tumor microenvironment contributes to immune escape from natural killer (NK) cell-mediated cytotoxicity. Heat shock protein 70 (Hsp70) and the stress-regulated major histocompatibility class I chain-related protein A and B (MICA/B) both serve as ligands for activated NK cells when expressed on the cell surface of tumor cells. Herein, we studied the effects of hypoxia and hypoxia-inducible factor-1α (HIF-1α) on the membrane expression of these NK cell ligands in H1339 with high and MDA-MB-231 tumor cells with low basal HIF-1α levels and its consequences on NK cell-mediated cytotoxicity. We could show that a hypoxia-induced decrease in the membrane expression of MICA/B and Hsp70 on H1339 and MDA-MB-231 cells, respectively, is associated with a reduced sensitivity to NK cell-mediated lysis. A knockdown of HIF-1α revealed that the decreased surface expression of MICA/B under hypoxia is dependent on HIF-1α in H1339 cells with high basal HIF-1α levels. Hypoxia and HIF-1α did not affect the MICA/B expression in MDA-MB-231 cells but reduced the Hsp70 membrane expression which in turn also impaired NK cell recognition. Furthermore, we could show that the hypoxia-induced decrease in membrane Hsp70 is independent of HIF-1α in MDA-MB-231. Our data indicate that hypoxia-induced downregulation of both NK cell ligands MICA/B and Hsp70 impairs NK cell-mediated cytotoxicity, whereby only MICA/B appears to be regulated by HIF-1α.

  3. 西伯利亚鲟热休克蛋白HSP70cDNA的克隆、序列分析和组织分布

    Institute of Scientific and Technical Information of China (English)

    田照辉; 徐绍刚; 王巍; 胡红霞; 董颖; 宋超

    2012-01-01

    采用普通PCR和RACE技术克隆了西伯利亚鲟Acipenser baerii热休克蛋白HSP70 cDNA的全序列,该序列全长为2 343 bp,其中5'非编码区为140 bp,3'非编码区为256 bp,可读编码框(ORF)为1 947bp,编码为648个氨基酸。该氨基酸序列中含有HSP70家族的3个特征序列——IDLGTTYS、IFDLGGGTFD-VSIL和IVLVGGSTRIPKIQK,细胞质特征性保守序列为EEVD,C端重复序列为GGMP。该cDNA序列与其它生物的HSP70 cDNA序列一样具有很高的相似性。系统发育树显示,西伯利亚鲟与非洲爪蛙蟾Xenopus laevis、密西西比短吻鳄Alligator mississippiensis、美西螈Ambystoma mexicanum的亲缘关系较近。实时定量分析结果表明,水温为17.5℃时,西伯利亚鲟肝脏、鳃、脾脏、心脏、肌肉、中肠、性腺、脑8种组织中均有HSP70表达,其中HSP70在脾脏中的表达量最高,鳃中的次之,肝脏中最低(P〈0.05)。

  4. Cloning and phylogenetic analysis of HSP70 gene from cattle Theileria sergenti%牛瑟氏泰勒虫HSP70 cDNA片段的克隆与系统发育分析

    Institute of Scientific and Technical Information of China (English)

    曹世诺; 于龙政; 薛书江; 贾立军; 张守发

    2009-01-01

    从血液涂片检查为牛瑟氏泰勒虫阳性的病牛血液中提取总BNA.通过RT-PCR技术扩增出牛瑟氏泰勒虫HSP70基因,将其重组到pMD-18T Simple载体后进行克隆、序列测定及分析.结果表明该片段长1966 bp,编码620个氨基酸残基,将该基因片段序列与GenBank中13种已知梨形虫的相应序列进行比较分析,牛瑟氏泰勒虫吉林分离株与已报道的牛瑟氏泰勒虫亲缘关系最近,其次是环形泰勒虫,与马巴贝斯虫亲缘关系较远.

  5. Dexamethasone regulates CFTR expression in Calu-3 cells with the involvement of chaperones HSP70 and HSP90.

    Directory of Open Access Journals (Sweden)

    Luiz Felipe M Prota

    Full Text Available BACKGROUND: Dexamethasone is widely used for pulmonary exacerbation in patients with cystic fibrosis, however, not much is known about the effects of glucocorticoids on the wild-type cystic fibrosis channel transmembrane regulator (CFTR. Our aim was to determine the effects of dexamethasone treatment on wild-type CFTR expression. METHODS AND RESULTS: Dose-response (1 nM to 10 µM and time course (3 to 48 h curves were generated for dexamethasone for mRNA expression in Calu-3 cells using a real-time PCR. Within 24 h, dexamethasone (10 nM showed a 0.3-fold decrease in CFTR mRNA expression, and a 3.2-fold increase in αENaC mRNA expression compared with control groups. Dexamethasone (10 nM induced a 1.97-fold increase in the total protein of wild-type CFTR, confirmed by inhibition by mifepristone. To access surface protein expression, biotinylation followed by Western blotting showed that dexamethasone treatment led to a 2.35-fold increase in the amount of CFTR in the cell surface compared with the untreated control groups. Once protein translation was inhibited with cycloheximide, dexamethasone could not increase the amount of CFTR protein. Protein stability was assessed by inhibition of protein synthesis with cycloheximide (50 µg/ml at different times in cells treated with dexamethasone and in untreated cells. Dexamethasone did not alter the degradation of wild-type CFTR. Assessment of the B band of CFTR within 15 min of metabolic pulse labeling showed a 1.5-fold increase in CFTR protein after treatment with dexamethasone for 24 h. Chaperone 90 (HSP90 binding to CFTR increased 1.55-fold after treatment with dexamethasone for 24 h, whereas chaperone 70 (HSP70 binding decreased 0.30 fold in an immunoprecipitation assay. CONCLUSION: Mature wild-type CFTR protein is regulated by dexamethasone post transcription, involving cotranslational mechanisms with HSP90 and HSP70, which enhances maturation and expression of wild-type CFTR.

  6. 子痫前期患者胎盘组织中HSP70和NF-κB的表达及意义%Expression of HSP70 and NF-κB in the placental trophoblasts of preg-nant women complicated with preeclampsia

    Institute of Scientific and Technical Information of China (English)

    林培红; 胡继芬

    2016-01-01

    Objective To investigate the difference and significance between the expression of HSP70 and NF-κB in the placenta tissues of normal pregnancy and preeclampsia patients, in order to find out the relation between HSP70 and NF-κB. Methods From December 2013 to January 2015, 65 pregnant patients in the First Affiliated Hospital of Fujian Medical University were selected. There were 25 healthy pregnant women (normal pregnant group), 17 mild preeclampsia patients (mild preeclampsia group) and 23 severe preeclampsia patients (severe preeclampsia group). The expressions of HSP70 and NF-κB in the placental tissues were measured using immunohistochemistry methods and re-al-time fluorescence quantitative PCR technique. Results HSP70 and NF-κB immunohistochemical results suggested that the staining degree of the normal pregnant group, the mild and severe preeclampsia group increased gradually. The differences of the expressions of HSP70 and NF-κB between the mild, severe preeclampsia group and the normal preg-nant group were significant (P 0.05); the results of HSP70 and NF-κB real-time fluorescence quantitative PCR suggested that, the level of the expression increased in mild preeclampsia group and severe preeclampsia group, as compared with the normal pregnant group, the difference between the mild preeclampsia group, severe preeclampsia group and the normal pregnant group was statistically significant (P0.05);实时荧光定量PCR结果提示:与正常妊娠组比较, HSP70及NF-κB在轻度子痫前期组及轻度子痫前期组中的表达水平增高, 轻度子痫前期组及重度子痫前期组与正常妊娠组两两比较差异有高度统计学意义(P<0.01).相关性分析结果显示,HSP70与NF-κB呈正相关(r=0.692, P<0.01). 结论 子痫前期患者胎盘组织中HSP70和NF-κB表达增加,可能与子痫前期的发生、发展有关.

  7. Induction of specific humoral and cellular immune responses in a mouse model following gene fusion of HSP70C and Hantaan virus Gn and S0.7 in an adenoviral vector.

    Directory of Open Access Journals (Sweden)

    Linfeng Cheng

    Full Text Available Heat shock proteins (HSPs display adjuvant functions when given as fusion proteins to enhance vaccination efficiency. To evaluate enhanced potency of Hantaan virus (HTNV glycoprotein (GP and nucleocapsid protein (NP immunogenicity by heat shock protein 70 (HSP70, a recombinant adenovirus rAd-GnS0.7-pCAG-HSP70C expression vector was developed by genetically linking the HSP70 C-terminal gene (HSP70 359-610 aa, HSP70C to the Gn and 0.7 kb fragment of the NP (aa1-274-S0.7. C57BL/6 mice were immunized with these recombinant adenoviral vectors. A series of immunological assays determined the immunogenicity of the recombinant adenoviral vectors. The results showed that rAd-GnS0.7-pCAG-HSP70C induced a stronger humoral and cellular immune response than other recombinant adenoviruses (rAd-GnS0.7-pCAG and rAd-GnS0.7 and the HFRS vaccine control. Animal protection experiments showed that rAd-GnS0.7-pCAG-HSP70C was effective at protecting C57BL/6 mice from HTNV infection. The results of the immunological experiments showed that HSP70C lead to enhanced vaccine potency, and suggested significant potential in the development of genetically engineered vaccines against HTNV.

  8. Influence of atorvastatin on plasma HSP70 and HSF1 in patients with cute coronary syndrome%阿托伐他汀对急性冠脉综合征患者血浆中HSP70、HSF1的影响

    Institute of Scientific and Technical Information of China (English)

    王水侠; 刑坤

    2012-01-01

    目的:探讨阿托伐他汀对急性冠脉综合征(ACS)患者血浆中HSP70、HSF1的影响.方法:选取正常者39例,阿托伐他汀治疗组48例,酶联免疫吸附法(ELISA)测定样本血浆中HSP70、HSF1表达的变化.结果:(1)给药前,阿托伐他汀治疗组血浆中HSP70、HSF1表达明显高于对照组(P<0.05);(2)给药后,阿托伐他汀治疗组患者血浆中HSP70、HSF1表达较给药前升高明显(P<0.01).结论:阿托伐他汀可通过诱导HSP70表达增多发挥抗ACS作用.

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

    Science.gov (United States)

    Walsh, Naomi; Larkin, AnneMarie; Swan, Niall; Conlon, Kevin; Dowling, Paul; McDermott, Ray; Clynes, Martin

    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.

  10. 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.

  11. 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.

  12. 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.

  13. Carboxyl terminus of Hsp70-interacting protein (CHIP) is required to modulate cardiac hypertrophy and attenuate autophagy during exercise.

    Science.gov (United States)

    Willis, Monte S; Min, Jin-Na; Wang, Shaobin; McDonough, Holly; Lockyer, Pamela; Wadosky, Kristine M; Patterson, Cam

    2013-12-01

    The carboxyl terminus of Hsp70-interacting protein (CHIP) is a ubiquitin ligase/cochaperone critical for the maintenance of cardiac function. Mice lacking CHIP (CHIP-/-) suffer decreased survival, enhanced myocardial injury and increased arrhythmias compared with wild-type controls following challenge with cardiac ischaemia reperfusion injury. Recent evidence implicates a role for CHIP in chaperone-assisted selective autophagy, a process that is associated with exercise-induced cardioprotection. To determine whether CHIP is involved in cardiac autophagy, we challenged CHIP-/- mice with voluntary exercise. CHIP-/- mice respond to exercise with an enhanced autophagic response that is associated with an exaggerated cardiac hypertrophy phenotype. No impairment of function was identified in the CHIP-/- mice by serial echocardiography over the 5 weeks of running, indicating that the cardiac hypertrophy was physiologic not pathologic in nature. It was further determined that CHIP plays a role in inhibiting Akt signalling and autophagy determined by autophagic flux in cardiomyocytes and in the intact heart. Taken together, cardiac CHIP appears to play a role in regulating autophagy during the development of cardiac hypertrophy, possibly by its role in supporting Akt signalling, induced by voluntary running in vivo.

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

    Directory of Open Access Journals (Sweden)

    Eneida Torres Cabra

    2014-06-01

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

  15. Correlation between HSP70 and c-F0S expression and apoptosis in rats undergolng transient focal cerebral ischemla and reperfusion

    Institute of Scientific and Technical Information of China (English)

    Chen Liynn; Li Yizhao; Hah Danchun

    2000-01-01

    OBJECTIVE To compare the induction of c-Fos and HSP70 and the presence of apoptosis and the influence of Ginkgo biloba extract (EGb761) in transient focal cerebral ischemic reperfusion rats.BACKGROUND Proto ancogene activation and induction of heat shock protein (HSP) occur in response to cerebral ischemia, but the correlation between these proteins and apoptosis remains uncertain. METHODS Healthy wistar rats were randomized to the normal control group(Group A, n=4), the Sham-operated control group(Group B, n=4), the ischemia and reperfusion group(Group C, n=24), the EGb761 pre-treated ischemia and reperfusion group(Group D, n=24). The rats of Group C and Group D were subjected to transient left middle cerebral artery occlusion (MCAO) as described by Zea longa for 1 hour. RESULTS Immunohistochemical analysis revealed no c-Fos or HSP70-immunoreactivity in Group A and Group B rats. However, in Group C rats, c-Fos was expressed in ipsilateral superficial cortical layers at 1 hour after reperfusion. At 6 hours, c-Fos immunoreactivities were increased in the ipsilateral cortex and were present in the contralateral cortex, while HSP70 were induced beginning in the ipsilateral neurons of MCA distribution. At 12 hours, the expression of c-Fos reached top in superficial cortical layers. At 24 hours HSP70 immunoreactivities reached top both in ipsilateral cortex and in ipsilateral striatum. At 3 days after recirculation, HSP70 expression decreased. c-Fos expression disappeared at day 7 and HSP70 expression only occured in endothelial cells. TUNEL staining showed that there was no cell apoptosis in Gronp A or in Group B. However, in Group C, TUNEL-positive neurons were observed in the border of the penumbra-like area that surrounds the ischemic core at 6 hours following reperfusion and then the number of TUNEL-positive cells reduced gradually. The changes in expression of HSP70 and c-Fos at different time points in Group D was in accord with in Group C, but the number of

  16. Survival and hsp70 gene expression in Plutella xylostella and its larval parasitoid Diadegma insulare varied between slowly ramping and abrupt extreme temperature regimes.

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    Md Habibullah Bahar

    Full Text Available BACKGROUND: In nature, insects have evolved behavioural and physiological adaptations to cope with short term exposure to extreme temperatures. Extreme heat events may increase as a result of climate change; this in turn will affect insect population dynamics. We examined the effect of abrupt and ecologically relevant gradual exposure to high temperatures on the survival and hsp70 gene expression in diamondback moth (DBM adults and the parasitoid Diadegma insulare, as well as in parasitized and non-parasitized DBM larvae. PRINCIPAL FINDINGS: Tolerance to high temperatures in DBM adults was higher than in D. insulare adults. There was no difference in the survival of DBM adults between abrupt and ramped increases from 25 to 38°C; however, at 40°C survival was higher when the temperature increased gradually. In contrast, more D. insulare adults survived when the temperature was ramped rather than shifted abruptly to both 38 and 40°C. There was no heat stress effect of up to 40°C on the survival of either parasitized or non-parasitized DBM larvae. In adults of both species, more hsp70 expression was observed when temperatures increased abruptly to 38°C compared to ramping. In contrast, at 40°C significantly more expression was found in insects exposed to the ramping rather than the abrupt regime. Hsp70 expression level was in agreement with adult survival data and appears to be a good indicator of stress levels. In parasitized and non-parasitized larvae, hsp70 expression was significantly higher after abrupt shifts compared to ramping at both temperatures. CONCLUSIONS/SIGNIFICANCE: Hsp70 gene expression was responsive to extreme temperatures in both DBM and D. insulare, which may underlie the ability of these insects to survive in extreme temperatures. Survival and hsp70 expression upon abrupt changes are distinctly different from those after ramping indicating that experimental protocol must be considered before extrapolating laboratory

  17. Morphological and Molecular Description of Phytophthora insolita Isolated from Citrus Orchard in India

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    Swapnil Bawage

    2013-01-01

    Full Text Available Citrus, an important cash crop in India, is adversely affected by Phytophthora nicotianae, P. palmivora, and P. citrophthora. Phytophthora insolita is known to be associated with citrus and reported for the first time in India. It is a rare and poorly characterized Phytophthora species, as its natural host and pathogenic impact are unclear. Previously, it was reported only in Taiwan and China; so to confirm our suspected isolate is P. insolita, regions of internal transcribed spacer, elongation factor, beta-tubulin, and cytochrome oxidase genes were sequenced. This study provides description of the lone Indian P. insolita isolate with respect to molecular identity, morphology, mating behaviour, and pathogenicity.

  18. 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.

  19. Specific Binding of Tetratricopeptide Repeat Proteins to Heat Shock Protein 70 (Hsp70) and Heat Shock Protein 90 (Hsp90) Is Regulated by Affinity and Phosphorylation.

    Science.gov (United States)

    Assimon, Victoria A; Southworth, Daniel R; Gestwicki, Jason E

    2015-12-01

    Heat shock protein 70 (Hsp70) and heat shock protein 90 (Hsp90) require the help of tetratricopeptide repeat (TPR) domain-containing cochaperones for many of their functions. Each monomer of Hsp70 or Hsp90 can interact with only a single TPR cochaperone at a time, and each member of the TPR cochaperone family brings distinct functions to the complex. Thus, competition for TPR binding sites on Hsp70 and Hsp90 appears to shape chaperone activity. Recent structural and biophysical efforts have improved our understanding of chaperone-TPR contacts, focusing on the C-terminal EEVD motif that is present in both chaperones. To better understand these important protein-protein interactions on a wider scale, we measured the affinity of five TPR cochaperones, CHIP, Hop, DnaJC7, FKBP51, and FKBP52, for the C-termini of four members of the chaperone family, Hsc70, Hsp72, Hsp90α, and Hsp90β, in vitro. These studies identified some surprising selectivity among the chaperone-TPR pairs, including the selective binding of FKBP51/52 to Hsp90α/β. These results also revealed that other TPR cochaperones are only able to weakly discriminate between the chaperones or between their paralogs. We also explored whether mimicking phosphorylation of serine and threonine residues near the EEVD motif might impact affinity and found that pseudophosphorylation had selective effects on binding to CHIP but not other cochaperones. Together, these findings suggest that both intrinsic affinity and post-translational modifications tune the interactions between the Hsp70 and Hsp90 proteins and the TPR cochaperones.

  20. A Novel Strategy for TNF-Alpha Production by 2-APB Induced Downregulated SOCE and Upregulated HSP70 in O. tsutsugamushi-Infected Human Macrophages

    Science.gov (United States)

    Liang, Jui-Lin; Tsai, Ming-Hsien; Yen, Chia-Jung; Li, Hsiu-Wen; Chiu, Siou-Jin; Chang, Chung-Hsing; Huang, Yaw-Bin; Lin, Ming-Wei; Yoshioka, Tohru

    2016-01-01

    Orientia (O.) tsutsugamushi-induced scrub typhus is endemic across many regions of Asia and the Western Pacific, where an estimated 1 million cases occur each year; the majority of patients infected with O. tsutsugamushi end up with a cytokine storm from a severe inflammatory response. Previous reports have indicated that blocking tumor necrosis factor (TNF)-α reduced cell injury from a cytokine storm. Since TNF-α production is known to be associated with intracellular Ca2+ elevation, we examined the effect of store-operated Ca2+ entry (SOCE) inhibitors on TNF-α production in O. tsutsugamushi-infected macrophages. We found that 2-aminoethoxydiphenyl borate (2-APB), but not SKF96365, facilitates the suppression of Ca2+ mobilization via the interruption of Orai1 expression in O. tsutsugamushi-infected macrophages. Due to the decrease of Ca2+ elevation, the expression of TNF-α and its release from macrophages was repressed by 2-APB. In addition, a novel role of 2-APB was found in macrophages that causes the upregulation of heat shock protein 70 (HSP70) expression associated with ERK activation; upregulated TNF-α production in the case of knockdown HSP70 was inhibited with 2-APB treatment. Furthermore, elevated HSP70 formation unexpectedly did not help the cell survival of O. tsutsugamushi-infected macrophages. In conclusion, the parallelism between downregulated Ca2+ mobilization via SOCE and upregulated HSP70 after treatment with 2-APB against TNF-α production was found to efficiently attenuate an O. tsutsugamushi-induced severe inflammatory response. PMID:27472555

  1. Effects of heavy metals on ultrastructure and Hsp70 induction in Lemna minor L. exposed to water along the Sarno River, Italy.

    Science.gov (United States)

    Basile, A; Sorbo, S; Cardi, M; Lentini, M; Castiglia, D; Cianciullo, P; Conte, B; Loppi, S; Esposito, S

    2015-04-01

    The effects of freshwater pollution in the highly contaminated river Sarno (Campania, Southern Italy) have been evaluated using bags containing the aquatic plant Lemna minor (Lemnacee, Arales), in order to determine morpho-physiological modifications as a response to pollutants. The exposition of Lemna bags for 7 days on three different sites along the river path showed alterations in chloroplasts and vacuoles shape and organization. Moreover, some specimens were exposed in vitro at the same heavy metal (HM) concentrations measured in the polluted sites of the river, and compared with data from the bag experiment; to verify the dose and time dependent effects, samples were exposed to HM in vitro at concentrations ranging from 10(-6) to 10(-4)M up to 7 days. Transmission electron microscopy (TEM) observations on in vitro plants confirmed that ultrastructural alterations affected most of plastids and the shape of different subcellular structures, namely vacuoles; in in vitro stressed specimens, Heat Shock Proteins 70 (Hsp70) levels changed, in dependence of changing levels of HM measured in different sites along the river path. Thus L. minor exhibited a possible correlation between the levels of HM pollution and Hsp70 occurrence; interestingly, the data presented showed that copper specifically increased Hsp70 levels at concentrations detected in polluted river waters, whereas cadmium and lead did not; on the other side, the latter represent highly toxic elements when specimens were exposed to higher levels in vitro. The effects of specific elements in vitro are compared to those observed in bags exposed along the river path; thus results are examined in order to propose L. minor as an organism able to be utilized to monitor heavy metals pollution; the possibility of using Hsp70s as specific markers of HM pollution is discussed. PMID:25618632

  2. A platform to view huntingtin exon 1 aggregation flux in the cell reveals divergent influences from chaperones hsp40 and hsp70.

    Science.gov (United States)

    Ormsby, Angelique R; Ramdzan, Yasmin M; Mok, Yee-Foong; Jovanoski, Kristijan D; Hatters, Danny M

    2013-12-27

    Our capacity for tracking how misfolded proteins aggregate inside a cell and how different aggregation states impact cell biology remains enigmatic. To address this, we built a new toolkit that enabled the high throughput tracking of individual cells enriched with polyglutamine-expanded Htt exon 1 (Httex1) monomers, oligomers, and inclusions using biosensors of aggregation state and flow cytometry pulse shape analysis. Supplemented with gel filtration chromatography and fluorescence-adapted sedimentation velocity analysis of cell lysates, we collated a multidimensional view of Httex1 aggregation in cells with respect to time, polyglutamine length, expression levels, cell survival, and overexpression of protein quality control chaperones hsp40 (DNAJB1) and hsp70 (HSPA1A). Cell death rates trended higher for Neuro2a cells containing Httex1 in inclusions than with Httex1 dispersed through the cytosol at time points of expression over 2 days. hsp40 stabilized monomers and suppressed inclusion formation but did not otherwise change Httex1 toxicity. hsp70, however, had no major effect on aggregation of Httex1 but increased the survival rate of cells with inclusions. hsp40 and hsp70 also increased levels of a second bicistronic reporter of Httex1 expression, mKate2, and increased total numbers of cells in culture, suggesting these chaperones partly rectify Httex1-induced deficiencies in quality control and growth rates. Collectively, these data suggest that Httex1 overstretches the protein quality control resources and that the defects can be partly rescued by overexpression of hsp40 and hsp70. Importantly, these effects occurred in a pronounced manner for soluble Httex1, which points to Httex1 aggregation occurring subsequently to more acute impacts on the cell. PMID:24196953

  3. A Platform to View Huntingtin Exon 1 Aggregation Flux in the Cell Reveals Divergent Influences from Chaperones hsp40 and hsp70*

    Science.gov (United States)

    Ormsby, Angelique R.; Ramdzan, Yasmin M.; Mok, Yee-Foong; Jovanoski, Kristijan D.; Hatters, Danny M.

    2013-01-01

    Our capacity for tracking how misfolded proteins aggregate inside a cell and how different aggregation states impact cell biology remains enigmatic. To address this, we built a new toolkit that enabled the high throughput tracking of individual cells enriched with polyglutamine-expanded Htt exon 1 (Httex1) monomers, oligomers, and inclusions using biosensors of aggregation state and flow cytometry pulse shape analysis. Supplemented with gel filtration chromatography and fluorescence-adapted sedimentation velocity analysis of cell lysates, we collated a multidimensional view of Httex1 aggregation in cells with respect to time, polyglutamine length, expression levels, cell survival, and overexpression of protein quality control chaperones hsp40 (DNAJB1) and hsp70 (HSPA1A). Cell death rates trended higher for Neuro2a cells containing Httex1 in inclusions than with Httex1 dispersed through the cytosol at time points of expression over 2 days. hsp40 stabilized monomers and suppressed inclusion formation but did not otherwise change Httex1 toxicity. hsp70, however, had no major effect on aggregation of Httex1 but increased the survival rate of cells with inclusions. hsp40 and hsp70 also increased levels of a second bicistronic reporter of Httex1 expression, mKate2, and increased total numbers of cells in culture, suggesting these chaperones partly rectify Httex1-induced deficiencies in quality control and growth rates. Collectively, these data suggest that Httex1 overstretches the protein quality control resources and that the defects can be partly rescued by overexpression of hsp40 and hsp70. Importantly, these effects occurred in a pronounced manner for soluble Httex1, which points to Httex1 aggregation occurring subsequently to more acute impacts on the cell. PMID:24196953

  4. Nucleotides Flanking the Start Codon in hsp70 mRNAs with Very Short 5'-UTRs Greatly Affect Gene Expression in Haloarchaea.

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    Wenchao Chen

    Full Text Available Leaderless translation is prevalent in haloarchaea, with many of these leaderless transcripts possessing short 5'-untranslated regions (UTRs less than 10 nucleotides. Whereas, little is known about the function of this very short 5'-UTR. Our previous studies determined that just four nucleotides preceded the start codon of hsp70 mRNA in Natrinema sp. J7, with residues -3A and +4G, relative to the A of the ATG start codon, acting as the preferred bases around the start codon of all known haloarchaeal hsp70 genes. Here, we examined the effects of nucleotides flanking the start codon on gene expression. The results revealed that shortening and deletion of the short 5'-UTR enhanced transcript levels; however, it led to significant reductions in overall translational efficiency. AUG was efficiently used as start codons, in both the presence and absence of short 5'-UTRs. GUG also could initiate translation, even though it was so inefficient that it would not be detected without considerably elevated transcript. Nucleotide substitutions at position -4 to +6 were shown to affect gene expression by transcript and/or translational levels. Notably, -3A and A/U nucleotides at position +4~+6 were more optimal for gene expression. Nucleotide transversions of -3A to -3C and +4G to +4T with hsp70 promoter from either Haloferax volcanii DS70 or Halobacterium salinarum NRC-1 showed the same effects on gene expression as that of Natrinema sp. J7. Taken together, our results suggest that the nucleotides flanking the start codon in hsp70 mRNAs with very short 5'-UTRs play an important role in haloarchaeal gene expression.

  5. Generation of the First TCR Transgenic Mouse with CD4+ T Cells Recognizing an Anti-inflammatory Regulatory T Cell-Inducing Hsp70 Peptide

    Science.gov (United States)

    Jansen, Manon A. A.; van Herwijnen, Martijn J. C.; van Kooten, Peter J. S.; Hoek, Aad; van der Zee, Ruurd; van Eden, Willem; Broere, Femke

    2016-01-01

    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 (Hsp) 70-derived peptide B29 and its mouse homologs mB29a and mB29b induced anti-inflammatory responses. Furthermore, B29 induced antigen-­specific 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 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. PMID:27014269

  6. Generation of the First TCR Transgenic Mouse with CD4(+) T Cells Recognizing an Anti-inflammatory Regulatory T Cell-Inducing Hsp70 Peptide.

    Science.gov (United States)

    Jansen, Manon A A; van Herwijnen, Martijn J C; van Kooten, Peter J S; Hoek, Aad; van der Zee, Ruurd; van Eden, Willem; Broere, Femke

    2016-01-01

    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 (Hsp) 70-derived peptide B29 and its mouse homologs mB29a and mB29b induced anti-inflammatory responses. Furthermore, B29 induced antigen--specific 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 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. PMID:27014269

  7. Analyses of promoter-proximal pausing by RNA polymerase II on the hsp70 heat shock gene promoter in a Drosophila nuclear extract.

    OpenAIRE

    Li, B.; Weber, J. A.; Chen, Y; Greenleaf, A L; Gilmour, D S

    1996-01-01

    Analyses of Drosophila cells have revealed that RNA polymerase II is paused in a region 20 to 40 nucleotides downstream from the transcription start site of the hsp70 heat shock gene when the gene is not transcriptionally active. We have developed a cell-free system that reconstitutes this promoter-proximal pausing. The paused polymerase has been detected by monitoring the hyperreactivity of thymines in the transcription bubble toward potassium permanganate. The pattern of permanganate reacti...

  8. Caries Experience among Females aged 16–21 in Punjab, India and its Relationship with Lifestyle and Salivary HSP70 Levels

    OpenAIRE

    Kaur, Rabinder; Kataria, Hardeep; Kumar, Sushil; Kaur, Gurcharan

    2010-01-01

    Objectives: The objective of the current study was to assess the prevalence and means of dental caries experience and the prevalence of calculus among college students and their awareness of general dental health. The correlation between the experience of caries and extracellular levels of heat shock protein-70 (eHSP70) was studied in saliva samples of these students to ascertain whether eHSP level changes as a result of increased caries experience of these subjects. Methods: The current stud...

  9. 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.

  10. Allele-specific PCR genotyping of the HSP70 gene polymorphism discriminating the green and red color variants sea cucumber (Apostichopus japonicus)

    Institute of Scientific and Technical Information of China (English)

    Jung-Ha Kang; Ki Hwan Yu; Jung-Youn Park; Chul-Min An; Je-Cheon Jun; Sang-Jun Lee

    2011-01-01

    Color variation is a well-known feature of sea cucumbers (Apostichopus japonicus),which are classified into three groups based on their colors of red,green and black.It is also one of the most important traits related to how they taste,and it thereby affects their market price.Attempts were made to identify single-nucleotide polymorphisms (SNPs) and to analyze differences associated with SNP genotypes between green and red color variants using HSP70 as the target gene.The HSP70 gene,which is found universally in organisms from bacteria to humans,is one of the most evolutionarily conserved genes and the most widely studied biomarker of stress response.DNA fragments of 1074 bp covering a partial sequence of the sea cucumber HSP70 gene,were amplified from both red and green variants,and subsequently analyzed for the presence of SNPs.Twenty-seven polymorphic sites in total,including heterozygous sites,were observed.Of these,six sites were found to be significantly different SNP genotypes between green and red variants.Furthermore,PCR with an internal primer designed to include an allelespecific SNP at the 3' end (site 443) showed differentiation between the two variants,100% and 4.2% amplification in green and red variants,respectively.The validated SNPs may serve as informative genetic markers that can be used to distinguish variants at the early developmental stage,prior to color differentiation.

  11. 温室粉虱和烟粉虱3个隐种中热激蛋白基因hsp70和hsp90含量的比较分析%Comparative analysis of the copy numbers of hsp70 and hsp90 in Trialeurodes vaporariorum and three cryptic species of Bemisia tabaci complex (Hemiptera: Aleyrodidae)

    Institute of Scientific and Technical Information of China (English)

    韦姣; 吕志创; 王韧; 万方浩

    2014-01-01

    [目的]昆虫适应新环境的能力与其对温度的耐受能力密切相关.热激蛋白HSP70和HSP90具有提高生物体温度耐受性的功能.烟粉虱Bemisia tabaci (Gennadius)不同隐种和不同种粉虱对温度的适应能力有差异,这与它们的热激蛋白基因拷贝数的差异可能相关.[方法]利用实时荧光定量PCR方法,检测入侵型烟粉虱MED隐种和MEAM1隐种、本地型烟粉虱AsiaⅡ1隐种以及温室粉虱Trialeurodes vaporariorum (Westwood)基因组DNA中热激蛋白基因hsp70和hsp90的拷贝数.[结果]不同种类的粉虱和烟粉虱不同隐种体内的hsp70和hsp90的含量均有较大差异,其中hsp70和hsp90拷贝数在入侵型烟粉虱MED和MEAM1隐种中含量较其他两种均高,而在土著种AsiaⅡ1隐种中含量最低,在温室粉虱中居中.此外,相同物种雌雄成虫hsp70和hsp90的拷贝数也不同,雌虫hsp70和hsp90拷贝数约为雄虫的2倍.[结论]不同种粉虱及烟粉虱不同隐种的hsp70和hsp90的拷贝数可能与其耐热性差异相关.本研究为解释不同种粉虱、烟粉虱不同隐种及其不同性别的耐热性差异机制提供了进一步的依据.

  12. 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)

    Alderson, T Reid; Kim, Jin Hae; Cai, Kai; Frederick, Ronnie O; Tonelli, Marco; Markley, John L

    2014-11-25

    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 (1)H(N) line widths in its two-dimensional (1)H-(15)N 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 (L(387)LLDVIPLS(395)) stimulated its ATPase activity and induced widespread NMR chemical shift perturbations indicative of a binding interaction in the trans configuration. PMID:25372495

  13. Responsiveness of different citrus genotypes to the Xanthomonas citri ssp. citri-derived pathogen-associated molecular pattern (PAMP) flg22 correlates with resistance to citrus canker.

    Science.gov (United States)

    Shi, Qingchun; Febres, Vicente J; Jones, Jeffrey B; Moore, Gloria A

    2015-06-01

    The bacterial agent of citrus canker disease (Xanthomonas citri ssp. citri, Xcc) has caused tremendous economic losses to the citrus industry around the world. Pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI) is important to plant immunity. In this study, we compared the defence responses of citrus canker-resistant and citrus canker-susceptible genotypes to the Xcc-derived PAMP flg22 (Xflg22) by analysing the expression of 20 citrus defence-associated genes. We showed that, in the most resistant genotype, 'Nagami' kumquat, there was significant induction of several defence genes (EDS1, NDR1, PBS1, RAR1, SGT1, PAL1, NPR2 and NPR3) as early as 6 h and up to 72 h after Xflg22 treatment. At the other end of the spectrum, highly susceptible 'Duncan' grapefruit showed no induction of the same defence genes, even 120 h after treatment. Citrus genotypes with partial levels of resistance showed intermediate levels of transcriptional reprogramming that correlated with their resistance level. Xflg22 also triggered a rapid oxidative burst in all genotypes which was higher and accompanied by the induction of PTI marker genes (WRKY22 and GST1) only in the more resistant genotypes. Pretreatment with Xflg22 prior to Xcc inoculation inhibited bacterial growth in kumquat, but not in grapefruit. A flagellin-deficient Xcc strain (XccΔfliC) showed greater growth increase relative to wild-type Xcc in kumquat than in grapefruit. Taken together, our results indicate that Xflg22 initiates strong PTI in canker-resistant genotypes, but not in susceptible ones, and that a robust induction of PTI is an important component of citrus resistance to canker.

  14. Construction of Eukaryotic Coexpression Vector Containing Mycobacterium Tuberculosis Heat Shock Protein 70 and Herpes Simplex Virus Thymidine Kinase Genes%mtHSP70/HSV-TK双基因真核共表达质粒的构建

    Institute of Scientific and Technical Information of China (English)

    曾曙光; 刘启才; 王素文; 徐平平; 章锦才; 张积仁

    2011-01-01

    目的 构建结核杆菌热休克蛋白70(mycobacterium tuberculosis heat-shock proteins 70,mtHSP70)和自杀基因单纯疱疹病毒-胸苷激酶(herpes simplex virus-thymidinekinase,HSV-TK)双基因真核共表达质粒pCMV-mtHSP70-IRES-TK.方法 登录Genbank查询HSV-TK和mtHSP70基因mRNA序列,应用引物设计软件DNA club分别设计扩增HSV-TK和mtHSP70基因全长cDNA的特异引物.以pcDNA3-TK质粒或mtHSP70质粒DNA为模板,分别采用各自的引物,用prime STAR HS DNA Polymerase进行聚合酶链反应(polymerase chain reaction,PCR),获得HSV-TK和mtHSP70基因cDNA片断,连接到T载体pMD18-T上,转化大肠杆菌TG1后用HSV-TK和mtHSP70基因的特异引物进行菌落PCR,确定含有阳性mtHSP70或HSV-TK重组质粒并将阳性pMD18T-TK重组质粒和pMD18T-mtHSP70重组质粒进行上、下游测序,测序结果与基因序列进行同源比较.选定正确的质粒用于HSV-TK和mtHSP70基因的亚克隆,构建质粒PCMV-mtHSP70-IRES-TK,所得阳性克隆摇菌后少量提取质粒,分别用NotⅠ单酶切及EcoRⅠ和XhoⅠ双酶切进行酶切鉴定.结果 构建的pCMV-mtHSP70-IRES-TK质粒分别用NotⅠ单酶切及EcoRⅠ/XhoⅠ双酶切进行酶切鉴定,酶切产物电泳见特异酶切图谱,确定为重组质粒pCMV-mtHSP70-IRES-TK.结论 本实验为后续研究mtHSP70/HSV-TK双基因的协同抗肿瘤作用奠定了实验基础.%Objective To construct an eukaryotic plasmid PCMV-mtHSP70-IRES-TK that contains and expresses both mycobacterium tuberculosis heat shock protein 70 (mtHSP70) gene and suicide gene HSV-TK for the experiments. Methods Login on genbank and search for mRNA sequences of HSV-TK and mtHSP70 genes. Specific primers for the amplification of full length cDNA of HSV-TK and mtHSP70 were designed by primer designing software DNA club. pcDNA3-TK plasmid and mtHSP70 plasmid DNA were used as template. PCR was performed with respective primers and prime STAR HS DNA polymerase, cDNA segments of HSV-TK and mtHSP70

  15. 钙/钙调素依赖性蛋白激酶Ⅱ对热休克小鼠胚胎成纤维细胞HSP70表达的影响%Calcium/Calmodulin-dependent Protein Kinase Ⅱ Contributes to HSP70 Expression in Mouse Embryonic Fibroblasts

    Institute of Scientific and Technical Information of China (English)

    孙琪; 丛霞; 索佳佳; 曹荣峰; 姜忠玲; 崔凯; 高善颂; 田文儒

    2012-01-01

    The objectives of the study were to verify if the CaMK II (Ca +/calmodulin-dependent protein kinase II ) influences the expression of HSP70 gene in the mouse embryonic fibroblasts (MEF) with heat shock and to clarify its mechanism. The fibroblasts were heat-treated at 37t ,39^ and 41°C individually before the expressions of both CaMK II and HSF1 were detected by RT-PCR at 0. 5 ,1 ,1. 5 and 2 h of heat shock ,respectivelly. Moreover, the fibroblasts were divided into control group and myr-AIP group randomly,and cultured at 371 and 39°C respectively. The expressions of both HSP70 and HSF1 were detected. The expressions of HSF1 and CaMK II mRNA in the MEF treated at 39^ for 1 h were both extremely significantly higher (P <0. 01) than that of the control group. There was a significantly (P <0. 05) decrease of HSP70 expression in the MEF cultured at 39t treated with myr-AIP compared with its blank control. However, there was no difference in HSP70 exoression between the MEF cultured at 37X1 with or without myr-AIP. The expressions of HSF1 in the MEF treated with myr-AIP were not significantly influenced. However,the p-HSFl expression in the MEF treated with myr-AIP was significantly (P <0. 05) decreased. Moderate heat shock increases both expressions of CaMK FJ and HSF1 in mouse embryonic fibroblasts. CaMK II participates in both heat shock response and HSP70 gene expression by p-HSFl.%为证实热休克小鼠胎儿成纤维细胞中钙/钙调素依赖性蛋白激酶Ⅱ(Ca2+/calmodulin-dependent protein kinaseⅡ,CaMKⅡ)对热休克蛋白70 (Heat shock protein70,HSP70)基因表达的影响及其作用机理,将体外培养的小鼠胎儿成纤维细胞(Mouse Embryonic Fibroblasts,MEF)随机分为39℃和41℃热处理组(分别处理0.5,1,1.5,2h)和常温对照组(37℃),测定各组细胞CaMKⅡ和HSF1 mRNA的量.此外,将培养的细胞分为37℃和39℃CaMKⅡ特异性抑制剂(myr-AIP)处理组和相应的空白对照组,分别测定各组细胞HSP70、HSF1

  16. Specific anti-tumor immune responses of dendritic cells pulsed with recombinant human rhHSP70 and freeze-thaw cellular lysates derived from breast cancer%rhHSP70联合冻融抗原修饰树突状细胞诱导的抗乳腺癌作用*

    Institute of Scientific and Technical Information of China (English)

    李斌; 陈鹏; 郑建云

    2013-01-01

      目的:利用rhHSP70联合树突状细胞递呈肿瘤抗原的特性提高细胞毒T淋巴细胞(CTLs)对乳腺癌细胞的杀伤活性。方法:外周血单个核细胞体外经GM-CSF和IL-4诱导产生树突状细胞,负载冻融抗原肽的同时加入新型热休克蛋白(rhHSP70),不同分组分别诱导自体CTLs产生。ELISA测定CTLs杀伤活性和细胞因子的分泌。结果:冻融抗原肽致敏的DCs促进CTLs增殖,上调CTLs中CD3+和CD8+T细胞群及Th1型细胞因子的分泌;体外实验中具有对人乳腺癌细胞MCF-7的杀伤活性,在加入rhHSP70后效果更加明显,并能显著增强CTLs对肿瘤细胞的杀伤率。结论:rhHSP70联合肝癌冻融抗原修饰DCs,能够促进DCs的成熟,增强DCs刺激淋巴细胞增殖的能力,诱导的CTLs在体外对乳腺癌细胞能产生高效杀伤力。rhHSP70增强DCs抗肿瘤能力的机制可能与其促进DCs成熟有关。%Objective:This work aims to use the characteristics of dendritic cells (DCs) pulsed with recombinant human HSP70, which can present and process tumor antigens, to enhance the killing activity of cytotoxic t lymphocytes (CTLs) against breast neoplasms. Methods:Autologous DCs were isolated from peripheral blood mononuclear cells and then stimulated in vitro with granulocyte macrophage-colony stimulating factor and interleukin-4. The DCs were loaded with A549 tumor cell freeze-thaw lysate, and rhHSP70 was added as an immune adjuvant. The specific groups were subjected to tumor-specific cytotoxic assay, enzyme-linked immunosorbent assay, and fluores-cence-activated cell sorting. Results:DCs pulsed with A549 tumor cell lysate enhanced the growth expansion of CTLs, upregulated CD40 and CD80 populations in CTLs, and augmented Th1 cytokines. In addition, the cytotoxicity of specific CTLs against A549 was highly enhanced. The above indications became more obvious after the addition of rhHSP70. Conclusion:DCs pulsed with freeze-thaw cell

  17. 中耳细菌感染诱导的急性期HSP-70相关反应的免疫组织化学研究%Immunocytochemical research of HSP-70 correlative response in acute stage induced by bacterial infected middle ear

    Institute of Scientific and Technical Information of China (English)

    周永清; 邹静; 毕爱芳; 尚耀东

    2001-01-01

    探讨中耳细菌感染急性期时,哺乳动物中、内耳热休克反应的部位,以及中耳细菌感染诱生的热休克蛋白是否可能引发内耳自身免疫损伤。运用中耳注射肺炎克雷伯杆菌制成豚鼠中耳急性感染动物模型,分别于接种后第1、3、5、7天处死动物取材。应用免疫组化技术,研究了中耳粘膜和耳蜗表达热休克蛋白70(HSP-70)的部位。结果表明:正常状态下,中耳粘膜表层的上皮细胞和内耳膜迷路血管纹、螺旋韧带、Corti氏器均有弱的阳性反应,感染应激后,上述同样部位均有强的阳性显色。不同取材时段显示的阳性位置无差异。说明在中耳急性细菌感染期,中耳粘膜和内耳组织均表达了同源HSP-70蛋白分子,这些同源HSP-70为引发内耳自身免疫损伤提供了物质基础。%To illustrate the location of HSP-70 (heat\\|shock protein\\|70)correlative response in acute stage between middle and inner ear when mammalian middle ear infected by bacteria, and investigate the possibility to cause autoimmune inner ear injure by HSP-70 that induced by otitis media. An animal model of acute otitis media was established by inoculating Klebsiella pneumoniae (K.p) into middle ear of guinea pigs. Animals were sacrificed 1,3,5 and 7days after inoculation.Investigation of the locations of HSP-70 in middle ear mucosa and cochlea structures were detected by immunocytochemical technique .The results showed that light stain of HSP-70 existed in the superficial epithelium cells of normal mucosa and cochlea structures such as stria vascularis, spiral ligament and Corti′s organ.Heavy stain appeared in the same sites in the infected ears . No different positive sites was observed in all animals.Our findings suggested that HSP-70 were expressed in middle ear mucosa and inner ear structures during acute middle ear infective stage.The HSP-70 provide a material base for causing autoimmune inner ear injure

  18. Transcriptional responses of xenobiotic metabolizing enzymes, HSP70 and Na+/K+ -ATPase in the liver of rabbitfish (Siganus oramin) intracoelomically injected with amnesic shellfish poisoning toxin.

    Science.gov (United States)

    Wang, Lin; Liang, Xu-Fang; Huang, Yan; Li, Shi-Ying; Ip, Kok-Chao

    2008-06-01

    Amnesic shellfish poisoning toxin domoic acid (DA) is a marine neurotoxin that accumulates in fish and shellfish, and has been implicated to be involved in human and marine wildlife mortality. The transcriptional responses of cytochrome P-450 1A (CYP1A), glutathione S-transferase alpha (GSTA), glutathione S-transferase rho (GSTR), heat shock protein 70 (HSP70), and Na(+)/K(+)-ATPase alpha 1 (ATP1A1) in the liver of rabbitfish (Siganus oramin) intracoelomically injected with DA, were investigated. Experimental fish were administered with one injection of DA (2 microg/g wet weight) or PBS as control. After 24 h, fish were killed and hepatic RNA was isolated. Partial cDNA of rabbitfish CYP1A, GSTA, GSTR, HSP70, ATP1A1, and beta-actin were obtained by PCR using degenerate primers. Using beta-actin as an external control, the relative liver CYP1A, GSTA, GSTR, HSP70, and ATP1A1 mRNA abundance of rabbitfish were determined by semi-quantitative RT-PCR within the exponential phase. The ratio CYP1A/beta-actin mRNA (%) of exposure group was determined to be 148.92+/-12.69, whereas the ratio of control group was 82.3+/-8.35, indicating that CYP1A was induced significantly in rabbitfish following DA exposure (P0.05). The induction of hepatic CYP1A in response to DA suggests a potential role for fish phase I xenobiotic metabolizing enzyme in DA metabolism.

  19. Cloning, Prokaryotic Expression of Echinococcus granulosus Heat Shock Protein 70 and Preparation of It's Antiserum%细粒棘球绦虫Hsp70基因的克隆、表达及抗体制备

    Institute of Scientific and Technical Information of China (English)

    赵莉; 薛晶; 石保新; 陈皓斐; 张文宝; 马正海; 张壮志; 张旭; 古努尔·吐尔逊; 米晓云; 金映红

    2012-01-01

    [Objective] The objective of the experiment is to express and purify E. granulosus (Eg) heat shock protein 70 (EgHsp70) in E. coli and prepare the antibody against E. granulosus. [ Methods] EgHsp70 gene was amplified by PCR and cloned into prokaryotic expression vector pMAL-p2x, and the recombinant plasmid was transformed into E. coil BL-21. The soluble expression conditions of fusion protein were optimized by induction with different concentrations, of IPTG different temperatures and cultivation times. The expressed fusion protein was purified by Mal-tag Magnetic Beads. To prepare the anti-serum, New Zealand white rabbits were immunized with purified EgHsp70 protein via hypodermic and volar. Western blot was used to determine the serum's specificity against EgHsp70 and native proteins. The serum titers were analyzed by ELISA. [Results] Full-length of EgHsp70 gene had an open reading frame of 765 bps encoding a protein mass of 68.6 kD. Restriction endonuclease analysis and DNA sequencing showed that EgHsp70 was cloned into the plasmid pMAL-p2x. Based on the optimization experiments, it was concluded that the best soluble expression conditions for the EgHsp70 protein are using 0.3 mmol· L-1 IPTG when bacterial cells growing to OD600 0.6 and induced for 4 h at 30℃. ELISA and Western blotting showed that the titers of the anti-serum were above 1 : 256 000, and the anti-serum could specifically bind with EgHsp70 protein and native proteins. [Conclusion] The EgHsp70 fusion protein was obtained by expressing in E.coli and purifying, and the antibody against EgHsp70 was prepared with the fusion protein immunized New Zealand white rabbits. This work will provide an antigen and detection antibody for further study on the EgHsp70 function. The protein is immunogenic and can be a vaccine candidate against Echinococcus infection.%[目的]克隆细粒棘球绦虫(Echinococcus granulosus,E.g)热休克蛋白家族基因Hsp70,在原核细胞中表达、纯化Hsp70蛋白并制

  20. Phylogenetic and Molecular Variability Studies Reveal a New Genetic Clade of Citrus leprosis virus C

    Science.gov (United States)

    Ramos-González, Pedro Luis; Chabi-Jesus, Camila; Guerra-Peraza, Orlene; Breton, Michèle Claire; Arena, Gabriella Dias; Nunes, Maria Andreia; Kitajima, Elliot Watanabe; Machado, Marcos Antonio; Freitas-Astúa, Juliana

    2016-01-01

    Citrus leprosis virus C (CiLV-C) causes a severe disease affecting citrus orchards in the Western hemisphere. This study reveals the molecular variability of the virus by analyzing four genomic regions (p29, p15, MP and RNA2-intergenic region) distributed over its two RNAs. Nucleotide diversity (π) values were relatively low but statistically different over the analyzed genes and subpopulations, indicating their distinct evolutionary history. Values of πp29 and πMP were higher than those of πp15 and πRNA2–IR, whereas πMP was increased due to novel discovered isolates phylogenetically clustered in a divergent clade that we called SJP. Isolate BR_SP_SJP_01 RNA1 and RNA2 sequences, clade SJP, showed an identity of 85.6% and 88.4%, respectively, with those corresponding to CiLV-C, the type member of the genus Cilevirus, and its RNA2 5′-proximal region was revealed as a minor donor in a putative inter-clade recombination event. In addition to citrus, BR_SP_SJP_01 naturally infects the weed Commelina benghalensis and is efficiently transmitted by Brevipalpus yothersi mites. Our data demonstrated that negative selection was the major force operating in the evaluated viral coding regions and defined amino acids putatively relevant for the biological function of cilevirus proteins. This work provides molecular tools and sets up a framework for further epidemiological studies. PMID:27275832

  1. 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

  2. 热应激预处理影响鼠背轴型皮瓣HSP70、HSF1表达的实验研究%Study of Heat Stress Pretreatment Effect on Expressions of HSP70,HSF1 in Axial Flap of Rats

    Institute of Scientific and Technical Information of China (English)

    刘哲伟; 苏开新; 陈军; 姚建; 李辉; 王海华; 赵鑫

    2014-01-01

    目的探讨热应激预处理(HSP)对缺血皮瓣组织的保护作用及其与皮瓣中热休克蛋白70(HSP70)和热休克因子1(HSF1)的关系。方法将96只雄性Sprague Dawley 大鼠随机分为缺血组与实验组(HSP+缺血组)。缺血组:皮瓣模型制备后,阻断皮瓣蒂部第1h、2h、4h、6h、8h、10h共6个时间点,每时间点后观测皮瓣存活面积、HSP70、HSF1的变化,并进行统计学分析;实验组( HSP+缺血组):先予以加热预处理,之后与缺血组相同。结果单纯缺血组皮瓣成活率下降率较实验组(HSP+缺血组)明显,差异有显著性(P<0.05),热应激预处理各组皮瓣的HSP70、HSF1合成量与皮瓣存活率呈正相关(r=0.7577,P<0.01)。结论热应激预处理能够改善皮瓣组织的存活率,HSP70、HSF1在热应激预处理改善皮瓣存活率的保护作用中发挥着重要的作用。%Objective This study was to reveal the continuous changes of HSP70 and HSF1 in the delayed and pro-tection of heat stress pretreatment on ischemic preconditioning axial flap in rats. Methods Animals were divided into two groups randomly:HSP+IPC group,ischemic pretreatment group( IPC) . In IPC group,rats skin flaps received ischemic pretreatment for 1 h,2 h,4 h,6 h,8 h,10 h,and the survival rate of the skin flaps,the contents of HSP70 and HSF1 were examined at the 6 time points. HSP+IPC group was given heat stress pretreatment and then the same treatment as IPC group. Results The survival rates of IPC descends more significantly than that of HSR+IPC. The survival rates of skin flaps have positive correlation with the contents of HSP70 and HSF1(r=0. 7577,P<0. 01). Conclusions Heat stress pretreatment can improve the survival rate of ischemia flap and the mechanism of the protection can be related to the func-tion of HSP70 and HSF1.

  3. HSP70在化学性缺氧诱导毛细胞损伤中的作用%Roles of heat shock protein 70 in the cellular injury induced by chemical hypoxia in auditory cells

    Institute of Scientific and Technical Information of China (English)

    曹磊; 张世平; 胡亿文

    2010-01-01

    目的:探讨热休克蛋白70(heat shock protein 70,HSP70)在氯化钴(cobalt chloride,CoCl2)诱导听细胞损伤中的作用.方法:应用CoCl2处理HEI-0C1听细胞,建立化学性缺氧诱导耳蜗外毛细胞损伤的体外模型.细胞计数试剂盒-8(cel]count kit 8,CCK-8)比色法检测细胞存活率,免疫印迹法(Western b1ot)检测HSP70的蛋白表达.结果:在150~750 μmol/L浓度范围内,不同浓度的CoCl2处理听细胞24 h可引起明显的细胞毒性,使细胞存活率呈剂量依赖性地降低.300 μmol/LCoCl2作用听细胞30~240min可时间依赖性地上调HSP70的蛋白表达.HSP70抑制剂槲皮素(quercetin,Quer)可在蛋白水平下调CoCl2引起的听细胞内HSP70表达增加.Ouer通过抑制HSP70的表达可加重CoCl2诱导的听细胞缺氧性损伤.结论:适应性HSP70表达增加可保护听细胞对抗缺氧诱导的损伤.

  4. 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. PMID:25245143

  5. 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. PMID:20444508

  6. Cytoplasmic Hsp70 promotes ubiquitination for endoplasmic reticulum-associated degradation of a misfolded mutant of the yeast plasma membrane ATPase, PMA1.

    Science.gov (United States)

    Han, Sumin; Liu, Yu; Chang, Amy

    2007-09-01

    Cells have a variety of strategies for dealing with misfolded proteins. Heat shock response involves transcriptional induction of chaperones to promote and/or correct folding, and also activation of the ubiquitin/proteasome system to degrade defective proteins. In the secretory pathway, it is primarily luminal misfolded or unassembled proteins that trigger the unfolded protein response which, like heat shock, induces chaperones and components of the endoplasmic reticulum (ER)-associated degradation (ERAD) pathway. To understand cellular response to a misfolded polytopic membrane protein of the secretory pathway, we studied Pma1-D378S, a model ERAD substrate. Expression of misfolded Pma1 induces heat shock response in the absence of increased temperature. Overexpression of HSF1, the transcription factor that mediates heat shock response, increases degradation of Pma1-D378S without temperature upshift. Nevertheless, efficient Pma1-D378S degradation occurs in an hsf1 mutant that maintains basal transcription levels but cannot mediate transcriptional activation. Thus, heat shock protein induction enhances but is not necessary for ERAD. The Ssa group of cytoplasmic Hsp70 chaperones is required for ERAD of both Pma1-D378S and another transmembrane ERAD substrate, Ste6*. In the absence of Ssa chaperones, ubiquitination of both substrates is impaired, resulting in stabilization. We suggest a role for Hsp70 cytoplasmic chaperones in recognition by the endoplasmic reticulum-associated ubiquitination machinery.

  7. 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.

  8. 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

    International Nuclear Information System (INIS)

    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.

  9. 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.

  10. 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.

  11. 乐至黑山羊与藏山羊HSC70、HSP70.1和HSF1基因的mRNA表达差异分析%Analysis of mRNA Differential Expression of HSC70、HSP70.1 and HSF1 Gene in Lezhi Black Goat and Tibetan Goat

    Institute of Scientific and Technical Information of China (English)

    牛惠然; 字向东; 任陶; 王永; 任称罗尔日; 朱万岭; 付锡三

    2013-01-01

    In order to investigate the relationship between eco-adaptation and heat shock protein70, the heat shock cognate protein 70 (HSC70), heat shock protein 70. 1 (HSP70. 1) and the heat shock transcription factor 1 (HSF1) genes in five tissue samples of Lezhi black goat and Tibetan goat were determined by qPCR. The results showed that the expression of HSC70 mRNA in Tibetan goat's heart was 4. 5 times to that of the Lezhi black goat (P<0. 05) , but in pituitary, Lezhi black goat is 3. 7 times to Tibetan goat. Its expression in the ovary, liver, uterus, was not different between the two breeds. The expression of HSF1 mRNA in Tibetan goat ovarian, heart was 7. 9, 3. 1 times to that of the Lezhi black goat (F< 0. 05), however, in the liver and uterus tissue samples, Lezhi black goat was in high expression levels than Tibetan goat (6. 3 fold, 10. 4 fold; P<0. 05). The expression of HSP70. 1 mRNA in Lezhi black goat pituitary was 6. 8 times to Tibetan goat (P<0. 05). It could be concluded that the difference in the expression of HSC70, HSP70. 1 and HSF1 wight be related to the eco-environment and rearing methods (grazing) in Lezhi black goat and Tibetan goat.%试验旨在利用qPCR技术对热休克同源蛋白(HSC70)、热休克蛋白70.1(HSP70.1)和热休克转录因子1(HSF1)基因在乐至黑山羊和藏山羊五个组织样中的表达量进行研究,以探讨热休克蛋白家族70 (HSP70)与山羊生态适应性的关系.结果表明,HSC70mRNA在藏山羊心脏中的表达量是乐至黑山羊中的4.5倍(P<0.05),但在脑垂体中乐至黑山羊的表达量是藏山羊的3.7倍(P<0.05),在卵巢、肝脏和子宫中两品种间无显著差异;HSF1 mRNA在藏山羊卵巢、心脏中的表达量分别是乐至黑山羊的7.9倍和3.1倍(P<0.05),在肝脏和子宫中乐至黑山羊的表达量却分别是藏山羊的6.3倍和10.4倍(P<0.05);HSP70.1 mRNA在乐至黑山羊脑垂体中的表达量是藏山羊的6.8倍(P<0.05).说明HSC70、HSP70.1

  12. Fractionated radiotherapy is the main stimulus for the induction of cell death and of Hsp70 release of p53 mutated glioblastoma cell lines

    International Nuclear Information System (INIS)

    Glioblastoma multiforme (GBM) is the most common primary brain tumor in adults. Despite a multimodal therapy consisting of resection followed by fractionated radiotherapy (RT) combined with the chemotherapeutic agent (CT) temozolomide (TMZ), its recurrence is almost inevitable. Since the immune system is capable of eliminating small tumor masses, a therapy should also aim to stimulate anti-tumor immune responses by induction of immunogenic cell death forms. The histone deacetylase inhibitor valproic acid (VPA) might foster this. Reflecting therapy standards, we applied in our in vitro model fractionated RT with a single dose of 2Gy and clinically relevant concentrations of CT. Not only the impact of RT and/or CT with TMZ and/or VPA on the clonogenic potential and cell cycle of the glioblastoma cell lines T98G, U251MG, and U87MG was analyzed, but also the resulting cell death forms and release of danger signals such as heat-shock protein70 (Hsp70) and high-mobility group protein B1 (HMGB1). The clonogenic assays revealed that T98G and U251MG, having mutated tumor suppressor protein p53, are more resistant to RT and CT than U87MG with wild type (WT) p53. In all glioblastoma cells lines, fractionated RT induced a G2 cell cycle arrest, but only in the case of U87MG, TMZ and/or VPA alone resulted in this cell cycle block. Further, fractionated RT significantly increased the number of apoptotic and necrotic tumor cells in all three cell lines. However, only in U87MG, the treatment with TMZ and/or VPA alone, or in combination with fractionated RT, induced significantly more cell death compared to untreated or irradiated controls. While necrotic glioblastoma cells were present after VPA, TMZ especially led to significantly increased amounts of U87MG cells in the radiosensitive G2 cell cycle phase. While CT did not impact on the release of Hsp70, fractionated RT resulted in significantly increased extracellular concentrations of Hsp70 in p53 mutated and WT glioblastoma

  13. 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

  14. HSP70和NF-κB在大鼠肾缺血再灌注损伤中的表达及洛伐他汀的干预研究%Intervention of Lovastatin in the expression of HSP70 and NF-κB after renal ischemia-reperfusion injury in rats

    Institute of Scientific and Technical Information of China (English)

    李鹏; 郝雷; 高进; 傅亮

    2013-01-01

    目的 探讨洛伐他汀(Lovastatin,Lov)对大鼠肾缺血再灌注损伤(RIRI)热休克蛋白70 (HSP70)和核因子-κB (NF-κB)表达的影响.方法 90只Wistar大鼠随机分为3组:假手术(Sham)组、缺血再灌注模型(IR)组、洛伐他汀(Lov)组.IR组和Lov组大鼠采用夹闭肾动脉60 min后再灌注的方法制备RIRI模型,Lov组于动脉夹闭前给予洛伐他汀2mg/(kg·d)灌胃,各组大鼠在再灌注4、12、24 h时分别处死10只取标本.测定各组大鼠的血肌酐(Scr)、尿素氮(BUN)水平,免疫组化法检测肾组织HSP70和NF-κB表达,镜下观察肾组织形态学改变.结果 与Sham组大鼠相比,IR组大鼠在再灌注4、12、24 h时点的Scr和BUN含量均显著升高;肾组织HSP70和NF-κB表达均明显增强,差异均有统计学意义(P均<0.05).与IR组相比,Lov组大鼠在各时点的Scr和BUN含量均显著降低(P均<0.01);HSP70表达均升高(P均<0.05);NF-κB表达减少,再灌注4、12 h时,两者差异有统计学意义(P<0.05);镜下病理改变减轻.结论 Lov可通过增加肾脏HSP70、下调NF-κB达而抑制炎症反应过程,减轻肾脏损害,对大鼠RIRI起到一定的保护作用.%Objective To investigate the possible effects of Lovastatin on the expression of HSP70 and NF-KB in rats after renal ischemia-reperfusion injury (RIRI). Methods Ninety Wistar rats were randomly divided into 3 groups, sham-operated group, IR group and Lov group. The model of RIRI in IR and Lov groups was established by the clamping of the kidney artery for 60 minutes. Lovastatin was given 2mg/(kg·d) by oral gavage to the rats in the Lov group before clamping of the kidney artery. Ten rats in each group were executed at 4, 12, and 24 hours after reperfusion. The concentrations of serum creatinine (Cr) and urea nitrogen (BUN) were detected, and the expression of HSP70 and NF-KB in renal tissues was detected by immunohistochemistry. The pathologic changes of renal tissues were observed by microscope

  15. 亚硝酸盐急性胁迫对中华绒螯蟹幼体相关免疫指标和应激蛋白(HSP70)表达的影响%Effects of Acute Nitrite Exposure on Immunity Indicators and HSP70 Expression in Chinese Mitten-hand Crab (Eriocheir sinensis)

    Institute of Scientific and Technical Information of China (English)

    洪美玲; 陈立侨; 孙新谨; 李二超; 顾顺樟; 禹娜

    2011-01-01

    为探讨亚硝酸盐急性胁迫对中华绒螯蟹机体免疫力的影响,以及机体在胁迫过程中的应对策略,分别测定了幼蟹(平均体重3.25 g)在10、20、30、40 mg L-1业硝酸盐暴露组和对照组中血细胞总数(Total hemocyte counts,THC)、超氧化物歧化酶( Superoxide dismutase,SOD)活力、丙二醛(Malodiadchydc,MDA)含量以及热休克蛋白 70( Heat stress protein 70,HSP70)表达量在受胁迫2d内的动态变化.结果表明:1)低浓度亚硝酸盐(10 mg L-1)胁迫组THC在胁迫3h时显著高于对照组,肝胰腺中SOD活性在胁迫3h和6h时亦显著提高,表现出一定的“毒物兴奋效应”2)但随着暴露时间的延长和亚硝酸盐浓度的升高,幼蟹THC减少,肝胰腺SOD活性下降,而MDA含量升高3)对照组中有少量的HSP70表达.幼蟹经10、20 mg L-1亚硝酸盐处理后,鳃组织中HSP70表达量在胁迫6h后达到最大,而经30、40 mg L-1 亚硝酸盐处理后,HSP70表达量在胁迫3h后达到最大.各浓度组所诱导的HSP70最大表达量与水中亚硝酸盐的浓度呈正相关.以上结果说明亚硝酸盐胁迫可通过加剧组织中脂质过氧化的方式导致机体免疫力下降,肝胰腺中MDA的含量变化可作为业硝酸盐胁迫的有效评价指标之一,而HSP70表达量的变化可作为早期预警指标应用于亚硝酸盐胁迫评价中.图2表1参36%Nitrite is an intermediate product either in the bacterial nitrification of ammonia, or in the bacterial denitrification of nitrate. The concentration of nitrite may reach 50 mg L-1 in intensive aquacultural systems due to an imbalance in biofilter resulting from overcrowding effect and poor water recirculation. High levels of nitrite in water were potential factors triggering stress on aquatic organisms. The sensitive indicators of stress evaluation for the Chinese mitten-hand crab Eriocheir smensis were selected to investigate the effect of ambient nitrite on the immunity response. In this experiment, the

  16. 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.

  17. BAG-1/SODD, HSP70, and HSP90 are potential prognostic markers of poor survival in node-negative breast carcinoma.

    Science.gov (United States)

    Davidson, Ben; Valborg Reinertsen, Kristin; Trinh, Don; Reed, Wenche; Bøhler, Per Johannes

    2016-08-01

    The objective of this study was to analyze the expression and clinical role of 13 signaling molecules in a large cohort of breast carcinoma patients with long follow-up period. Breast carcinomas (n=410) were analyzed for protein expression of phosphorylated mitogen-activated protein kinases (p-ERK, p-JNK, p-p38) and phosphoinositide 3-kinase signaling pathway proteins (p-AKT, p-mTOR, p-p70S6K); the BAG family proteins BAG-1 and BAG-4/SODD; the antiapoptotic protein Bcl-2; the inhibitor of apoptosis family member Survivin; and the heat shock protein family members HSP27, HSP70, and HSP90. Protein expression was studied for association with clinicopathological parameters and survival. Significantly higher expression of p-AKT (Pmarkers in node-negative breast carcinoma that merit further research. PMID:27038683

  18. 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.

  19. Expression of HSP70 family and HSP90 family in skeletal muscles of transport stressed pigs%运输应激猪骨骼肌中热应激蛋白HSP70和HSP90家族的表达

    Institute of Scientific and Technical Information of China (English)

    鲍恩东; Sultan,KR; 等

    2001-01-01

    To demonstrate the expression of heat shock protein s(HSP)in skeletal muscles and to estimate the relationship between expression of HSP and pork quality, using Western blot four kinds of HSP which belongs to HSP70 family and HSP90 family respectively in porcine skeletal muscle tissues were determined after a long distance transport.All four HSP,namely HSP70,HSP72,HSP86 and HSP90,were regularly detected in M.longissimus dorsi and M.gluteus medius of both the transported and the cont rol group.The expression of HSP in control pigs showed that HSPs have some indep endent physiological from stress response undetermined functions.after 6 h trans port,the induction of the HSP70 and HSP72 in skeletal muscle tissues was slightly elevated(P>0.05).However,expression of the HSP 86 and the HSP90,especially HSP90,markedly decreased(P<0.01)in skelet al muscles.The results are implied that the reduction tendency of HSP90 fa mily is relative to pork quality and may serve as a potential marker for stress response.%利用Western blot技术,检测长途运输试验猪骨骼肌(背最长肌和臀大肌)中分属于HSP70家族和HSP90家族的4种应激蛋白(HSP70,HSP 72,HSP86和HSP90)的表达。所有运输应激猪和对照猪肌肉组织中均检测到了上述4种HSP。对照猪组织中HSP的表达说明HSP除了在受应激的细胞内行使生理功能外,还具有独立于应激刺激应答以外的其它作用。6 h的长途运输后,HSP的表达量明显不同,HSP70和HSP72在肌肉组织中的表达虽然有一定的增加,但统计学分析差异不显著(P>0.05);而HSP86和HSP90在骨骼肌中的表达明显下降,尤以HS P90下降最为显著(P<0.01)。提示HSP90家族成员与肉品品质相关,可能作为判断应激损伤的指征。

  20. 大鼠再生肝中hsbp1、hsf1、hsf2、hsp70表达水平改变的分析%ANALYSIS OF CHANGES ABOUT hsbp1, hsf1, hsf2 AND hsp70'S EXPRESSION LEVELS IN RAT'S REGENERATING LIVER

    Institute of Scientific and Technical Information of China (English)

    苏丽娟; 常翠芳; 韩鸿鹏; 马辉; 徐存拴

    2006-01-01

    在克隆了大鼠热休克因子结合蛋白1基因(hsbp1)全长cDNA基础上,进一步分析它在肝再生中作用.用SD纯系大鼠为材料,按Higgens等方法建立大鼠部分肝切除(PH)模型;用原位杂交等方法分析hsbp1在肝再生中表达变化;用基因表达谱芯片分析hsbp1、hsf1、hsf2和hsp70在肝再生中表达变化.原位杂交和基因表达谱芯片分析表明,PH后6h和66-144h,hsp1表达发生了有意义上调;8-16h,hsf1表达发生了有意义上调;2-16h,hsf2表达发生了有意义上调;0.5-24h,hsp70表达发生了有意义上调.假手术(只打开腹腔和翻动肝叶,但不进行部分肝切除)后0.5-2h,hsbp1表达发生了有意义下调;8-16h,hsf1表达发生了有意义上调;0-144h,hsf2未发生有意义表达变化;0.5-30h,hsp70表达发生了有意义上调.根据实验结果推测,PH后hsbp1表达上调可增加细胞内HSBP1量,促进生长、发育、分化相关基因表达和再生肝的组织结构功能重建;(假)手术后hsbp1表达下调可减少细胞内HSBP1量,有利于HSF1上调hsp70表达,提高机体和肝脏抗损伤能力.

  1. Identification and Molecular Characterization of Nuclear Citrus leprosis virus, a Member of the Proposed Dichorhavirus Genus Infecting Multiple Citrus Species in Mexico.

    Science.gov (United States)

    Roy, Avijit; Stone, Andrew L; Shao, Jonathan; Otero-Colina, Gabriel; Wei, Gang; Choudhary, Nandlal; Achor, Diann; Levy, Laurene; Nakhla, Mark K; Hartung, John S; Schneider, William L; Brlansky, Ronald H

    2015-04-01

    Citrus leprosis is one of the most destructive diseases of Citrus spp. and is associated with two unrelated virus groups that produce particles primarily in either the cytoplasm or nucleus of infected plant cells. Symptoms of leprosis, including chlorotic spots surrounded by yellow haloes on leaves and necrotic spots on twigs and fruit, were observed on leprosis-affected mandarin and navel sweet orange trees in the state of Querétaro, Mexico. Serological and molecular assays showed that the cytoplasmic types of Citrus leprosis virus (CiLV-C) often associated with leprosis symptomatic tissues were absent. However, using transmission electron microscopy, bullet-shaped rhabdovirus-like virions were observed in the nuclei and cytoplasm of the citrus leprosis-infected leaf tissues. An analysis of small RNA populations from symptomatic tissue was carried out to determine the genome sequence of the rhabdovirus-like particles observed in the citrus leprosis samples. The complete genome sequence showed that the nuclear type of CiLV (CiLV-N) present in the samples consisted of two negative-sense RNAs: 6,268-nucleotide (nt)-long RNA1 and 5,847-nt-long RNA2, excluding the poly(A) tails. CiLV-N had a genome organization identical to that of Orchid fleck virus (OFV), with the exception of shorter 5' untranslated regions in RNA1 (53 versus 205 nt) and RNA2 (34 versus 182 nt). Phylogenetic trees constructed with the amino acid sequences of the nucleocapsid (N) and glycoproteins (G) and the RNA polymerase (L protein) showed that CiLV-N clusters with OFV. Furthermore, phylogenetic analyses of N protein established CiLV-N as a member of the proposed genus Dichorhavirus. Reverse-transcription polymerase chain reaction primers for the detection of CiLV-N were designed based on the sequence of the N gene and the assay was optimized and tested to detect the presence of CiLV-N in both diseased and symptom-free plants. PMID:25423071

  2. Identification and Molecular Characterization of Nuclear Citrus leprosis virus, a Member of the Proposed Dichorhavirus Genus Infecting Multiple Citrus Species in Mexico.

    Science.gov (United States)

    Roy, Avijit; Stone, Andrew L; Shao, Jonathan; Otero-Colina, Gabriel; Wei, Gang; Choudhary, Nandlal; Achor, Diann; Levy, Laurene; Nakhla, Mark K; Hartung, John S; Schneider, William L; Brlansky, Ronald H

    2015-04-01

    Citrus leprosis is one of the most destructive diseases of Citrus spp. and is associated with two unrelated virus groups that produce particles primarily in either the cytoplasm or nucleus of infected plant cells. Symptoms of leprosis, including chlorotic spots surrounded by yellow haloes on leaves and necrotic spots on twigs and fruit, were observed on leprosis-affected mandarin and navel sweet orange trees in the state of Querétaro, Mexico. Serological and molecular assays showed that the cytoplasmic types of Citrus leprosis virus (CiLV-C) often associated with leprosis symptomatic tissues were absent. However, using transmission electron microscopy, bullet-shaped rhabdovirus-like virions were observed in the nuclei and cytoplasm of the citrus leprosis-infected leaf tissues. An analysis of small RNA populations from symptomatic tissue was carried out to determine the genome sequence of the rhabdovirus-like particles observed in the citrus leprosis samples. The complete genome sequence showed that the nuclear type of CiLV (CiLV-N) present in the samples consisted of two negative-sense RNAs: 6,268-nucleotide (nt)-long RNA1 and 5,847-nt-long RNA2, excluding the poly(A) tails. CiLV-N had a genome organization identical to that of Orchid fleck virus (OFV), with the exception of shorter 5' untranslated regions in RNA1 (53 versus 205 nt) and RNA2 (34 versus 182 nt). Phylogenetic trees constructed with the amino acid sequences of the nucleocapsid (N) and glycoproteins (G) and the RNA polymerase (L protein) showed that CiLV-N clusters with OFV. Furthermore, phylogenetic analyses of N protein established CiLV-N as a member of the proposed genus Dichorhavirus. Reverse-transcription polymerase chain reaction primers for the detection of CiLV-N were designed based on the sequence of the N gene and the assay was optimized and tested to detect the presence of CiLV-N in both diseased and symptom-free plants.

  3. 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

    Directory of Open Access Journals (Sweden)

    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.

  4. 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

    Directory of Open Access Journals (Sweden)

    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

  5. Versatile members of the DNAJ family show Hsp70 dependent anti-aggregation activity on RING1 mutant parkin C289G

    Science.gov (United States)

    Kakkar, Vaishali; Kuiper, E. F. Elsiena; Pandey, Abhinav; Braakman, Ineke; Kampinga, Harm H.

    2016-01-01

    Parkinson’s disease is one of the most common neurodegenerative disorders and several mutations in different genes have been identified to contribute to the disease. A loss of function parkin RING1 domain mutant (C289G) is associated with autosomal-recessive juvenile-onset Parkinsonism (AR-JP) and displays altered solubility and sequesters into aggregates. Single overexpression of almost each individual member of the Hsp40 (DNAJ) family of chaperones efficiently reduces parkin C289G aggregation and requires interaction with and activity of endogenously expressed Hsp70 s. For DNAJB6 and DNAJB8, potent suppressors of aggregation of polyglutamine proteins for which they rely mainly on an S/T-rich region, it was found that the S/T-rich region was dispensable for suppression of parkin C289G aggregation. Our data implies that different disease-causing proteins pose different challenges to the protein homeostasis system and that DNAJB6 and DNAJB8 are highly versatile members of the DNAJ protein family with multiple partially non-overlapping modes of action with respect to handling disease-causing proteins, making them interesting potential therapeutic targets. PMID:27713507

  6. Methoxychlor-induced alteration in the levels of HSP70 and clusterin is accompanied with oxidative stress in adult rat testis.

    Science.gov (United States)

    Vaithinathan, S; Saradha, B; Mathur, P P

    2009-01-01

    Methoxychlor, an organochlorine pesticide, has been reported to induce abnormalities in male reproductive tract. However, the insight into the mechanisms of gonadal toxicity induced by methoxychlor is not well known. We investigated whether treatment with methoxychlor would alter the levels of stress proteins, heat shock proteins (HSP), and clusterin (CLU), and oxidative stress-related parameters in the testis of adult male rats. Animals were exposed to a single dose of methoxychlor (50 mg/kg body weight) orally and were terminated at various time points (0, 3, 6, 12, 24, and 72 h) using anesthetic ether. The levels of HSP70, CLU, and the activities of superoxide dismutase (SOD), catalase, and lipid peroxidation levels were evaluated in a 10% testis homogenate. A sequential reduction in the activities of catalase and SOD with concomitant increase in the levels of thiobarbituric acid reactive substance (TBARS) was observed. These changes elicited by methoxychlor were very significant between 6-12 h of posttreatment. Immunoblot analysis of HSP revealed the expression of HSP72, an inducible form of HSP, at certain time points (3-24 h) following exposure to methoxychlor. Similarly, the levels of secretory CLU (sCLU) were also found to be elevated between 3-24 h of treatment. The present data demonstrate methoxychlor-elicited increase in the levels of inducible HSP72 and sCLU, which could be a part of protective mechanism mounted to reduce cellular oxidative damage.

  7. Bos indicus cattle possess greater basal concentrations of HSP27, alpha B-crystallin, and HSP70 in skeletal muscle in vivo compared with cattle.

    Science.gov (United States)

    Mullins, C R; Zerby, H N; Fitzpatrick, L A; Parker, A J

    2016-01-01

    The objectives of the present study were to evaluate the basal concentrations of heat shock proteins (HSP) between and cattle and to determine if HSP basal concentrations change as an animal matures. A total of 40 cattle were used in a 2 × 2 factorial design to evaluate the effects of genotype and age (heifers and mature cows) on basal concentrations of heat shock protein 27 (HSP27), α B-crystallin (Cryab), and heat shock protein 70 (HSP70). Each experimental group of 10 animals was sampled on a separate day over a period of 4 wk during July 2014. A muscle sample was collected from the longissimus thoracis (LT) and concentrations of HSP were quantified using ELISA. There were no significant differences in HSP concentration for the interaction between age and genotype or for age alone. cattle had greater ( cattle. The results of this study show that basal in vivo HSP concentrations differ between and cattle. However, further studies are needed to investigate the relationship between HSP concentrations and meat tenderness with respect to genotypes to see if HSP concentrations account for at least some variability in tenderness differences.

  8. Fasted and postprandial response of serum physiological response, hepatic antioxidant abilities and HSP70 expression in Wuchang bream (Megalobrama amblycephala fed different dietary carbohydrate levels

    Directory of Open Access Journals (Sweden)

    Chuanpeng Zhou

    2014-12-01

    Full Text Available The effect of dietary carbohydrate (CHO level on serum physiological response, hepatic antioxidant abilities and heat shock protein 70 (HSP70 expression of Wuchang bream (Megalobrama amblycephala was studied. Two isonitrogenous (28.56% crude protein and isolipidic (5.28% crude lipid diets were formulated to contain 30% or 53% wheat starch. Diets were fed for 90 days to fish in triplicate tanks (28 fish per tank. At the end of feeding trial, significantly higher serum triglyceride level, insulin level, cortisol level, and malondialdehyde (MDA content were observed in fish fed the 53% CHO diet, while significantly lower serum total protein content, alkaline phosphatase activity, superoxide dismutase activity and total antioxidative capacity were found in fish fed the 53% CHO diet compared with those fed the 30% diet. The relative level of hepatic heat shock protein 70 mRNA was significantly higher in the 53% CHO group than that in the 30% CHO at 6, 12 and 48 h after feeding. Ingestion of 53% dietary CHO impacts the nonspecific immune ability and causes metabolic stress in Megalobrama amblycephala.

  9. CHIP, a carboxy terminus HSP-70 interacting protein, prevents cell death induced by endoplasmic reticulum stress in the central nervous system.

    Science.gov (United States)

    Cabral Miranda, Felipe; Adão-Novaes, Juliana; Hauswirth, William W; Linden, Rafael; Petrs-Silva, Hilda; Chiarini, Luciana B

    2014-01-01

    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 signaling. CHIP emerge as a candidate for clinical intervention in neurodegenerative diseases associated with ER stress.

  10. Effect of Benzo[a]pyrene or/and Heat on Levels of HSP70 and HSP90 β in Mice Brain%苯并[a]芘和高温及其联合作用对小鼠脑组织HSP70和HSP90 β水平的影响

    Institute of Scientific and Technical Information of China (English)

    涂白杰; 邬堂春; 贺涵贞

    2004-01-01

    目的研究苯并[a]芘(BaP)、高温及其联合作用对小鼠脑组织热应激蛋白70(HSP70)与热应激蛋白90β(HSP90 β)表达的影响.方法将56只昆明种小鼠随机分为7组(每组8只), 即空白对照组、溶剂对照组、低剂量BaP单独染毒组、高剂量BaP单独染毒组、单独高温处理组、高温处理+低剂量BaP染毒组、高温处理+高剂量BaP染毒组.低、高剂量BaP染毒分别给予BaP 0.5mg/kg和5mg/kg,用植物油作溶剂,每周4次腹腔注射,溶剂对照组用植物油作平行处理,空白对照组不做处理.高温处理为每周4次,每次41.5~42.0℃处理2 h.实验8周后取各组小鼠脑组织制作混合匀浆,Westernblot法检测HSP70和HSP 90 β水平.结果 HSP70在高温单独作用、低剂量BaP单独作用、高温与低剂量BaP联合作用下表达明显升高.HSP90 β仅在高温与低剂量BaP联合作用及高温与高剂量BaP联合作用下表达升高.结论不同的热应激蛋白在不同条件下对高温、BaP染毒等有害因素对脑组织的损害起保护和标志作用.

  11. The Mechanism of Calcium and Calcium-activated Calmodulin Modulating the Expression of Hsp70%钙-钙调素信号通路调节热应激蛋白70表达的机制

    Institute of Scientific and Technical Information of China (English)

    齐晓楠; 赵园; 田文儒

    2015-01-01

    随着温室效应的加剧,全球气候变暖已经成为现代畜牧业生产体系所面临的严峻挑战.高温炎热气候已成为影响畜禽生长繁殖的一种主要的非生物胁迫.热应激刺激时,动物体产生一系列生理变化来提高机体的耐受性,而本文旨在阐述钙(calcium,Ca2+)-钙调蛋白(calcium-activated calmodulin,CaM)参与的耐热信号的转导机制,即热应激蛋白(heat shock proteins,Hsps)的表达机制.热应激时,细胞内Ca2+浓度上升,激活CaM及其表达量的增加,随即CaM作用于其下游的2种靶蛋白,即2条信号转导支路来提高热激转录因子l(heat shock factor 1,HSFl)的转录活性,进而调节热应激蛋白70(Hsp70)的表达.热应激时,Hsps表达的2条具体通路:①Ca2+-CaM激活下游的靶蛋白—钙/钙调素依赖性蛋白激酶Ⅱ(Ca2+/calmodulin-dependent protein kinaseⅡ,CaMKⅡ),CaMKⅡ通过促进HSF1(230)的磷酸化来提高HSF1的转录活性,进而诱导Hsp70的表达;②Ca2+-CaM激活下游另一个靶蛋白Hsp70,Hsp70通过与增多的CaM形成复合物,释放HSF1单体,增多的HSF1形成有活性的HSF1三聚体,并入核与热激元件(heat shockelement,HSE)结合,使HSF1具有转录活性,进而诱导Hsp70的表达.本文系统完整地综述了Ca2+-CaM-HSF1-Hsp70热应激信号系统,在感受、传导和响应热应激刺激时的机制.

  12. 基于18S rRNA和HSP70基因序列的隐孢子虫种系发育分析%Phylogentic Analysis of the Cryptosporidium Based on 18S rRNA Gene and HSP70 Gene Sequences

    Institute of Scientific and Technical Information of China (English)

    王进产; 菅复春; 张龙现; 宁长申; 闫文朝; 孙铭飞; 仇书兴; 卢庆斌

    2007-01-01

    为阐明河南区域隐孢子虫分子流行病学特点,用PCR技术扩增分离虫株的18S rRNA基因全序列和HSP70基因序列,并对扩增片段进行测序.用PAUP 4.0和TREEPUZZLE 4.1构建进化树,试图从分子水平证明河南省不同地区不同宿主来源隐孢子虫的遗传特征,以阐明隐孢子虫病的分子流行病学特点.通过18S rRNA基因全序列和HSP70基因序列分析,其结果:河南人源隐孢子虫分离株为Cryptosporidium parvum鼠基因型;河南鹿源隐孢子虫分离株为C. parvum鹿基因型;河南猪源隐孢子虫的2个分离株均为C.parvum猪基因I型,即C. suis;河南鹌鹑源的隐孢子虫2个分离株分别为C. baileyi和C. meleagridis;河南乌鸡源隐孢子虫和鸵鸟源隐孢子虫分离株均为C. baileyi;河南牛源隐孢子虫分离株为C. andersoni.

  13. Occurrence, molecular characterisation, and pathogenicity of Neoscytalidium dimidiatum on Citrus in Italy

    NARCIS (Netherlands)

    Polizzi, G.; Aiello, D.; Castello, I.; Vitale, A.; Groenewald, J.Z.; Crous, P.W.; Gentile, A.; La Malfa, S.

    2011-01-01

    During 2008 and 2009, a new disease was noticed in eastern Sicily (Italy) in two re-grafted citrus orchards (Citrus sinensis (L.) Osbeck) on sour orange rootstock. Symptoms consisted of blight of vigorously growing shoots and a sooty canker on shoots and rootstock trunks. A Scytalidium-like fungus w

  14. Occurrence, molecular characterisation, and pathogenicity of Neoscytalidium dimidiatum on citrus in Italy

    NARCIS (Netherlands)

    Polizzi, G.; Aiello, D.; Castello, I.; Vitale, A.; Groenewald, J.Z.; Crous, P.W.

    2011-01-01

    During 2008 and 2009, a new disease was noticed in eastern Sicily (Italy) in two re-grafted citrus orchards (Citrus sinensis (L.) Osbeck) on sour orange rootstock. Symptoms consisted of blight of vigorously growing shoots and a sooty canker on shoots and rootstock trunks. A Scytalidium-like fungus w

  15. Effect of Ganoderma lucidum spores on cytochrome C, mitochondrial Ca2+, HSP70 and BDNF in brain of epilepsy rats%灵芝孢子粉对癫痫大鼠脑组织细胞色素C、线粒体钙、HSP70和BDNF的影响

    Institute of Scientific and Technical Information of China (English)

    王淑秋; 王淑湘; 李晓捷; 姜志梅; 秦晓玉; 韩玉泽; 刘蕾; 刘君星; 王芳芳; 梁衍峰

    2011-01-01

    目的:研究灵芝孢子粉对癫痫大鼠脑组织超氧化物歧化酶(SOD)、丙二醛(MDA)、总抗氧化能力(T-AOC)、细胞色素C、热休克蛋白70(HSP70)、线粒体Ca2+和脑源性神经营养因子(BDNF)的影响,探讨灵芝孢子粉对癫痫大鼠脑细胞的保护作用机制.方法:用亚惊厥剂量的戊四氮(PTZ)复制癫痫慢性模型,采用火焰原子吸收法测定脑组织线粒体Ca2+的含量,分光光度计比色法测定SOD的活性、MDA和细胞色素C含量,免疫组织化学方法检测HSP70和BDNF的表达情况.结果:与癫痫模型组相比,灵芝孢子粉组线粒体内细胞色素C、线粒体Ca2+的含量和HSP70的表达显著增高,而胞浆内细胞色素C含量则明显降低;脑组织SOD和T-AOC的活性明显增加,而MDA含量明显降低.大脑皮质和海马区BDNF阳性细胞数癫痫模型组明显高于正常对照组 (P<0.05);灵芝孢子粉组皮质和海马区BDNF阳性神经元数目进一步增多,与癫痫模型组相比,差异显著(P<0.05).结论:灵芝孢子粉能明显降低癫痫发作对脑细胞线粒体造成的损伤,其作用机制可能是通过清除自由基、增强脑组织的抗氧化能力,从而减轻自由基对线粒体膜的损伤作用,恢复线粒体的能量代谢,减轻脑细胞的损伤与凋亡.%AIM : To investigate the effects of Ganoderma lucidum spores on superoxide dismutase( SOD ),malondialdehyde( MDA ),total antioxidative capacity( T - AOC ), cytochrome C, heat - shock protein 70 ( HSP70 ), mitochondrial Ca2+ and hrain - derived neurotrophic factor( BDNF ) in the brain tissues of epilepsy rats.METHODS : The rat hronic epilepsy model was hy intraperitoneal injection of pentetrazole( PTZ ) at a subconvulsant dose ( 32 mg/kg ).Flame atomic absorption method was used to detect the content of mitochondrial Ca2 + , and spectrophotometer colorimetry was used to measure SOD activity,MDA content,T - AOC and cytochrome C levels in rat brain tissues.HSP70 and BDNF were de termined

  16. Protective effect of SjC23-Hsp70 DNA vaccine and interleukin-12 on Schistosoma japonicum infection in water buffalos%日本血吸虫SjC23-Hsp70 DNA疫苗与IL-12对水牛保护性作用的研究

    Institute of Scientific and Technical Information of China (English)

    胡平成; 夏大; 崔虹艳; 张苹芳; 何永康; 喻鑫玲; 孙振球

    2012-01-01

    目的:研究日本血吸虫中国大陆株23 kD膜蛋白-热休克蛋白(SjC23-Hsp70)DNA疫苗联合佐剂白细胞介素12(IL-12)质粒DNA对水牛的免疫保护作用.方法:将血吸虫病非流行区8~10月龄健康水牛45头随机分为A组(SjC23-Hsp70+IL-12)、B组(SjC23+IL- 12)和C组(pVAX+IL-12),每组15头.每头牛经肩部肌注免疫3次,每次间隔28 d.末次免疫后28 d,每头牛感染日本血吸虫尾蚴1000条.解剖前2天及当天分别收集粪便1次,用定量法检测虫卵和毛蚴数.攻击感染后56天解剖所有水牛,经胸主动脉灌冲法收集成虫,计数成虫数,检测每克肝组织虫卵数.结果:A,B组与C组相比,分别获得45.70%和26.61%的减雌率,44.51%和25.84%的减虫率,41.10%和31.63%的减粪卵率,48.11%和38.07%的减毛蚴率及43.39%和31.95%的减肝卵率.A组的5个率均比B组高(P<0.05).结论:用SjC23-Hsp70 DNA疫苗和IL-12联合免疫水牛可获得明显的免疫保护作用.%Objective: To determine the immune-protective effect of Japan Schistosoma (Chinese mainland strain) 23 kD membrane protein-heat shock protein (SjC23-Hsp70) DNA vaccine plus adjuvant-induced interleukin-12 (IL-12) plasmid DNA on Schistosoma japonicum infection in water buffalos. Methods: Forty-five health water buffalos (8-10 months old) in non-endemic area of schistosomiasis were randomly assigned into group A (SjC23-Hsp70+IL-12, 300 μg), group B (SjC23+IL-12, 300 μg) and group C (pVAX+IL-12, 300 μg), 15 in each group. Each buffalo was immuned by shoulder intramuscular injection for 3 times, at an interval of 28 days. Twenty-eight days after the last immunization, each buffalo was infected with 1000 Japan cercariae of Schistosoma. Fecal examinations were conducted 2 days and 1 day before the perfusion, and on the day of perfusion. The number of hatching miracidia and eggs per gram feces was recorded. Fifty-six days after the infection, the buffalos were sacrificed and perfused via the descending

  17. 热应激预处理影响鼠背轴型皮瓣HSP70、HSF1表达的实验研究

    Institute of Scientific and Technical Information of China (English)

    刘哲伟; 苏开新; 陈军; 姚建; 李辉; 王海华; 赵鑫

    2014-01-01

    目的探讨热应激预处理(HSP)对缺血皮瓣组织的保护作用及其与皮瓣中热休克蛋白70(HSP70)和热休克因子1(HSF1)的关系。方法将96只雄性Sprague Dawley大鼠随机分为缺血组与实验组(HSP+缺血组)。缺血组:皮瓣模型制备后,阻断皮瓣蒂部第1 h、2 h、4 h、6 h、8 h、10 h共6个时间点,每时间点后观测皮瓣存活面积、HSP70、HSF1的变化,并进行统计学分析;实验组(HSP+缺血组):先予以加热预处理,之后与缺血组相同。结果单纯缺血组皮瓣成活率下降率较实验组(HSP+缺血组)明显,差异有显著性(P〈0.05),热应激预处理各组皮瓣的HSP70、HSF1合成量与皮瓣存活率呈正相关(r=0.7577,P〈0.01)。结论热应激预处理能够改善皮瓣组织的存活率,HSP70、HSF1在热应激预处理改善皮瓣存活率的保护作用中发挥着重要的作用。

  18. 热休克蛋白70过表达对骨骼肌细胞内 ATP水平的影响%Effect of over-expression Hsp70 on the intracellular ATP level in C2C12 cells

    Institute of Scientific and Technical Information of China (English)

    王磊; 王尊; 刘跃飞; 顾一煌

    2013-01-01

    Objective To examine the effect of over-expression Hsp70 on the intracellular ATP level in C2C12 cells. Methods Hsp70 gene was amplified from pAT153 plasmids and then cloned into pTRE2hyg vector. After the transfection of recombinant plasmids of pTRE2hyg-Hsp70 into the C2C12 cells, the expression of Hsp70 was examined by Western blot. Furthermore, the intracellular ATP level was evaluated in C2C12 cells at different time points (0 d, 3 d, 7 d) during cell culture. Results Compared with controls, the intracellular ATP level was significantly increased (P<0.05) at different time points,(14.5 ± 2.87)nmol/mg protein (3 d), (15.3 ± 3.12) nmol/mg protein(7 d) after transfection. Conclusion The C2C12 cells of over-expression Hsp70 can increase the intracellular ATP level, indicating that Hsp70 may play a role in the metabolism in skeletal muscle cells.%目的:探讨热休克蛋白70(heat shock protein 70,Hsp70)过表达对骨骼肌细胞(C2C12)内ATP水平的影响。方法通过构建重组pTRE2hyg-Hsp70质粒,稳定转染C2C12细胞系,建立Hsp70过表达的C2C12细胞系。分别在转染后细胞培养的不同时间点(0 d,3 d,7 d),检测细胞内ATP的水平。结果 Hsp70过表达的C2C12细胞系在培养的3 d,7 d,细胞内ATP的水平分别达到(14.5±2.87)nmol/mg蛋白质、(15.3±3.12)nmol/mg 蛋白质,与对照组相比明显增高(P<0.05)。结论 Hsp70过表达的骨骼肌细胞可以提高细胞内的ATP水平,提示Hsp70对骨骼肌细胞的能量代谢产生影响。

  19. 海豚链球菌感染对不同品系罗非鱼血液生化指标和肝脏HSP70 mRNA表达的影响%Studies on blood biochemical indices and expression of hepatic HSP70 mRNA of different tilapia strains artificially challenged with Streptococcus iniae

    Institute of Scientific and Technical Information of China (English)

    强俊; 杨弘; 王辉; 徐跑; 柒壮林; 何杰

    2012-01-01

    In order to establish the superiority, four different tilapia strains (GIFT tilapia, New GIFT tilapia, Egypt Nile tilapia and Red tilapia) were compared in the study. At day 100 tilapias in the treatment group were intraperitoneally injected with Streptococcus iniae suspension (2.95×108/mL) to evaluate disease resistance of four different tilapia strains against S. iniae in terms of the serum biochemical parameters and hepatic HSP70 mRNA expression before and after infection. Another set of twenty fish taken from each tank was challenged with S. iniae using the same method as above, and the cumulative mortality of 4 strains was recorded at different times post infection. The results showed that GIFT tilapia and New GIFT tilapia were more sensitive to S. iniae at 96 h after infection, cumulative mortalities were 36.67% and 38.33% respectively. Egypt Nile tilapia was less sensitive to pathogen and no death occurred during the experiment. The levels of serum cortisol (COR) and glucose (GLU) and expression levels of HSP70 mRNA in GIFT tilapia, New GIFT tilapia and Red tilapia strains were highly significant(P<0.05), and the activities of glutamic-pyruvic transaminase (ALT), glutamic-oxaloacetic transaminase (AST)and lysozyme (LSZ) also increased, while serum alkaline phosphatase (AKP), triglyceride and cholesterol levels were significantly lower than those prior to infection (P<0.05). By utilizing the energy from resolved hepatin and lipid, the synthesis of hepatic HSP70 mRNA and levels of LSZ and globulin in Egypt Nile tilapia were improved, and thus the non-specific immunity strengthened. It is suggested that in the course of tilapia breeding, effective combination of the disease resistance with growth be conducted so as to guarantee sustainable development of the tilapia industry.%以吉富罗非鱼、新吉富罗非鱼、埃及尼罗罗非鱼和红罗非鱼为研究对象,饲养100 d后,进行海豚链球菌(2.95×108 CFU/mL)感染试验,分析攻毒前后各品

  20. Identification of Citrus Reticulata cv.Chachiensis by ISSR molecular markers%利用ISSR分子法标记鉴别新会茶枝柑

    Institute of Scientific and Technical Information of China (English)

    邓锋; 林向华; 梁蔚阳

    2012-01-01

    Objective To identify Citrus Reticulata cv. Chachiensis from Xinhui by ISSR molecular markers from different species of Citrus Reticulata. Methods 28 ISSR primers were analyzed by PCR with total DNA extracted from fresh leaves of 16 samples of Citrus Citrus Reticulata cv. Chachiensis , reticulata ' Shiyueju' , reticulata ' Yangshan' ,reticulata' Gong' and reticulata' Lu'. Results Two ISSR primers amplified from Citrus Reticulata cv. Chachiensis products showed different electrophoresis bands with other species of Citrus Reticulata. Conclusion ISSR method provides a quick, reliable molacular marker technique for identification of Citrus Reticulata cv. Chachiensis.%目的 研究利用ISSR分子标记方法在核酸分子水平上鉴别新会茶枝柑.方法 从28条ISSR引物中筛选合适的引物,对从新会茶枝柑、砂糖桔、阳山桔、贡柑、芦柑共16个新鲜叶片样品中提取的总DNA进行PCR扩增及电泳分析,寻找特征位点.结果 新会茶枝柑所有样品与其中2条ISSR引物扩增出较为明显的特征条带,可区别于其他品种.结论 ISSR作为一种简便有效的分子标记方法,可用于新会茶枝柑的鉴别.

  1. Occurrence, molecular characterisation, and pathogenicity of Neoscytalidium dimidiatum on Citrus in Italy

    OpenAIRE

    Polizzi, G.; Aiello, D.; Castello, I.; Vitale, A.; Groenewald, J.Z.; Crous, P.W.; A. Gentile; LA MALFA, S

    2011-01-01

    During 2008 and 2009, a new disease was noticed in eastern Sicily (Italy) in two re-grafted citrus orchards (Citrus sinensis (L.) Osbeck) on sour orange rootstock. Symptoms consisted of blight of vigorously growing shoots and a sooty canker on shoots and rootstock trunks. A Scytalidium-like fungus was isolated consistently from symptomatic tissues on 2% potato-dextrose agar (PDA). The mycelium was composed of branched, septate, brown hyphae which disarticulated into 0-1-septate phragmospores....

  2. Occurrence, molecular characterisation, and pathogenicity of Neoscytalidium dimidiatum on citrus in Italy

    OpenAIRE

    Polizzi, G.; Aiello, D.; Castello, I.; Vitale, A.; Groenewald, J.Z.; Crous, P.W.

    2011-01-01

    During 2008 and 2009, a new disease was noticed in eastern Sicily (Italy) in two re-grafted citrus orchards (Citrus sinensis (L.) Osbeck) on sour orange rootstock. Symptoms consisted of blight of vigorously growing shoots and a sooty canker on shoots and rootstock trunks. A Scytalidium-like fungus was isolated consistently from symptomatic tissues on 2% potato-dextrose agar (PDA). The mycelium was composed of branched, septate, brown hyphae which disarticulated into 0-1-septate phragmospores....

  3. Gymnastics of molecular chaperones.

    Science.gov (United States)

    Mayer, Matthias P

    2010-08-13

    Molecular chaperones assist folding processes and conformational changes in many proteins. In order to do so, they progress through complex conformational cycles themselves. In this review, I discuss the diverse conformational dynamics of the ATP-dependent chaperones of the Hsp60, Hsp70, Hsp90, and Hsp100 families. PMID:20705236

  4. Application of Bioinformatics in the Study of HSP70 Recombinant Vaccine of Echinococcus Granulosus%生物信息学技术在细粒棘球蚴热休克蛋白70重组疫苗研究中的应用

    Institute of Scientific and Technical Information of China (English)

    丁淑琴; 刘宏鹏; 张爱君; 张彩芳; 赵巍

    2009-01-01

    目的 通过基因序列分析,寻找细粒棘球蚴(Echinococcus granulosus,Eg)热休克蛋白基因(Heat shock pm-tein7,HSP70),为包虫病防治筛选新的候选疫苗抗原.方法 ①登录 GenBank 公共数据库,检索 EgHSP70目的基因序列.②对克隆基因进行测序,结果输入DNAStax、BioSun分析软件和互联网 SWISS-MODEI 数据库,对重组 EgHSP70 的特性、抗原表位及构象进行预测分析.结果 ①获得了具有完整开放阅读框的 EgHSP70核苷酸序列.②DNAStar分析软件推算氨基酸序列结果显示,EgHSP70由133个理论氨基酸组成,分子量约为14.53kDa.DNAStar、BioSun分析软件确定了重组 EgHSP70 可能的抗原表位.与其他动物热休克蛋白70氨基酸的同源性分析结果显示EgHSP70具有高度的保守性.结论 生物信息学技术在 EgHSP70 重组疫苗研究中有一定的理论和应用价值.

  5. The Interactions of TPR Domains of Ppp5c/TTC16 with Hsp70/90 and Their Influence on the Cell Cycle%Ppp5c和TTC16基因的TPR结构域与Hsp70、Hsp90蛋白的相互作用及对细胞周期的影响

    Institute of Scientific and Technical Information of China (English)

    刘德康; 李蕾; 陈霞; 唐超; 李建民

    2012-01-01

    目的:研究Ppp5c及TTC16基因的TPR (tetratricopeptide repeat)结构域短片段和Hsp70及Hsp90家族蛋白的相互做用,及其过表达时细胞周期的影响.方法:通过生物信息学的分析及PCR的方法,克隆Ppp5c及TTC16基因的TPR结构域以及HSPAIA、HsP90AA1的全长基因,并连入酵母双杂交载体,通过ClonTech的酵母双杂交实验体系研究蛋白和蛋白之问的相互作用.把Ppp5c及TIC16基因的TPR结构域克隆入真核表达栽体,构架稳定表达Ppp5c及TTC16基因的TPR结构域的MCF-7细胞系,并通过流式细胞实验观察细胞周期.结果:Ppp5c及TTC16基因的TPR结构域能与HSPAIA或HsP90AAI发生相互作用.Ppp5c及TTC16基因的TPR结构域在MCF-7中的过表达能严重影响细胞周期,引起细胞凋亡和S期阻滞.结论:本实验初步揭示了不同蛋白的TPR结构域在与Hsp70及Hsp90蛋白的相互作用性质的异同点以及其过表达对细胞周期的影响,为全面理解TPR结构域的功能、PPP5c以及TTC16蛋白在细胞内的功能奠定了前期实验基础.%Objective: To investigate the interactions between the TPR (tetratricopeptide repeat) domains of Ppp5c/TTC16 and protein, of Hsp70/Hsp90, to study the effect of the overexpression of TPR domains of Ppp5c/TTC16 on cell cycle in MCF-7 cell lines. Methods: After being analyzed by online informatics tools, the TPR domains of Ppp5c/TTC16 and full-length of HSPA1A/HSP90AA1 genes were respectively sub-cloned into two separate vectors of the yeast two-hybrid system provided by ClonTech to confirm the protein-protein interactions. The TPR domains of Ppp5c/TTC16 were sub-cloned into eukaryotic expression vectors which were stably transfected into MCF-7 cell-lines, whose cell cycle was then examined by flow cytometry. Results: The TPR domains of Ppp5c/TTC16 interacted with proteins of HSPA1A/HSP90AA1. A large amount of apoptosis was detected when the TPR domains of Ppp5c over-expressed in MCF-7 cell line, and the cell cycle was

  6. EFFECTS OF ACUTE THERMAL STRESS ON HSP70 MRNA, PHYSIOLOGY AND NONSPECIFIC IMMUNITY IN SIBERIAN STURGEON (ACIPENSER BAERII)%急性热应激对西伯利亚鲟HSP70mRNA表达、血清皮质醇和非特异性免疫的影响

    Institute of Scientific and Technical Information of China (English)

    田照辉; 徐绍刚; 王巍; 胡红霞; 马国庆

    2013-01-01

    The effect of acute thermal stress on HSP1Q mRNA expression, physiology and nonspecific immunity were investigated in Siberian sturgeon. In the experiment, ten fish were transferred froml7.5℃ to 27.5℃ water, then data were measured at lh and 3h. Fish in 17.5℃ water were taken as Oh group. Respiratory rate increased from 80-90 breaths/min to 210 breaths/min, then decreased to 180 breaths/min after 0.5h. Among the three tissues including gill, spleen and brain, the HSP70 mRNA expression in gill rose most quickly after lh stress, and was about 1.63 fold compared with the Oh group (P<0.05), then remained this level to 3h; The HSP10 mRNA expression in spleen and brain had nearly no change in lh thermal stress, then rose quickly from lh to 3h in the two tissues, especially in brain at 3h rose most quickly to about 1.77 fold compared with the lh group (P<0.05). Compared with the Oh group, serum cortisol increased quickly and was 5.14 fold (PO.05) at lh, then decreased quickly to 2.1 fold at 3h. Spleen macrophage respiratory burst achieved the maximum (P<0.05) at lh, then reduced to the Oh group level. Serum complement C3 increased at lh and decreased significantly at 3h (P<0.05). Serum lysozyme activity firstly elevated then decreased but had no significant differences. Serum SOD activity decreased when thermal stress time prolonged, and achieved the minimum at 3h (P<0.05). Serum MDA content decreased when thermal stress time prolonged and had no significant difference. The results showed that lh acute thermal stress promoted the nonspecific immunity and tolerance, but 3h acute thermal stress decreased it.%为研究西伯利亚鲟(Acipenser baerii)对急性热应激的抗逆机理,将体质量为(155.47±19.50) g的鱼从17.5 ℃迅速转至27.5 ℃水中,在1h和3h取样测定HSP70 mRNA表达变化、血清皮质醇和非特异性免疫指标.结果显示:急性热应激时鳃、脾和脑的HSP70 mRNA表达量升高,具有组织特异性,热应激1h时

  7. Study on the expression of heat shock protien 70 gene and protein in adult lymphocyte of human acute leukemia%成人急性淋巴细胞白血病HSP70蛋白及mRNA的表达研究

    Institute of Scientific and Technical Information of China (English)

    吴青; 陈燕; 李新刚

    2005-01-01

    目的研究热休克蛋白70(HSP70)在成人急性淋巴细胞白血病中的表达.方法RT-PCR法和Westrn blot法检测急性淋巴细胞白血病化疗前、化疗后及正常人HSP70 mRNA及HSP70蛋白的表达.结果成人急性淋巴细胞白血病化疗前HSP70 mRNA及HSP70的蛋白表达明显低于正常人(P<0.05),化疗后明显高于正常人(P<0.05).结论化疗后能明显提高急性淋巴细胞白血病患者HSP70基因转录和蛋白的表达,而对癌细胞则起保护作用.

  8. 逆针灸"关元"穴对更年期大鼠脾脏热休克蛋白70及其mRNA表达和血清白介素-2、肿瘤坏死因子-α含量的影响%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-α in Menopausal Rats

    Institute of Scientific and Technical Information of China (English)

    李晓泓; 王洪彬; 徐莉莉; 宋晓琳; 郑玲; 何玉伟; 张露芬

    2009-01-01

    目的:观察逆针灸"关元"穴对自然更年期大鼠免疫功能的影响.方法:经阴道筛查出动情周期紊乱的10月龄SD雌性大鼠160只,随机分为自然10、12、14、16月龄组,逆灸12、14、16月龄组和逆针12、14、16月龄组.另选3.5月龄健康SD雌性大鼠16只为年轻对照组.逆灸、逆针各组从大鼠10月龄开始,每周灸或针"关元"穴2次,共8周.采用免疫组化法检测脾脏热休克蛋白70(HSP 70),原位杂交法检测HSP 70mRNA表达,放射免疫法测定血清白介素-2(IL-2)、肿瘤坏死因子-α(TNF-α)含量.结果:更年期大鼠随着月龄的增长脾脏HSP 70HSP 70 mRNA的表达呈先升后降的变化,血清IL-2含量波动性降低,TNF-α含量先降后升.与自然同月龄组相比,逆灸、逆针各月龄组HSP 70HSP 70 mRNA表达和IL-2含量均呈增加趋势,其中逆灸、逆针12月龄组HSP 70HSP 70 mRNA,逆灸14、16月龄组HSP 70,逆针14月龄组HSP 70 mRNA的表达差异具有统计学意义(P<0.01或P<0.05),IL-2含量逆灸14月龄组明显升高(P<0.05);TNF-α含量逆灸、逆针12月龄组均明显升高(P<0.01),逆灸16月龄组明显降低(P<0.05).同月龄逆灸、逆针组相比,逆针12月龄组HSP 70 mRNA显著升高(P<0.05).结论:逆针灸"关元"穴对调整更年期免疫功能紊乱具有一定作用,其机制可能与调节脾脏组织细胞HSP 70及其基因的表达,启动了机体内在的抗病与应变能力,从而调节血清免疫因子IL-2、TNF-α的含量有关.%Objective To observe the influence of preventive acupuncture (PA) and preventive moxibustion (PM) at "Guanyuan" (CV 4) on the immune function in natural climacteric rats. Methods 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

  9. H-ras transfection of the rat kidney cell line NRK-52E results in increased induction of c-fos, c-jun and hsp70 following sulofenur treatment.

    Science.gov (United States)

    Gu, H; Smith, M W; Phelps, P C; Berezesky, I K; Merriman, R L; Boder, G B; Trump, B F

    1996-09-10

    The effect of the antineoplastic drug sulofenur on the induction of the immediate-early genes (IEG) c-fos and c-jun and the stress gene hsp70 was compared in the rat kidney epithelial-like cell line NRK-52E and a derivative H-ras-transfected (H/1.2NRK-52E) cell line. Fold induction for each gene after sulofenur (500 microM) treatment was greater in H/1.2NRK-52E. The maximum increases for NRK-2E and H/1.2NRK-52E were as follows: c-fos, approximately 10-fold and approximately 18-fold; c-jun, approximately 2.5-fold and approximately 3.6-fold; hsp70, approximately 13-fold and approximately 30-fold. In cells loaded with EGTA/AM or treated in low or no Ca2+ HBSS, c-fos induction was reduced similarly in both cell types. However, inhibition of protein kinases with staurosporin and calphostin C reduced c-fos by 80% in NRK-52E but by only 10-20% in H/1.2NRK.52E. These results indicate that sulofenur-induced IEG elevation is Ca(2+)-dependent and that the requirement for protein kinase C activation is bypassed in H-ras-transfected cells.

  10. Detection and molecular identification protocols for Phyllosticta citricarpa from citrus matter

    Directory of Open Access Journals (Sweden)

    Mariette Truter

    2012-03-01

    Full Text Available Strict quarantine measures for the export of South African citrus fruit to European and US markets require the development of sensitive and accurate detection methods for the pathogen Phyllosticta citricarpa – a fungus causing citrus black spot disease. Because of the presence of other, non-pathogenic Phyllosticta species, rapid and accurate verification of the Phyllosticta species present on exported citrus fruit is important to producers, exporters and regulatory authorities to prevent unnecessary losses. We have analysed over 800 samples collected over 7 years and have compared sample preparation and detection protocols applied in different environments: nurseries, production systems including phytosanitary inspections in orchards, pack houses and export terminals in order to compile protocols for the detection of P. citricarpa. Standard procedures of sample preparation and DNA extraction were adapted to suit diverse inoculum sources. Low pathogen numbers in symptomless green leaves, for example, obliged the use of a wet-dry enrichment technique constituting the stimulation of fungal growth for easier detection. Physical maceration was adapted for sturdy material using liquid nitrogen or bead beating. The use of a two-step polymerase chain reaction (PCR with nested primers significantly increased both the sensitivity and the specificity of the PCR performed on soil samples, overcoming problems with relatively impure DNA extracts and low pathogen numbers. The assays have proven to be highly consistent, thereby providing a reliable, reproducible and highly sensitive detection and diagnostic service to the southern African citrus industries in order to sustain market access.

  11. Molecular characterization of Wolbachia strains associated with the invasive Asian citrus psyllid Diaphorina citri in Brazil.

    Science.gov (United States)

    Guidolin, A S; Cônsoli, F L

    2013-02-01

    Wolbachia is a symbiont intensively studied due to its ability to interfere with their host's reproduction, and it has been recently proposed as an alternative tool to control insect pests or vectors of diseases. The Asian citrus psyllid Diaphorina citri is an important pest of citrus since it vectors the bacterium that causes the "Huanglongbing" disease in citrus. The frequency and diversity of Wolbachia associated with D. citri is unknown, limiting the utilization of Wolbachia as an alternative strategy for insect management. Thus, we aimed to determine the natural rate of infection, to characterize the Wolbachia strains associated with this psyllid by "multilocus sequencing typing" (MLST) and wsp analysis, and to verify the association of the symbiont to particular genotypes of the host. Analysis indicated Wolbachia infects 100 % of all specimens tested from all 15 sampled populations. MLST revealed the occurrence of five new sequence types (STs) of Wolbachia, while analysis based on the wsp sequences indicated only four different types of Wolbachia. ST-173 was predominant, while the remaining STs were population specific. Analysis of the host-symbiont relationship did not reveal any particular association of Wolbachia and haplotypes or a decrease in nucleotide diversity of D. citri in populations in which more than one ST was recorded. The consequences of the diversity of STs reported are still unknown, but the fact that Wolbachia infection is fixed and that there is one ST with a broad distribution highlights the use of this symbiont as an alternative strategy to control D. citri.

  12. Echinococcus granulosus 2 heat shock protein 70 gene: construction of recombinant plasmid, prokaryotic expression, purification and characterization%细粒棘球蚴2HSP70重组质粒的构建、原核表达、纯化和初步鉴定

    Institute of Scientific and Technical Information of China (English)

    丁淑琴; 王洁; 王淑静; 张焱; 赵巍

    2006-01-01

    目的 构建细粒棘球蚴(Echinococcus granulosus,EG)2热休克蛋白70(heat shock protein,2HSP70)表达重组质粒,原核表达及纯化重组蛋白,并对重组蛋白进行免疫学特性的初步鉴定.方法 从EG2HSP70/pGEM-T/JM109质粒中分离EG2HSP70目的基因,将此片段重组入表达载体pGEX-6P-1后转化入大肠杆菌BL21,鉴定后以IPTG诱导表达,通过SDS-PAGE检测其表达水平,用亲和层析纯化并分离重组蛋白,利用Western blot来鉴定重组蛋白的免疫学特性.结果 酶切鉴定及测序分析表明,成功构建细粒棘球蚴pGEX-6P-1/EG2HSP70重组质粒;IPTG诱导表达后,融合蛋白占菌体总蛋白的39%,占裂解物上清总蛋白的70.4%,表明重组的EG2HSP70基因能在BL21中高效表达,表达的蛋白大部分以可溶性状态存在.亲和层析法纯化重组蛋白EG2HSP70,通过Western blot证实该重组蛋白能被细粒棘球蚴囊壁免疫兔血清识别.结论 成功构建细粒棘球蚴中国大陆株pGEX-6P-1/EG2HSP70重组质粒,重组的EG2HSP70能够高效表达,表达的EG2HSP70具有免疫学活性.

  13. A comparative study of survival, metabolism, immune indi-cators and HSP70 expression in three kinds of shell colors Japanese scallop Mizuhopecten yessoensis under high tem-perature stress%高温下3种壳色虾夷扇贝存活率、代谢率、免疫酶活力及HSP70表达的比较研究

    Institute of Scientific and Technical Information of China (English)

    郝振林; 刘京哲; 唐雪娇; 湛垚垚; 田莹; 丁君; 杨立猛; 常亚青

    2015-01-01

    以 3 种壳色虾夷扇贝(Mizuhopecten yessoensis)(枫叶贝、红贝、白贝)为实验材料, 将在 15℃暂养的 3 种壳色虾夷扇贝分别驯化至 20 , 22 , 24 和 26 ℃ 4 个温度梯度(升温幅度为 1 ℃/d), 待各处理组达到对应温度梯度后暂养 7 d, 测定比较 3 种壳色虾夷扇贝的存活率、耗氧率和排氨率、抗氧化酶活力和热激蛋白 HSP70 表达等指标.结果表明: 随着温度的升高, 3 种壳色的虾夷扇贝的存活率呈先上升后下降趋势, 其中, 白壳色虾夷扇贝的存活率在同一处理组中最高; 3 种壳色的虾夷扇贝耗氧和排氨率均随温度升高呈现先升高后降低的趋势, 同一温度处理组下白壳色虾夷扇贝的代谢率始终高于红贝和枫叶贝; 另外, 总抗氧化能力(total-antioxidant capacity, T-AOC)、超氧化物歧酶(activity of superoxide dismutase, SOD)、过氧化氢酶(Catalase, CAT)、过氧化物酶(Peroxidase, POD) 4 种免疫酶的活性随温度升高也呈现先升高后降低的趋势, 且同一温度处理组下白壳色虾夷扇贝的免疫酶活力始终高于红贝和枫叶贝; 3 种壳色虾夷扇贝鳃 HSP70 的表达模式基本相同, 随着温度的逐渐升高, HSP70 持续表达, 到 26℃时表达量最高, 且白壳色扇贝 HSP70 的相对表达量最高为 5.07.综上, 白壳色虾夷扇贝在高温应激下表现出较强的抵抗和耐受能力, 因此, 可将其作为后期耐高温型虾夷扇贝新品系的重要培育材料.%Three shell colors of Mizuhopecten yessoensis were used to test the survival rate, oxygen consumption rate, ammonia-N excretion rate, catalase (CAT) and activity of superoxide dismutase (SOD) activities, to-tal-antioxidant capacity (T-AOC), peroxidase (POD) contents, and HSP70 under short-term high temperature. The scallops reared in 15℃ were acclimatized to the different temperature levels 20, 22, 24 and 26℃ (temperature increased 1℃ per day), and then maintained for 7 days. The

  14. 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.

    Science.gov (United States)

    Jensen, Frank B; Gerber, Lucie; Hansen, Marie N; Madsen, Steffen S

    2015-07-01

    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-nitrosated compounds. Nitrite uptake was slightly higher in hypoxia than normoxia, and high internal nitrite levels extensively converted blood hemoglobin to methemoglobin and nitrosylhemoglobin. Hypoxia increased inducible NOS (iNOS) mRNA levels in the gills, which was overruled by a strong inhibition of iNOS expression by nitrite in both normoxia and hypoxia, suggesting negative-feedback regulation of iNOS gene expression by nitrite. A similar inhibition was absent for neuronal NOS. Branchial NKA activity stayed unchanged, but mRNA levels of the nkaα1a subunit increased with hypoxia and nitrite, which may have countered an initial NKA inhibition. Nitrite also increased hsp70 gene expression, probably contributing to the cytoprotective effects of nitrite at low concentrations. Nitrite displays a concentration-dependent switch between positive and negative effects similar to other signaling molecules. PMID:25908056

  15. Molecular characterization of a citrus yellow vein clearing virus strain from China.

    Science.gov (United States)

    Zhen, Song; Kurth, Elizabeth G; Peremyslov, Valera V; Changyong, Zhou; Dolja, Valerian V

    2015-07-01

    The complete nucleotide sequence of an isolate of citrus yellow vein clearing virus from Yunnan, China (CYVCV-RL), was determined following whole-genome amplification by RT-PCR. The CYVCV-RL genome was 7529 nt in length, excluding the 3' poly (A) tail, and contained six open reading frames (ORFs), resembling that of viruses belonging to the genus Mandarivirus in the family Alphaflexiviridae. Sequence analysis showed that the CYVCV-RL shared the greatest nucleotide sequence identity with the CYVCV-Y1 (JX040635) isolate from Turkey for the whole genome (97.1%), 5' UTR (98.7%), 3' UTR (100.0%), and each of six ORFs (96.5% to 97.8%), suggesting that there is apparent genetic stability among CYVCV isolates of different geographic origin. PMID:25913691

  16. Molecular characterization of a citrus yellow vein clearing virus strain from China.

    Science.gov (United States)

    Zhen, Song; Kurth, Elizabeth G; Peremyslov, Valera V; Changyong, Zhou; Dolja, Valerian V

    2015-07-01

    The complete nucleotide sequence of an isolate of citrus yellow vein clearing virus from Yunnan, China (CYVCV-RL), was determined following whole-genome amplification by RT-PCR. The CYVCV-RL genome was 7529 nt in length, excluding the 3' poly (A) tail, and contained six open reading frames (ORFs), resembling that of viruses belonging to the genus Mandarivirus in the family Alphaflexiviridae. Sequence analysis showed that the CYVCV-RL shared the greatest nucleotide sequence identity with the CYVCV-Y1 (JX040635) isolate from Turkey for the whole genome (97.1%), 5' UTR (98.7%), 3' UTR (100.0%), and each of six ORFs (96.5% to 97.8%), suggesting that there is apparent genetic stability among CYVCV isolates of different geographic origin.

  17. Genetic diversity and phylogenetic analysis of Citrus (L) from north-east India as revealed by meiosis, and molecular analysis of internal transcribed spacer region of rDNA.

    Science.gov (United States)

    Hynniewta, Marlykynti; Malik, Surendra Kumar; Rao, Satyawada Rama

    2014-12-01

    The north-eastern region of India is reported to be the center of origin and rich in diversity of Citrus (L.) species, where some wild and endangered species namely Citrus indica, Citrus macroptera, Citrus latipes, Citrus ichagensis and Citrus assamensis exist in their natural and undisturbed habitat. In order to have comprehensive information about the extent of genetic variability and the occurrence of cryptic genomic hybridity between and within various Citrus species, a combined approach involving morphological, cytogenetical and molecular approaches were adopted in the present study. Cytogenetic approaches are known to resolve taxonomic riddles in a more efficient manner, by clearly delineating taxa at species and sub species levels. Male meiotic studies revealed a gametic chromosome number of n = 9, without any evidence of numerical variations. Bivalents outnumbered all other types of associations in pollen mother cells (PMCs) analyzed at diplotene, diakinesis and metaphase I. Univalents were frequently encountered in nine species presently studied, though their presence appropriately did not influence the distributional pattern of the chromosomes at anaphases I and II. The molecular approaches for phylogenetic analysis based on sequence data related to ITS 1, ITS 2 and ITS 1 + 5.8 s + ITS 2 of rDNA using maximum parsimony method and Bayesian inference have thrown light on species inter-relationship and evolution of Citrus species confirming our cytogenetical interpretations. The three true basic species i.e. Citrus medica, Citrus maxima and Citrus reticulata with their unique status have been resolved into distinct clades with molecular approaches as well. C. indica which occupies a unique position in the phylogenetic ladder of the genus Citrus has been resolved as a distinct clade and almost behaving as an out-group. The presences of quadrivalents in C. indica also echo and support its unique position. From our study it is amply clear that C

  18. Genetic diversity and phylogenetic analysis of Citrus (L from north-east India as revealed by meiosis, and molecular analysis of internal transcribed spacer region of rDNA

    Directory of Open Access Journals (Sweden)

    Marlykynti Hynniewta

    2014-12-01

    Full Text Available The north-eastern region of India is reported to be the center of origin and rich in diversity of Citrus (L. species, where some wild and endangered species namely Citrus indica, Citrus macroptera, Citrus latipes, Citrus ichagensis and Citrus assamensis exist in their natural and undisturbed habitat. In order to have comprehensive information about the extent of genetic variability and the occurrence of cryptic genomic hybridity between and within various Citrus species, a combined approach involving morphological, cytogenetical and molecular approaches were adopted in the present study. Cytogenetic approaches are known to resolve taxonomic riddles in a more efficient manner, by clearly delineating taxa at species and sub species levels. Male meiotic studies revealed a gametic chromosome number of n = 9, without any evidence of numerical variations. Bivalents outnumbered all other types of associations in pollen mother cells (PMCs analyzed at diplotene, diakinesis and metaphase I. Univalents were frequently encountered in nine species presently studied, though their presence appropriately did not influence the distributional pattern of the chromosomes at anaphases I and II. The molecular approaches for phylogenetic analysis based on sequence data related to ITS 1, ITS 2 and ITS 1 + 5.8 s + ITS 2 of rDNA using maximum parsimony method and Bayesian inference have thrown light on species inter-relationship and evolution of Citrus species confirming our cytogenetical interpretations. The three true basic species i.e. Citrus medica, Citrus maxima and Citrus reticulata with their unique status have been resolved into distinct clades with molecular approaches as well. C. indica which occupies a unique position in the phylogenetic ladder of the genus Citrus has been resolved as a distinct clade and almost behaving as an out-group. The presences of quadrivalents in C. indica also echo and support its unique position. From our study it is amply

  19. Hsp70/C D80 DNA vaccine inhibits asthma by regulating the balance of Th1/Th2/Treg/Th1 7 in an acute mouse model%HSP70/C D80 DNA疫苗通过调节Th1/Th2/Treg/Th17细胞对小鼠急性哮喘的抑制作用

    Institute of Scientific and Technical Information of China (English)

    李燕; 谢敏; 史小玲; 王晓燕; 唐利; 钟森; 陈庄

    2014-01-01

    Objective To explore the mechanism of Th1/Th2/Treg/Th17 rebalance in an acute asthma mouse model treated with heat shock protein70 (Hsp70)/C D80 DNA vaccine.Methods Forty BALB/c mice were divided into the control group,asthma-model group,pVXA1-vector group and vaccine treatment group,10 in each group.They were intraperitoneally injected into ovalbumin (OVA)to build acute asthma model and treated with DNA vaccine.The air-way responsiveness was evaluated after inhaling the methacholine (Mch).The IgE content in serum was detected by ELISA.The pathological change of lung tissue was observed by HE and AB-PAS staining.The expressions of interferon-γ(IFN-γ),interleukin-4 (IL-4),ransforming growth factor beta (TGF-β)and interleukin-17 (IL-17)in BALF were measured by ELISA.The expressions of T-bet,GATA binding protein 3 (GATA-3 ),forkheah box protein3 (Foxp3 ) and retinoid related orphan receptor gamma t (RORγt)in lung tissue were detected by Real-time PCR.Results Com-pared with asthmatic mouse,mouse treated with vaccine exhibited the decreased airway responsiveness,IgE (all P<0.05)and inflammatory infiltration of lung tissues,repressed proliferation of goblet cells,and increased IFN-γ/IL-4, TGF-β/IL-17,T-bet/GATA-3 and Foxp3/RORγt (all P<0.05).Conclusion The Hsp70/C D80 DNA vaccine plays a therapeutic role in asthma by restoring the balance of Th1/Th2/Treg/Th17 cell.%目的:考察热休克蛋白70(HSP70)/C D80 DNA疫苗治疗小鼠急性哮喘的疗效,研究该过程中Th1/Th2/Treg/Th17再平衡的作用机制。方法 BALB/c小鼠40只随机分为空白对照组(空白组),哮喘模型组(模型组), pVAX1(+)空载体对照组(空载组)和 HSP70/C D80疫苗治疗组(治疗组),每组10只。鸡卵清蛋白(OVA)激发致敏小鼠构建急性哮喘模型,HSP70/C D80 DNA疫苗进行治疗。吸入性支气管激发试验测定气道反应性,ELISA法测定血清中IgE的含量,HE和AB-PAS染色观察肺组织病理学

  20. Physiological and Molecular Responses to Excess Boron in Citrus macrophylla W.

    Directory of Open Access Journals (Sweden)

    Mary-Rus Martínez-Cuenca

    Full Text Available This work provides insight into several mechanisms involved in boron (B regulation pathway in response to high B conditions in Citrus. The study was carried out in Citrus macrophylla W. (Cm seedlings cultured "in vitro" in media with 50 or 400 μM H3BO3 (control, Ct, and B-excess, +B, plants, respectively. Growth parameters, B concentration, leaf chlorophyll (Chl concentration, the expression of the main putative genes involved in B transport and distribution, and leaf and root proline and malonaldehyde (MDA concentrations, were assessed. Excess B led to high B concentration in +B plants (3.8- and 1.4-fold in leaves and roots, respectively when compared with Ct ones. However, a minor effect was recorded in the plant (incipient visual symptoms, less than 27% reduction in root growth and 26% decrease in Chl b concentration. B toxicity down-regulated by half the expression level of putative B transporter genes NIP5 and PIP1. CmBOR1 gene was not repressed in +B plants and B accumulated in the shoots. High B level increased the transcripts of putative gene TIP5, involved in B transport across the tonoplast, by 3.3- and 2.4-fold in leaves and roots, respectively. The activity of V-PPiase proton pump, related with the electrochemical gradient in the vacuole, was also enhanced in +B organs. B toxicity up-regulated putative BOR4 gene (2.1- and 2.7-fold in roots and leaves, respectively, which codifies for an active efflux B transporter. Accordingly, B was located in +B plants preferently in an insoluble form on cell walls. Finally, excess B caused a significant rise in proline concentration (51% and 34% in roots and leaves, respectively, while the MDA level did not exceed 20%. In conclusion, Cm tolerance to a high B level is likely based on the synergism of several specific mechanisms against B toxicity, including: 1/ down-regulation of NIP5 and PIP1 boron transporters; 2/ activation of B efflux from cells due to the up-regulation of putative BOR4 gene

  1. Molecular characterization of two carboxylesterase genes of the citrus red mite, Panonychus citri (Acari: Tetranychidae).

    Science.gov (United States)

    Zhang, Kun; Niu, Jin-Zhi; Ding, Tian-Bo; Dou, Wei; Wang, Jin-Jun

    2013-04-01

    The citrus red mite, Panonychus citri, is known for its ability to rapidly evolve resistance to insecticides/acaricides and to adapt to hosts that produce toxins. To get better insight into the detoxification mechanism of P. citri, two carboxylesterase (CarE) genes, PCE1 and PCE2, were isolated and characterized. PCE1 and PCE2 contained open reading frames of 1,653 and 1,392 nucleotides, encoding proteins of 550 and 463 amino acid residues, respectively. Phylogenetic analyses showed that PCE1 and PCE2 were most closely related to the CarE genes from other phytophagous mites. The transcriptional profiles of two CarE genes among developmental stages (egg, larva, nymph, adult female, and adult male), after exposing to four acaricides (avermectin, azocyclotin, pyridaben, and spirodiclofen) and acid rain were investigated using real-time quantitative PCR (qPCR). The results showed that during development, PCE1 was highly expressed at the egg stage, whereas PCE2 was abundantly expressed at the adult stage of males. The expression levels of PCE1 were highly induced upon exposure to acaricides and acid rain. On the other hand, the expression levels of PCE2 were increased after treatment with avermectin and pyridaben. These results suggest that PCE1 and PCE2 may have distinct roles in different developmental stages and participate in the detoxification of acaricides. PMID:23404785

  2. 脑损伤后热休克蛋白70及热休克因子Ⅰ基因转录的实验研究%Experimental research of the transcription of HSP70and HSF1 gene after traumatic brain injury

    Institute of Scientific and Technical Information of China (English)

    赵连旭; 徐小虎; 潘速跃; 张成

    2002-01-01

    目的为了解脑损伤后应激基因转录的状况.方法采用RT-PCR对伤后脑组织HSP70、HSF1 mRNA的数量进行测定.结果在脑损伤15 min后脑干HSP70、HSF1 mRNA水平显著升高,在损伤后迅速死亡的致死损伤组的鼠脑中HSP70、HSF1 mRNA水平亦显著升高.结论应激基因的转录水平在脑损伤后明显提高,脑应激蛋白合成增强,参与损伤与修复过程.同时由于HSP70、HSF1 mRNA对脑损伤的迅速反应能力,使之成为观察脑损伤的灵敏指标.

  3. 脑室内低温对家兔急性大脑中动脉缺血模型HSP70表达的影响%The effects of cerebral ventricle low temperature Oil the expression of heat shock protein 70 in middle cerebral artery occlusion (MCAO) rabbits

    Institute of Scientific and Technical Information of China (English)

    姚冰; 姚长义

    2007-01-01

    目的 观察脑室内低温对家兔急性大脑中动脉缺血模型热休克蛋白70((Heat shock protein 70,HSP70)表达的影响.方法 家兔16只,随机分为2组:对照组(常温生理盐水)、低温组(22℃低温生理盐水)各8只,应用线栓法建立大脑中动脉闭塞(MCA0)模型,进行脑室内穿刺低温液体灌注亚低温干预,24 h后采用SABC方法检测各组脑组织HSP70表达,TUNEL方法检测凋亡细胞.结果 低温组的TUNEL染色阳性细胞数和HSP70染色阳性细胞数明显低于及多于常温组,有显著性差异(P<0.05).结论 通过脑室内低温治疗,可以减少缺血区域凋亡细胞,并增加HSP70细胞的表达.同时改善了动物的神经系统症状.

  4. Effects of astragalus polysaccharides (APS) injection on the expression of heat stress proteins 70 (hsp70) gene in Oreochromis niloticus%注射黄芪多糖对吉富罗非鱼热应激蛋白基因表达的影响

    Institute of Scientific and Technical Information of China (English)

    汤菊芬; 吴灶和; 简纪常; 鲁义善; 王蓓

    2011-01-01

    将黄芪多糖(APS)用无菌生理盐水配制成2 mg/mL(低剂量)、20 mg/ mL(高剂量)两种针剂,腹腔注射吉富罗非鱼(Oreochromis niloticus).分别在注射后0h、24h、48 h、72 h和96 h采样提取吉富罗非鱼鳃、头肾、肝脏和脾脏组织中的总RNA,并反转录成cDNA,利用Real-time PCR方法对不同组织中的热应激蛋白(hsp70)基因表达进行定量分析.结果显示,在24h时,高剂量组吉富罗非鱼鳃、头肾和肝脏三个组织中hsp70基因的表达量显著高于同期对照组(P<0.05),之后表达量快速降低,低剂量组hsp70基因表达量仅在脾脏中有上调.实验表明,APS可通过诱导吉富罗非鱼hsp70基因表达来保护免疫细胞免受应激损伤,从而增强鱼体抵抗病原菌的能力.%Astragalus polysaccharides (APS) was dissolved in sterile saline at the doses of 2 mg/mL (low dose), 20 mg/mL (high dose) to inject Oreochromis niloticus intraperitoneally, and the effects of APS on gene expression of hsp70 in gill, head kidney, liver and spleen of 0. Niloticus were determined using quantitative real-time polymerase chain reac tion at 24 h, 48 h, 72 h and 96 h after intraperitoneal injection. The results revealed that the gene expression level of hsp70 in gill, head kidney and liver of high dose group increased significantly than those of control group (P <0. 05) at 24 h, but decreased quickly afterwards. The gene expression of hsplO was up-regulated only in the spleen in low dose group (P <0.05) at 24 h. These results indicated that APS could protect immune cell from heat injury and enhance the fish immunity against infection of bacterial pathogen by inducing the expression of hsp70 of 0. Niloticus.

  5. The destructive citrus pathogen, 'Candidatus Liberibacter asiaticus' encodes a functional flagellin characteristic of a pathogen-associated molecular pattern.

    Directory of Open Access Journals (Sweden)

    Huasong Zou

    Full Text Available Huanglongbing (HLB is presently the most devastating citrus disease worldwide. As an intracellular plant pathogen and insect symbiont, the HLB bacterium, 'Candidatus Liberibacter asiaticus' (Las, retains the entire flagellum-encoding gene cluster in its significantly reduced genome. Las encodes a flagellin and hook-associated protein (Fla of 452 amino acids that contains a conserved 22 amino acid domain (flg22 at positions 29 to 50 in the N-terminus. The phenotypic alteration in motility of a Sinorhizobium meliloti mutant lacking the fla genes was partially restored by constitutive expression of Fla(Las. Agrobacterium-mediated transient expression in planta revealed that Fla(Las induced cell death and callose deposition in Nicotiana benthamiana, and that the transcription of BAK1 and SGT1, which are associated with plant innate immunity, was upregulated. Amino acid substitution experiments revealed that residues 38 (serine and 39 (aspartate of Fla(Las were essential for callose induction. The synthetic flg22(Las peptide could not induce plant cell death but retained the ability to induce callose deposition at a concentration of 20 µM or above. This demonstrated that the pathogen-associated molecular pattern (PAMP activity of flg22 in Las was weaker than those in other well-studied plant pathogenic bacteria. These results indicate that Fla(Las acts as a PAMP and may play an important role in triggering host plant resistance to the HLB bacteria.

  6. 苯并芘处理对人支气管上皮细胞热休克蛋白70和着色性干皮病G蛋白表达的影响%Changes of Hsp70 and XPG in human bronchial epithelia cells treated with benzo[a] prrene

    Institute of Scientific and Technical Information of China (English)

    杨瑾; 刘晓勇; 邹云锋; 牛丕业; 段燕英; 邬堂春

    2010-01-01

    目的 探讨苯并芘(Benzo[a]pyrene,BaP)作用下的人支气管上皮细胞(16HBE)热休克蛋白70(Heat shock protein 70,Hsp70)和着色性干皮病G蛋白(Xeroderma pigmentosum group G,XPG)表达的时间效应特征,并分析二者之间可能存在的关联性.方法 用8 μmol/L BaP处理16HBE细胞0、1、2、4、8、12、24和48 h,以彗星实验检测细胞DNA损伤并以Olive尾矩值(Olive Tail Moments,OTM)评价DNA损伤程度,以Western-blot检测Hsp70和XPG的表达水平,并以激光共聚焦法检测二者的共定位.结果 染毒1~2h时OTM值变化率最大,与正常细胞相比,除染毒1h组外其余各组OTM值均显著性增高(P<0.01);Hsp70和XPG的表达随染毒时间逐渐增高,12 h时达到峰值,激光共聚焦结果显示BaP染毒细胞后Hsp70和XPG的结合在细胞核内增强.结论 在该试验条件下,XPG在执行核苷酸切除修复作用时可能有Hsp70的参与,其具体机制还有待其他实验进一步加以证实.

  7. 快速眼动睡眠剥夺后大鼠皮质及海马 HSP70表达的研究%Rapid eye movement sleep deprivation on expression of heat shock protein 70 in rat cortex and hippocampus

    Institute of Scientific and Technical Information of China (English)

    黄建欧; 田国红; 赵忠新; 贺斌; 黄流清

    2004-01-01

    目的探讨不同时间的快速眼动(REM)睡眠剥夺对大鼠皮质及海马各区的热休克蛋白70(HSP70)表达的影响及意义.方法60只Wistar大鼠,随机分为睡眠剥夺组(SD)、环境对照组(TC)和空白对照组(CC).其中SD组又分为1d、3d、5d、7d 4个时点.用改良多平台睡眠剥夺法进行REM睡眠剥夺,运用免疫组织化学方法观察REM睡眠剥夺后大鼠额叶、顶叶皮质及海马各区HSP70表达的分布规律及时空变化;同时结合蛋白质免疫印记(Western Blot)实验对额叶皮质及全海马HSP70蛋白作了选择性的半定量分析.结果REM睡眠剥夺后1 d脑内HSP70表达一过性增强,以后逐渐下降.Western Blot实验印证了这一结果.结论REM睡眠剥夺能够引起大鼠皮质及海马神经元内HSP70表达增强,可能是一种自身稳定调节的保护机制.

  8. La proteína de estrés calórico Hsp70 funciona como un indicador de adaptación de los bovinos a las zonas áridas

    Directory of Open Access Journals (Sweden)

    Bañuelos-Valenzuela, Rómulo, Sánchez-Rodríguez, Sergio H.

    2005-03-01

    Full Text Available El propósito del presente estudio fue seleccionar genotipos de ganado bovino que estén mas adaptados al estrés calórico en las zonas áridas, a través de la expresión de las proteínas de choque calórico (Hsp, las cuales, son constituyentes normales de las células y su síntesis se incrementa con la exposición a varias formas de estrés. Nuestro objetivo fue establecer, si la expresión de las Hsp de los linfocitos del ganado bovino explotado en la región, muestra diferencias raciales en respuesta al estés calórico. De los bovinos, se obtuvieron linfocitos de sangre periférica por gradientes de Ficoll-histopaque, después se cultivaron en medio RPMI a temperaturas de 38, 40, 42 y 44°C, durante 4 horas, posteriormente se lisaron, se obtuvieron extractos que se caracterizaron por PAGE-SDS 10%, se transfirieron a papel de nitrocelulosa y se probaron con anticuerpos monoclonales anti-Hsp 25, 60,70 y 90. Se encontró: El calor es un agente estresante en los linfocitos del ganado bovino explotado en la región, cuya respuesta es a través de la expresión de la proteína Hsp70. La cantidad de proteína Hsp70 expresada en los linfocitos esta en función al estrés calórico recibido. El nivel de adaptación de un organismo ante un estresor, esta en función a la cantidad de Hsp expresada, por lo tanto, las razas bovinas más adaptadas a la región en orden decreciente es: Holstein-Australiano, Criollo, Pardo Suizo, Holstein y Limousin.

  9. Isolation of a Latimeria menadoensis heat shock protein 70 (Lmhsp70) that has all the features of an inducible gene and encodes a functional molecular chaperone.

    Science.gov (United States)

    Modisakeng, Keoagile W; Jiwaji, Meesbah; Pesce, Eva-Rachele; Robert, Jacques; Amemiya, Chris T; Dorrington, Rosemary A; Blatch, Gregory L

    2009-08-01

    Molecular chaperones facilitate the correct folding of other proteins, and heat shock proteins form one of the major classes of molecular chaperones. Heat shock protein 70 (Hsp70) has been extensively studied, and shown to be critically important for cellular protein homeostasis in almost all prokaryotic and eukaryotic systems studied to date. Since there have been very limited studies conducted on coelacanth chaperones, the main objective of this study was to genetically and biochemically characterize a coelacanth Hsp70. We have successfully isolated an Indonesian coelacanth (L. menadoensis) hsp70 gene, Lmhsp70, and found that it contained an intronless coding region and a potential upstream regulatory region. Lmhsp70 encoded a typical Hsp70 based on conserved structural and functional features, and the predicted upstream regulatory region was found to contain six potential promoter elements, and three potential heat shock elements (HSEs). The intronless nature of the coding region and the presence of HSEs suggested that Lmhsp70 was stress-inducible. Phylogenetic analyses provided further evidence that Lmhsp70 was probably inducible, and that it branched as a clade intermediate between bony fish and tetrapods. Recombinant LmHsp70 was successfully overproduced, purified and found to be functional using ATPase activity assays. Taken together, these data provide evidence for the first time that the coelacanth encodes a functional molecular chaperone system.

  10. 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 Effect of acute heat stress on hepatic and cerebral messenger RNA heat shock protein 70 and heat shock protein 70 level of broiler chicks from 2 to 5 days old of different strains

    Directory of Open Access Journals (Sweden)

    Nelson José Laurino Dionello

    2001-10-01

    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.Eighty broiler chicks, Naked neck (Na/na and Hubbard-Pettersen strains, were exposed to acute heat stress (36-37°C, from 2 to 5 days of age, and body weight and cloacal temperature were measured in the beginning and at the end of the heat stress period (5 h. Birds were sacrificed at the end and liver and brain samples were collected and analyzed using Western Blotting and Northern Blotting to quantify Hsp70 levels and mRNA Hsp70 transcript, respectively. The brain tissue had higher Hsp70 level than liver tissue. Before heat stress it was not observed difference in the Hsp70

  11. Chaperoning Proteins for Destruction: Diverse Roles of Hsp70 Chaperones and their Co-Chaperones in Targeting Misfolded Proteins to the Proteasome

    Directory of Open Access Journals (Sweden)

    Ayala Shiber

    2014-07-01

    Full Text Available Molecular chaperones were originally discovered as heat shock-induced proteins that facilitate proper folding of proteins with non-native conformations. While the function of chaperones in protein folding has been well documented over the last four decades, more recent studies have shown that chaperones are also necessary for the clearance of terminally misfolded proteins by the Ub-proteasome system. In this capacity, chaperones protect misfolded degradation substrates from spontaneous aggregation, facilitate their recognition by the Ub ligation machinery and finally shuttle the ubiquitylated substrates to the proteasome. The physiological importance of these functions is manifested by inefficient proteasomal degradation and the accumulation of protein aggregates during ageing or in certain neurodegenerative diseases, when chaperone levels decline. In this review, we focus on the diverse roles of stress-induced chaperones in targeting misfolded proteins to the proteasome and the consequences of their compromised activity. We further discuss the implications of these findings to the identification of new therapeutic targets for the treatment of amyloid diseases.

  12. Molecular Characterization of Vitellogenin and Its Receptor Genes from Citrus Red Mite, Panonychus citri (McGregor

    Directory of Open Access Journals (Sweden)

    Rui Zhong

    2015-03-01

    Full Text Available The production and uptake of yolk protein play an important role in the reproduction of all oviparous organisms. Vitellogenin (Vg is the precursor of vitellin (Vn, which is the major egg storage protein, and vitellogenin receptor (VgR is a necessary protein for the uptake of Vg into developing oocytes. In this paper, we characterize the full-length Vg and VgR, PcVg1 and PcVgR, respectively, of the citrus red mite Panonychus citri (McGregor. The PcVg1 cDNA is 5748 nucleotides (nt with a 5553-nt open reading frame (ORF coding for 1851 amino acids (aa, and the PcVgR is 6090 nt, containing an intact ORF of 5673 nt coding an expected protein of 1891 aa. The PcVg1 aa sequence shows a typical GLCG domain and several K/RXXR cleavage sites, and PcVgR comprises two ligand-binding domains, two epidermal growth factor (EGF-like regions containing YWTD motifs, a transmembrane domain, and a cytoplasmic domain. An analysis of the aa sequences and phylogenetics implied that both genes were genetically distinct from those of ticks and insects. The transcriptional profiles determined by real-time quantitative PCR in different developmental stages showed that both genes present the same expressional tendencies in eggs, larvae, nymphs, and adults. This suggested that the biosynthesis and uptake of PcVg occurs coordinately. The strong reproductive capacity of P. citri has been hypothesized as an important factor in its resistance; consequently, understanding the molecular mechanisms regulating Vg and VgR are fundamental for mite control.

  13. Molecular characterization of vitellogenin and its receptor genes from citrus red mite, Panonychus citri (McGregor).

    Science.gov (United States)

    Zhong, Rui; Ding, Tian-Bo; Niu, Jin-Zhi; Xia, Wen-Kai; Liao, Chong-Yu; Dou, Wei; Wang, Jin-Jun

    2015-01-01

    The production and uptake of yolk protein play an important role in the reproduction of all oviparous organisms. Vitellogenin (Vg) is the precursor of vitellin (Vn), which is the major egg storage protein, and vitellogenin receptor (VgR) is a necessary protein for the uptake of Vg into developing oocytes. In this paper, we characterize the full-length Vg and VgR, PcVg1 and PcVgR, respectively, of the citrus red mite Panonychus citri (McGregor). The PcVg1 cDNA is 5748 nucleotides (nt) with a 5553-nt open reading frame (ORF) coding for 1851 amino acids (aa), and the PcVgR is 6090 nt, containing an intact ORF of 5673 nt coding an expected protein of 1891 aa. The PcVg1 aa sequence shows a typical GLCG domain and several K/RXXR cleavage sites, and PcVgR comprises two ligand-binding domains, two epidermal growth factor (EGF)-like regions containing YWTD motifs, a transmembrane domain, and a cytoplasmic domain. An analysis of the aa sequences and phylogenetics implied that both genes were genetically distinct from those of ticks and insects. The transcriptional profiles determined by real-time quantitative PCR in different developmental stages showed that both genes present the same expressional tendencies in eggs, larvae, nymphs, and adults. This suggested that the biosynthesis and uptake of PcVg occurs coordinately. The strong reproductive capacity of P. citri has been hypothesized as an important factor in its resistance; consequently, understanding the molecular mechanisms regulating Vg and VgR are fundamental for mite control. PMID:25739087

  14. Citrus Genomics

    OpenAIRE

    Talon, Manuel; Gmitter, Fred G.Jr.

    2008-01-01

    Citrus is one of the most widespread fruit crops globally, with great economic and health value. It is among the most difficult plants to improve through traditional breeding approaches. Currently, there is risk of devastation by diseases threatening to limit production and future availability to the human population. As technologies rapidly advance in genomic science, they are quickly adapted to address the biological challenges of the citrus plant system and the world's industries. The hist...

  15. Study on the relationship between HSP70 expression and DNA damage in human embryonic lung(HEL)cells by benzo(a)pyrene metabolites%苯并(a)芘代谢产物作用下人胚肺细胞热应激蛋白70表达与DNA损伤

    Institute of Scientific and Technical Information of China (English)

    高雅娟; 陈胜; 肖成峰; 王瑞波; 贺涵贞; 邬堂春

    2000-01-01

    Objective To investigate the relationship between HSP70 expression and DNA damage in human embryonic lung(HEL) cells treated by benzo(a)pyrene(BaP) metabolites in vitro. Methods HEL cells of unheated group were stimulated by 0, 10,50,100,200 μ moL/L BaP for 3 h incubation(induced by S9-mix in vitro). HEL cells of heated group were treated at 41 ℃ for 1 h,recovered 2 h at 37 ℃ ,then were stimulated by BaP at the same conditions as the untreated group. The DNA damage and HSP70 expression were determined by single cell gel assay and Western blot respectively. Results DNA damage in HEL cells induced by BaP was detectable at 50 μmol/L in untreated group, while DNA damage in heated group was detectable at10 μmol/L. The level of HSP70 expression decreased,but DNA damage of HEL cells reduced at the highest dose(200 μmol/L BaP ) compared with the control. Compared with the untreated group, the number of cells with “+ +” level DNA damage reduced( P 0.05). Conclusion The roles of HSP70 in the protection from DNA damage induced by BaP are not significant, which may be related to the inhibition of HSP70 expression by BaP.%目的探讨苯并(a)芘(BaP)代谢产物作用下,人胚肺细胞HSP70表达与DNA损伤的关系。方法选用正常人胚肺(HEL)二倍体细胞,进行如下处理:一般处理组,以0、10、50、100、200μmol/L BaP染毒3 h(体外经大鼠肝S9-mix活化诱导),单细胞凝胶电泳技术检测DNA损伤情况;热应激组,细胞预热应激(41℃1 h,37℃2 h),再以同样浓度的BaP染毒3 h,采用Westem blot和单细胞凝胶电泳技术检测HSP70表达与DNA损伤情况。结果一般处理组BaP在50μmol/L时可引起HEL细胞DNA的明显损伤,且随BaP染毒剂量增加,DNA损伤级别加重;接受预热应激的细胞经不同浓度BaP染毒3 h后,与对照组比,HSP70表达水平降低,BaP在10 μmol/L时可见明显的DNA损伤,随染毒剂量的增加,DNA损伤级别

  16. 艾灸对亚急性衰老大鼠松果体细胞数及热休克蛋白70表达的影响%Effect of Moxibustion on Quantity of Pinealocytes and Pineal HSP 70 Expression in Subacute Aging Rats

    Institute of Scientific and Technical Information of China (English)

    梁欣; 钟愉

    2011-01-01

    Objective To study the mechanism of moxibustion in postponing the process of aging. Methods Thirty Wistar rats were equally randomized into control, model and moxibustion groups. The subacute aging model was established by hypodermic injection of 25% D-galactose (125 mg/kg). Moxibustion was applied to bilateral "Shenshu" (GV23) and "Pishu" (GV 20) once everyday for 6 weeks. After slicing, the pineal gland tissue was stained with HE and insitu hybridization methods respectively for detecting the quantity of pinealocytes and the expression of heat shock protein (HSP 70).Results Compared with the control group, both the quantity of pinealocytes and the expression of HSP 70 in the pineal gland in the model group were downregulated significantly (P<0. 001, P<0. 01). Compared with model group, the quantity of pinealocytes and of HSP 70 mRNA in the pineal gland of moxibustion group were upregulated significantly( P<0. 01, P<0. 05). Conclusion Moxibustion can suppress aging induced decrease of pinealocyte number and HSP 70 expression in subacute aging rats, which may contribute to its effect in postponing aging.%目的:探讨艾灸延缓衰老的可能作用机制.方法:将30只实验大鼠随机分为空白组、模型组、艾灸组,每组10只.皮下注射25%D-半乳糖溶液(125 mg/kg)造成大鼠衰老模型.艾灸组选取双侧"脾俞""肾俞"进行艾灸治疗,每日治疗1次,每次每穴灸5 min,连续治疗6周.采用组织化学染色及多克隆杭体免疫组化技术,结合细胞医学图像分析及病理图文图像分析方法,测定各组大鼠松果体细胞数及HSP 70表达.结果:与空白组相比,模型组大鼠松果体细胞数显著减少(P<0.001),HSP 70表达显著降低(P<0.01).与模型组相比,艾灸组大鼠松果体细胞数显著增多(P<0.01),HSP 70表达显著增加(P<0.05).结论:艾灸可能通过上调脑区松果体中HSP 70表达、减缓机体衰老过程中松果体细胞减少的速度,在一定程度上维持松果体的正

  17. 加味七方胃痛颗粒对慢性萎缩性胃炎大鼠胃蛋白酶含量及HSP70蛋白表达的影响%Effect of Modified Qifang Weitong Granules on Pepsin Content and Heat Shock Protein 70 Expression in Rats with Chronic Atrophic Gastritis

    Institute of Scientific and Technical Information of China (English)

    唐梅文; 王伟; 李桂贤; 岳小红; 陈国忠; 邓嫦

    2011-01-01

    Objective To investigate the effects of Modified Qifan Weitong Granules(QWG) on pepsin content and heat shock protein 70(HSP70) expression in the gastric mucosa of rats with chronic atrophic gastritis(CAG).Methods CAG rat models were established by means of N-methyl-N'-nitro-N-nitrosoguanidine(MNNG), and the effect of QWG on relative indexes was observed and compared with that of Weifuchun Granules. Results Compared with the model group, the atrophy and intestinal metaplasia of gastric mucosa were relieved in large-and medium-dosage QWG groups and in Weifuchun Granules group, and the pepsin content and HSP70 expression were increased in large-, medium-and small-dosage QWG groups and in Weifuchun Granules group, the difference being significant(P< 0.01). Compared with Weifuchun Granules group, pepsin content and HSP70 expression in large-dosage QWG group were increased with statistical significance(P < 0.05). Conclusion The mechanism of QWG in preventing and treating CAG may be related with prevention of gastric mucosa through increasing pepsin con tent and HSP70 expression.%目的 探讨加味七方胃痛颗粒对慢性萎缩性胃炎(CAC)模型大鼠胃黏膜组织中胃蛋白酶含量及热休克蛋白70(HSP70)表达的影响.方法 采用化学致癌物N-甲基-N'-硝基-N-亚硝基胍(MNNG)建立CAG动物模型,以胃复春为对照组,观察加味七方胃痛颗粒对大鼠CAG相关指标的作用.结果 与模型组比较,加味七方胃痛颗粒高、中剂量组和胃复春组的胃黏膜萎缩程度、胃黏膜肠化生程度明显减轻(P<0.01);加味七方胃痛颗粒高、中、低剂量组和胃复春组胃蛋白酶的含量、HSP70蛋白表达显著增加(P<0.01);与胃复春组比较,加味七方胃痛颗粒高剂量组胃蛋白酶的含量、HSP70蛋白表达显著增加(P<0.05).结论 加味七方胃痛颗粒防治CAG的作用机制可能是通过升高胃蛋白酶的含量及促进HSP70的表达,从而发挥对黏膜保护作用.

  18. Molecular Cloning and Identification of a Heat Shock Cognate Protein 70 Gene, Thhsc70, in Thellungiella halophila

    Institute of Scientific and Technical Information of China (English)

    ZHANGXia; GUOShan-Li; YINHai-Bo; XIONGDong-Jin; ZHANGHui; ZHAOYan-Xiu

    2004-01-01

    Heat shock cognate proteins 70 (hsp70s) act as molecular chaperones. Some hsp70s are also expressed in unstressed plants, known as hsc70. To gain further knowledge about the hsc70, the Thellungiella halophila hsc70 (Thhsc70) gene that encoded the cytosolic hsc70 in salt cress (T.halophila (C.A.Mey.) O.E.Schulz) was identified. In unstressed plants the expression of Thhsc70was shown to be tissue-specific. The Thhsc70 gene was induced by heat and cold stresses, but almost not by salt and drought stresses. Overexpression of Thhsc7Ocould increase thermctolerance and chilling tolerance in transgenic Arabidopsis plants.

  19. 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

  20. Evaluation of molecular chaperons Hsp72 and neuropeptide Y as characteristic markers of adaptogenic activity of plant extracts.

    Science.gov (United States)

    Asea, Alexzander; Kaur, Punit; Panossian, Alexander; Wikman, Karl Georg

    2013-11-15

    We have previously demonstrated that ADAPT-232, a fixed combination of adaptogenic substances derived from Eleutherococcus senticosus root extract, Schisandra chinensis berry extract, Rhodiola rosea root extract stimulated the expression and release of neuropeptide Y (NPY) and molecular chaperone Hsp72 from isolated human neurolgia cells. Both of these mediators of stress response are known to play an important role in regulation of neuroendocrine system and immune response. We further demonstrated that ADAPT-232 induced release of Hsp70 is mediated by NPY, suggesting an existence of NPY-mediated pathway of activation of Hsp72 release into the blood circulation system. The objective of this study was to determine whether this pathway is common for adaptogens and whether NPY and/or Hsp72 can be considered as necessary specific biomarkers for adaptogenic activity. The release of NPY and Hsp72 from neuroglia cells in response to treatment with various plant extracts (n=23) including selected validated adaptogens, partly validated adaptogens, claimed but negligibly validated adaptogens and some other plant extracts affecting neuroendocrine and immune systems but never considered as adaptogens was measured using high throughput ELISA techniques. We demonstrated that adaptogens, e.g. R. rosea, S. chinensis and E. senticosus stimulate both NPY and Hsp70 release from neuroblastoma cells, while tonics and stimulants have no significant effect on NPY in this in vitro test. In the groups of partly validated adaptogens the effect of Panax ginseng and Withania somnifera was not statistically significant both on NPY and Hsp70 release, while the activating effect of Bryonia alba and Rhaponticum cartamoides was significant only on Hsp70. In contrast, all tested non-adaptogens, such as antiinflammatoty plant extracts Matricaria recutita, Pelargonium sidoides, Hedera helix and Vitis vinifera significantly inhibit Hsp70 release and have no influence on NPY release from neuroblastoma

  1. A water-soluble high molecular weight substance isolated from Hyuganatsu orange (Citrus tamurana, suspected to be a polysaccharide, inhibits rat osteoclast cell formation

    Directory of Open Access Journals (Sweden)

    Hiroko Hata

    2015-06-01

    Full Text Available Background: Osteoporosis is detrimental to aged women’s health care. We previously reported that Hyuganatsu orange (Citrus Tamurana contains active substances that inhibit osteoclast activities. Prior to conducting a human study, we sought to identify the biological active substance in the Hyuganatsu orange which suppresses osteoclast formation. Methods: We isolated five fractions from a Hyuganatsu orange extract according to molecular weight. Each fraction was tested to determine its suppressive effect on the formation of osteoclasts in rats. We also used high-performance liquid chromatography (HPLC, infra-red (IR, and 1H and 13C NMR spectroscopy to evaluate its chemical structure. Data was recorded as mean ± standard error of the mean. The Mann-Whitney test was used, and a p-value of <.05 was considered statistically significant. Results: The highest and lowest molecular weight fractions showed significant suppression activity on rat osteoclast formation (p < .05. The lowest molecular weight fraction was identified as hesperidin using thin layer chromatography. Additionally, IR absorption revealed that the highest molecular weight fraction was not a flavonoid. With regard to chemical structure, 1H and 13C NMR spectroscopy suggested that the highest molecular weight fraction had signals compatible with a polysaccharide such as galacturonic acid. Conclusions: Hyuganatsu orange contains a biological active substance other than hesperidin that may be a polysaccharide and may suppress osteoclast formation.

  2. Molecular Characterization of Vitellogenin and Its Receptor Genes from Citrus Red Mite, Panonychus citri (McGregor)

    OpenAIRE

    Rui Zhong; Tian-Bo Ding; Jin-Zhi Niu; Wen-Kai Xia; Chong-Yu Liao; Wei Dou; Jin-Jun Wang

    2015-01-01

    The production and uptake of yolk protein play an important role in the reproduction of all oviparous organisms. Vitellogenin (Vg) is the precursor of vitellin (Vn), which is the major egg storage protein, and vitellogenin receptor (VgR) is a necessary protein for the uptake of Vg into developing oocytes. In this paper, we characterize the full-length Vg and VgR, PcVg1 and PcVgR, respectively, of the citrus red mite Panonychus citri (McGregor). The PcVg1 cDNA is 5748 nucleotides (nt) with a ...

  3. Citrus leprosis research update

    Science.gov (United States)

    Citrus leprosis is one of the oldest citrus diseases, but is also one of the most important emerging citrus diseases in South and Central America, and it is apparently spreading northward towards the U.S. Research in our labs and by others has shown that citrus leprosis disease is caused by a compl...

  4. Expression of HSP70 in rabbit dental pulp after cavity preparation with Er,Cr:YSGG laser%Er,Cr:YSGG激光备洞对兔牙髓组织热休克蛋白70表达的影响

    Institute of Scientific and Technical Information of China (English)

    杨超; 贾兴亚; 安欣; 王抒智; 钟鸣

    2012-01-01

    目的 观察Er,Cr:YSGG激光用于牙体窝洞制备后兔牙髓的形态学变化及热休克蛋白70 (heat shock protein70 HSP70)的表达情况.方法 用Er,Cr:YSGG激光在家兔前牙制备V类洞并常规充填,普通高速牙钻备洞作对照,分别于术后4h、24h处死实验动物,组织切片经HE染色及免疫组化染色,镜下观察.结果 Er,Cr:YSGG激光备洞后牙髓轻到中度炎症反应,与牙钻对照组无明显差异.备洞后24h,激光组成牙本质细胞HSP70中到强阳性表达,成纤维细胞、血管内皮细胞中度阳性表达.牙钻组成牙本质细胞及相邻成纤维细胞强阳性表达,血管内皮细胞强阳性表达,对照组HSP70表达高于Er,Cr:YSGG激光实验组,差别有统计学意义(P<0.05).结论 Er,Cr:YSGG激光备洞对牙髓刺激小,有很好的临床使用价值.%Objective To observe the dental pulpal morphological changes and the expression of HSP70 to cavity preparation in rabbit incisors by the Er,Cr:YSGG laser. Methods A total of thirtyt-wo incisors of eight healthy rabbits were divided into two groups: Er,Cr:YSGG laser and conventional bur group.after the cavities preparation by the laser and the conventional bur in rabbit incisors, the cavities, were filled by normal way .The teeth were extracted 4 and 24 hours after operation respectively, after HE and immunohistochemical staining, the tissue sections were observed by microscopy. Results The histological examination showed that there was no appreciable difference in the manner in which dental pulp tissue responds to cavity preparation with either Er,Cr:YSGG laser or a conventional bur. 24h after the operation, the Er,Cr:YSGG laser group have a moderate to strong expression of HSP70 in Odontoblasts,and moderate expression in Fibroblast and endothelial cells.the expression of HSP70 in the control group was higher than that in the Er, Cr:YSGG laser group (P<0.05). Conclusion Cavity preparation by Er,Cr:YSGG laser causes no noticeable pulpal damage

  5. Morphological and molecular characterization of a Hirsutella species infecting the Asian citrus psyllid, Diaphorina citri Kuwayama (Hemiptera: Psyllidae), in Florida.

    Science.gov (United States)

    Meyer, Jason M; Hoy, Marjorie A; Boucias, Drion G

    2007-06-01

    The Asian citrus psyllid, Diaphorina citri Kuwayama, is an invasive pest that vectors citrus greening disease, which recently was detected in Florida. Mycosed adult D. citri were collected at four sites in central Florida between September 2005 and February 2006. Observation of the cadavers using scanning electron microscopy revealed that the pathogen had branched synnemata supporting monophiladic conidiogenous cells. A high-fidelity polymerase chain reaction (PCR) assay was used to amplify the 18S rRNA, 28S rRNA and beta-tubulin genes of the pathogen for phylogenetic analysis. The morphological and genetic data indicated that the pathogen was a novel isolate related to Hirsutella citriformis Speare. PCR assays using isolate-specific primers designed from the unique putative intron region of the beta-tubulin sequence distinguished the psyllid pathogen from five related Hirsutella species. The pathogen was maintained in vivo by exposing healthy D. citri to the synnemata borne on field-collected cadavers. Infected psyllids had an abundance of septate hyphal bodies in their hemolymph and exhibited behavioral symptoms of disease. In vitro cultures of the pathogen were slow-growing and produced synnemata similar to those found on mycosed D. citri. In laboratory bioassays, high levels of mortality were observed in D. citri that were exposed to the conidia-bearing synnemata produced in vivo and in vitro.

  6. Effects of Cold Stress on Immune System and hsp70 mRNA Expression in Altay Sheep%寒冷应激对阿勒泰羊细胞免疫及热休克蛋白70 mRNA表达的影响

    Institute of Scientific and Technical Information of China (English)

    杨莉; 黄钰; 王凤丽; 刘静; 齐亚银; 张莉

    2015-01-01

    为研究不同的寒冷应激温度对阿勒泰羊(Ovis aries)细胞免疫及热休克蛋白70的影响.实验采集阿勒泰羊寒冷应激前后肝、肺、脾、淋巴结组织及血清,采用实时荧光定量PCR中最大二阶导数法(2-△△Ct)对各组织中hsp70的表达量进行统计学分析,同时采用ELISA方法测定血清中白细胞介素-4(IL-4)及白细胞介素-2(IL-2)寒冷应激前后浓度变化.结果显示,在寒冷条件下阿勒泰羊各种组织中热休克蛋白hsp70的表达都有所增加,尤其是脾组织的表达增加幅度较大.ELISA方法测定阿勒泰羊在寒冷刺激后IL-4浓度发生显著下降(P=0.016),而IL-2在冷应激后变化不显著(P=0.502),出现轻微的下调.研究表明,寒冷应激条件下,较高水平的hsp70能够保护机体免受应激的损伤.而在冷应激过程中,机体的免疫系统受到抑制.

  7. Molecular characterization of the heat shock protein 70 gene in Mycoplasma ovipneumoniae.

    Science.gov (United States)

    Zhang, Bin; Han, Xiao; Yue, Hua; Tang, Cheng

    2013-10-01

    Mycoplasma ovipneumoniae is a species of mycoplasma bacteria that commonly infects the respiratory tract, causing respiratory disease in sheep and goats worldwide. In the current study, the 70-kDa heat shock protein (Hsp70) gene was cloned, sequenced and analyzed in 14 clinical isolates of M. ovipneumoniae. Results showed that, compared to the reference Y98 strain, the open-reading frames (ORFs) of Hsp70 gene in all isolates were 1818 base pairs (bp). Three nucleotides of TCA were inserted at 1,776 bp, resulting in insertion of the amino acid glutamine at amino acid position 593. The neighbor-joining trees, constructed using the Hsp70 gene, exhibited that the closest genetic relationship occurred between M. ovipneumoniae and Mycoplasma hyopneumoniae, which was consistent with the one based on the whole genome comparisons between these two mycoplasma species. Therefore, these results suggest that the Hsp70 gene, rather than 16S ribosomal RNA, was suitable as a potential molecular marker for evaluating the genetic relationship of M. ovipneumoniae with other bacterial species.

  8. Citrus breeding, genetics and genomics in Japan.

    Science.gov (United States)

    Omura, Mitsuo; Shimada, Takehiko

    2016-01-01

    Citrus is one of the most cultivated fruits in the world, and satsuma mandarin (Citrus unshiu Marc.) is a major cultivated citrus in Japan. Many excellent cultivars derived from satsuma mandarin have been released through the improvement of mandarins using a conventional breeding method. The citrus breeding program is a lengthy process owing to the long juvenility, and it is predicted that marker-assisted selection (MAS) will overcome the obstacle and improve the efficiency of conventional breeding methods. To promote citrus molecular breeding in Japan, a genetic mapping was initiated in 1987, and the experimental tools and resources necessary for citrus functional genomics have been developed in relation to the physiological analysis of satsuma mandarin. In this paper, we review the progress of citrus breeding and genome researches in Japan and report the studies on genetic mapping, expression sequence tag cataloguing, and molecular characterization of breeding characteristics, mainly in terms of the metabolism of bio-functional substances as well as factors relating to, for example, fruit quality, disease resistance, polyembryony, and flowering.

  9. Citrus breeding, genetics and genomics in Japan.

    Science.gov (United States)

    Omura, Mitsuo; Shimada, Takehiko

    2016-01-01

    Citrus is one of the most cultivated fruits in the world, and satsuma mandarin (Citrus unshiu Marc.) is a major cultivated citrus in Japan. Many excellent cultivars derived from satsuma mandarin have been released through the improvement of mandarins using a conventional breeding method. The citrus breeding program is a lengthy process owing to the long juvenility, and it is predicted that marker-assisted selection (MAS) will overcome the obstacle and improve the efficiency of conventional breeding methods. To promote citrus molecular breeding in Japan, a genetic mapping was initiated in 1987, and the experimental tools and resources necessary for citrus functional genomics have been developed in relation to the physiological analysis of satsuma mandarin. In this paper, we review the progress of citrus breeding and genome researches in Japan and report the studies on genetic mapping, expression sequence tag cataloguing, and molecular characterization of breeding characteristics, mainly in terms of the metabolism of bio-functional substances as well as factors relating to, for example, fruit quality, disease resistance, polyembryony, and flowering. PMID:27069387

  10. Effect of temperature on growth and HSP70 mRNA expres-sion of thermo-tolerant brood of Apostichopus japonicus%温度选择对刺参群体在不同温度下生长及热休克蛋白表达的影响

    Institute of Scientific and Technical Information of China (English)

    刘广斌; 杨红生; 刘石林

    2010-01-01

    高温(26℃)培育刺参(Apostichopus japonicus)幼体获得选择群体, 生长至1.03 g±1.30 g, 与对照组幼参在18、20、22和24℃温度下培育.二者的成活率随着温度升高而降低, 而选择组幼参24℃的成活率显著高于对照群体(P<0.05).不同温度下的特定生长率之间没有差异.TRIzol法提取刺参总RNA,获得刺参HSP70片断基因序列, 利用实时定量PCR方法检测,选择群体HSP70mRNA的表达量在正常温度和热激条件下都较对照组高, 但表达量升高的倍数相近, 选择组为1.99倍, 对照组为2.07倍.

  11. 76 FR 23449 - Citrus Canker, Citrus Greening, and Asian Citrus Psyllid; Interstate Movement of Regulated...

    Science.gov (United States)

    2011-04-27

    ... to movement. Citrus canker is a plant disease that is caused by the bacterium Xanthomonas citri subsp... quarantined for citrus canker are found in Sec. 301.75-6. Citrus greening, also known as Huanglongbing disease of citrus, is considered to be one of the most serious citrus diseases in the world. Citrus...

  12. 78 FR 63369 - Citrus Canker, Citrus Greening, and Asian Citrus Psyllid; Interstate Movement of Regulated...

    Science.gov (United States)

    2013-10-24

    ... citrus canker are found in Sec. 301.75-6. Citrus greening, also known as Huanglongbing disease of citrus, is considered to be one of the most serious citrus diseases in the world. Citrus greening is a... areas of the world where the disease is endemic, citrus trees decline and die within a few years and...

  13. Progress on Dissecting and Controlling the Citrus Huanglongbing Complex

    OpenAIRE

    Duan, Yongping; Zhou, Lijuan; Zhang, Muqing; Benyon, Lesley; Armstrong-Vahling, Cheryl; Hoffman, Michele; Hao, Guixia; Zou, Huasong; Doud, Melissa; Ding, Fang; Morgan, Kent

    2014-01-01

    Citrus huanglongbing (HLB) is a century-old and emerging disease that impedes citrus production worldwide. ‘Candidatus Liberibacter asiaticus’ (Las) is the globally prevalent species of HLB bacteria. Here we describe our molecular characterizations of Las, and our newly-developed control methods for citrus HLB. From a genomics standpoint, we revealed Las has a significantly reduced genome (1.26Mb) and unique features adapted to its intracellular life style.  Although the genome is small, Las ...

  14. Molecular Characterization and Genetic Diversity Analysis of Sweet Orange Citrus sinensis L. Osbeck Cultivars in Iraq Using RAPD Markers

    Directory of Open Access Journals (Sweden)

    Ali Saeed Atiyah AL-Janabi

    2016-03-01

    Full Text Available Sweet orange (Citrus sinensis L. Osbeck is one of the most important commercially cultivated fruit crops of Citrus. Genetic diversity and inter-relationship among 5 cultivars (Indian, Iraqi, Japanese, Syrian, Egyptian of C. sinensis were analyzed based on RAPD markers. Six primers generated reproducible and easily storable RAPD profiles with a number of amplified DNA fragments ranging from 6 to 14 fragment bands. The total number of amplicons detected was 51, including 14 fragments unique bands with average reached 2.8 fragments ̸ primers. While the number of polymorphic ranged from 0 to 8 with an average reached 4.4 fragments ̸ primers with the polymorphic percentage ranged from 0% to 57.1%. While the number of monomorphic ranged from 2 to 5 fragment bands and was total of the monomorphic 15 fragments with an average reached 3 fragments ̸ primers with the monomorphic percentage was 14.2 % to 83.3%. A maximum numbers of amplicons was amplified with primer OPS-238 reached 14 fragments while the minimum number of fragments was amplified with primer OPS-253 reached 6 fragments. The highest number of polymorphic bands reached 8 fragments was obtained with primer OPS-238 with high percentage 57.1%, while the highest number of monomorphic bands reached 5 fragments with high percentage 83.3% was obtained with primer OPS-253. RAPD markers detected genetic distance and similarity, amaximum genetic distance value was observed between Japanese (Jap and Syrian (Syr cultivars reached 0.530 with less similarity value reached 47%, a minimum genetic distance value was observed between sweet Iraqi (Irq and Indian (Ind cultivars reached 0.239 with high similarity value reached 76.1%. The similarity matrices were employed in the cluster analysis to generate a dendrogram using the UPGMA method. The cluster tree analysis showed that the sweet orange cultivars were broadly divided into two main groups A and B with similarity reached 50%. A group including

  15. Multiple molecular penumbras after focal cerebral ischemia.

    Science.gov (United States)

    Sharp, F R; Lu, A; Tang, Y; Millhorn, D E

    2000-07-01

    Though the ischemic penumbra has been classically described on the basis of blood flow and physiologic parameters, a variety of ischemic penumbras can be described in molecular terms. Apoptosis-related genes induced after focal ischemia may contribute to cell death in the core and the selective cell death adjacent to an infarct. The HSP70 heat shock protein is induced in glia at the edges of an infarct and in neurons often at some distance from the infarct. HSP70 proteins are induced in cells in response to denatured proteins that occur as a result of temporary energy failure. Hypoxia-inducible factor (HIF) is also induced after focal ischemia in regions that can extend beyond the HSP70 induction. The region of HIF induction is proposed to represent the areas of decreased cerebral blood flow and decreased oxygen delivery. Immediate early genes are induced in cortex, hippocampus, thalamus, and other brain regions. These distant changes in gene expression occur because of ischemia-induced spreading depression or depolarization and could contribute to plastic changes in brain after stroke. PMID:10908035

  16. Tracking the interplay between bound peptide and the lid domain of DnaK, using molecular dynamics.

    Science.gov (United States)

    Azoulay, Itzhaq; Kucherenko, Nataly; Nachliel, Esther; Gutman, Menachem; Azem, Abdussalam; Tsfadia, Yossi

    2013-01-01

    Hsp70 chaperones consist of two functional domains: the 44 kDa Nucleotide Binding Domain (NBD), that binds and hydrolyses ATP, and the 26 kDa Substrate Binding Domain (SBD), which binds unfolded proteins and reactivates them, utilizing energy obtained from nucleotide hydrolysis. The structure of the SBD of the bacterial Hsp70, DnaK, consists of two sub-domains: A β-sandwich part containing the hydrophobic cavity to which the hepta-peptide NRLLLTG (NR) is bound, and a segment made of 5 α-helices, called the "lid" that caps the top of the β-sandwich domain. In the present study we used the Escherichia coli Hsp70, DnaK, as a model for Hsp70 proteins, focusing on its SBD domain, examining the changes in the lid conformation. We deliberately decoupled the NBD from the SBD, limiting the study to the structure of the SBD section, with an emphasis on the interaction between the charges of the peptide with the residues located in the lid. Molecular dynamics simulations of the complex revealed significant mobility within the lid structure; as the structure was released from the forces operating during the crystallization process, the two terminal helices established a contact with the positive charge at the tip of the peptide. This contact is manifested only in the presence of electrostatic attraction. The observed internal motions within the lid provide a molecular role for the function of this sub-domain during the reaction cycle of Hsp 70 chaperones. PMID:23774839

  17. Tracking the Interplay between Bound Peptide and the Lid Domain of DnaK, Using Molecular Dynamics

    Directory of Open Access Journals (Sweden)

    Yossi Tsfadia

    2013-06-01

    Full Text Available Hsp70 chaperones consist of two functional domains: the 44 kDa Nucleotide Binding Domain (NBD, that binds and hydrolyses ATP, and the 26 kDa Substrate Binding Domain (SBD, which binds unfolded proteins and reactivates them, utilizing energy obtained from nucleotide hydrolysis. The structure of the SBD of the bacterial Hsp70, DnaK, consists of two sub-domains: A β-sandwich part containing the hydrophobic cavity to which the hepta-peptide NRLLLTG (NR is bound, and a segment made of 5 α-helices, called the “lid” that caps the top of the β-sandwich domain. In the present study we used the Escherichia coli Hsp70, DnaK, as a model for Hsp70 proteins, focusing on its SBD domain, examining the changes in the lid conformation. We deliberately decoupled the NBD from the SBD, limiting the study to the structure of the SBD section, with an emphasis on the interaction between the charges of the peptide with the residues located in the lid. Molecular dynamics simulations of the complex revealed significant mobility within the lid structure; as the structure was released from the forces operating during the crystallization process, the two terminal helices established a contact with the positive charge at the tip of the peptide. This contact is manifested only in the presence of electrostatic attraction. The observed internal motions within the lid provide a molecular role for the function of this sub-domain during the reaction cycle of Hsp 70 chaperones.

  18. 跑台运动训练对脑缺血损伤大鼠热休克蛋白70及C-MYC表达的影响%Effects of treadmill training on the expression of HSP70 and C-MYC in the brains of rats with focal cerebral ischemia

    Institute of Scientific and Technical Information of China (English)

    刘德山; 刘楠; 张逸仙; 杜厚伟; 陈荣华

    2010-01-01

    目的 探讨跑台运动训练对局灶性脑缺血大鼠神经功能恢复和脑缺血组织中热休克蛋白70(HSP70)及C-MYC表达的影响.方法 将42只清洁级成年雄性SD大鼠分为假手术组6只、模型组18只、运动组18只.模型组、运动组大鼠采用改良的Longs线栓法制备大脑中动脉闭塞(MCAO)脑缺血模型,运动组于造模成功后24 h采用跑台训练器进行运动训练,每周6 d,共2周,其余2组则置于普通笼内饲养,期间可自由活动、进食.采用修正的神经行为学评分方法评价模型组和运动组大鼠MCAO脑缺血后第3,7,14天的神经功能,并断头取脑,采用逆转录-多聚酶链反应(RT-PCR)法、免疫组织化学和Western blot法检测脑缺血组织中HSP70、C-MYC的表达.结果 运动组大鼠脑缺血第7,14天神经功能评分均明显优于模型组(P<0.05或0.01);运动组大鼠脑缺血第7,14天HSF70和C-MYC表达较模型组增强(P<0.05或0.01).结论 运动训练可促进脑缺血大鼠神经功能恢复,其机制可能与上调脑缺血组织中HSP70及C-MYC表达有关.%Objective To observe the effects of treadmill training on the recovery of neurological function and the expression of HSP70 and C-MYC in the brains of rats with focal cerebral ischemia. Methods Forty-two male adult Sprague-Dawley rats were randomly divided into a sham group ( n =6), a model group (n =18) and a treadmill exercise group (n=18). Focal cerebral ischemia was induced by right middle cerebral artery occlusion (MCAO) in the model group and exercise group using a modified version of Longa's method. The rats in the treadmill exercise group were given treadmill training 6 d per week for 2 weeks after 24 h of MCAO. By contrast, the rats in the sham group and the model group were reared in standard cages. Before the rats were sacrificed at the 3rd, 7th and 14th d after MCAO, their neurological functions were tested using modified neurological severity scores ( mNSS) , and the mRNA and

  19. Bone marrow mesenchymal stem cells with support of ultrasound-mediated microbubbles and bispecific antibody prevent myocardial fibrosis via HSP-70%超声微泡联合双特异性抗体对骨髓间充质干细胞干预心肌纤维化的实验研究

    Institute of Scientific and Technical Information of China (English)

    邓玮; 陈庆伟; 王志刚; 李兴升; 刘灏; 周玥; 李桂琼; 柯大智; 莫显刚

    2011-01-01

    Objective To explore the effects of transplantated bone marrow mesenchymal stem cells (MSCs) on myocardial fibrosis with the aid of ultrasound-mediated microbubbles (MB) and bispecific antibody(BiAb) combination.Methods With the aid of MB,isolated MSCs from male mice and the BiAb were transfused into female mice with isoproterenol-induced myocardial fibrosis via tail vein (MSCs + BiAb + MB group).BiAb was producted with anti-CD29 which can recognize MSCs and anti-myosin light chain antibody (AMLCA) which can specifically bind to injured myocardium.There were six groups investigated:MSC + BiAb + MB,MSC,BiAb,MB,MSC + BiAb,untreated,and control groups.Five weeks after treatment administration,the expressions of sex-determining region of Y-chromosome (SRY) in myocardium were detected by fluorescent quantitative PCR.The distribution of collagen was observed by sirius red staining.Heat shock protein-70 (HSP-70) in myocardium was detected by immunohistochemistry.Results The highest homing number of MSCs was in the MSCs + BiAb + MB group,MSCs + BiAb group ranked the second,and lowest in MSCs group.Compared to the untreated group,the MSCs + BiAb + MB,MSCs + BiAb and MSCs groups had less collagen deposition (P <0.05),and decreased level of HSP-70.Compared to those of the MSCs group,the collagen deposition were decreased in MSCs + BiAb + MB group (P <0.05).Conclusions MB and BiAb can promote the homing number of MSCs in mice.MB can further the homing rate and the repairing efficacy of MSCs.The homing MSCs can prevent the process of myocardial fibrosis.And HSP-70 was involved in the internal mechanism.%目的 探讨骨髓间充质干细胞(MSCs)在超声辐照微泡(MB)和双特异抗体(BiAb)的辅助下干预心肌纤维化的效果及可能机制.方法 在MB的辅助下,将雄性小鼠MSCs与BiAb输入异丙肾性心肌纤维化雌性小鼠(MSCs+BiAb+MB组).BiAb由能识别间充质干细胞的anti-CD29和能特异性结合损伤心肌的抗肌凝蛋白轻链抗

  20. Effects of Ultra-filtration Extract from Angelicae Sinensis Radix and Hedysarum Polybotrys on Expressions of HSP70 and eNOS in H2O2-induced Endothelial Cell Apoptosis%当归红芪超滤物对过氧化氢致内皮细胞凋亡中热休克蛋白70及内皮型一氧化氮合酶表达的影响

    Institute of Scientific and Technical Information of China (English)

    顾丽娟; 刘凯; 孙少伯; 刘国安; 李应东

    2015-01-01

    Objective To investigate the effects of ultra-filtration extract from the mixture of Angelicae Sinensis Radix and Hedysarum Polybotrys (UFE-AH) on the expressions of HSP70 and eNOS in H2O2-induced endothelial cell apoptosis. Methods H2O2 induced ECV-304 cell apoptosis to prepare models. The experiment was divided into normal control group, model group, simple medicine group, medicine intervention group, and all treatment groups received relevant medicine for intervention. Flow cytometry (FCM) was used to detect apoptosis and concentration of intracellular Ca2+;RT-PCR was used to detect the mRNA expression of HSP70 and eNOS;Western blot was used to detect the expression of HSP70 protein;Nitrale reduetase and spectrophotometric method were employed to detect the content of NO. Results Compared with normal control group, cell apoptosis rate, concentration of intracellular Ca2+, and expression of HSP70 increased significantly in model group (P<0.05);gene expression of eNOS mRNA and content of NO decreased (P<0.05). Compared with model group, cell apoptosis rate and concentration of intracellular Ca2+dropped in medicine intervention group (P<0.05);expressions of HSP70, eNOS mRNA and content of NO increased (P<0.05). Conclusion UFE-AH can confront H2O2-induced cell apoptosis H2O2 of ECV-304 human umbilical vein endothelial by increasing the expressions of HSP70 and eNOS and content of NO, and reducing the intracellular calcium overload.%目的:探讨当归红芪超滤物对过氧化氢(H2O2)致内皮细胞凋亡中热休克蛋白70(HSP70)及内皮型一氧化氮合酶(eNOS)表达的影响。方法 H2O2诱导 ECV-304人脐静脉内皮细胞凋亡制备模型,实验分为正常对照组、模型组、单纯药物组、药物干预组,并给予相应药物干预,流式细胞仪检测细胞凋亡及细胞内 Ca2+浓度,RT-PCR法检测HSP70、eNOS的基因表达,蛋白印迹法检测HSP70的蛋白表达,硝酸还原酶法及分光光度法检测一氧化氮(NO)的含量。

  1. A Rational Design Strategy for the Selective Activity Enhancement of a Molecular Chaperone toward a Target Substrate.

    Science.gov (United States)

    Aprile, Francesco A; Sormanni, Pietro; Vendruscolo, Michele

    2015-08-18

    Molecular chaperones facilitate the folding and assembly of proteins and inhibit their aberrant aggregation. They thus offer several opportunities for biomedical and biotechnological applications, as for example they can often prevent protein aggregation more effectively than other therapeutic molecules, including small molecules and antibodies. Here we present a method of designing molecular chaperones with enhanced activity against specific amyloidogenic substrates while leaving unaltered their functions toward other substrates. The method consists of grafting onto a molecular chaperone a peptide designed to bind specifically an epitope in the target substrate. We illustrate this strategy by describing Hsp70 variants with increased affinities for α-synuclein and Aβ42 but otherwise unaltered affinities for other substrates. These designed variants inhibit protein aggregation and disaggregate preformed fibrils significantly more effectively than wild-type Hsp70 indicating that the strategy presented here provides a possible route for tailoring rationally molecular chaperones for specific purposes.

  2. 76 FR 8603 - Citrus Seed Imports; Citrus Greening and Citrus Variegated Chlorosis

    Science.gov (United States)

    2011-02-15

    ... disease of citrus and referred to below as HLB) is considered to be one of the most serious citrus... world where the disease is endemic, citrus trees decline and die within a few years and may never.... Manjunath. Asian Citrus Psyllids (Sternorrhyncha: Psyllidae) and Greening Disease of Citrus: A...

  3. 糖尿病周围神经病变与NF-κB、TNF-α和HSP70的关系%Association of Diabetic Peripheral Neuropathy with NF-κB, TNF-αand the Heat Shock Protein 70

    Institute of Scientific and Technical Information of China (English)

    刘肖梅; 李才锐; 孙曙光

    2016-01-01

    糖尿病周围神经病变(DPN)是一种常见的糖尿病并发症,主要发病机制与高血糖引起的代谢紊乱、氧化应激、神经营养障碍及遗传因素等有关。氧化应激可以通过包括多元醇途径、己糖胺途径、蛋白激酶C途径的激活以及糖基化终末产物(AGES)的形成等代谢途径间接引起神经元和轴突的损伤,同时直接损伤了神经细胞。其中NF-κB、TNF-α和HSP70在糖尿病周围神经病变的氧化应激的过程中扮演了不同角色。%Diabetic peripheral neuropathy (DPN) is one of the common complications of diabetes, its pathogenesis shows a rela-tionship with some reasons,such as metabolic disorders, oxidative stress, nerve nutrition disorder, vascular injury and genetic factors, which caused by high blood sμgar. Oxidative stress can be achieved by some metabolic pathways, including polyol pathway, hexose amine way, the activation of protein kinase C pathway and the formation of advanced glycation end products(AGES), to indirect cause damage to neurons and axons.And they also can damage to nerve cells directly. The NF-κB, TNF-αand HSP70 play different roles in oxidative stress of diabetic peripheral neuropathy different links.

  4. 五味子乙素诱导的HSP27和HSP70对Con A诱导小鼠肝损伤的保护作用%Protective effect of Schisandrin B against Con A-induced liver injury in mice through inducing heat shock protein 27/70

    Institute of Scientific and Technical Information of China (English)

    李丽波; 王玉祥; 杨宏艳; 辛亚兵; 高涵

    2013-01-01

    目的 探讨五味子乙素(Sch B)对刀豆蛋白A(Concanavalin A,Con A)诱导小鼠肝损伤的保护作用与HSP27和HSP70的关系.方法 采用静脉注射Con A诱导小鼠免疫性肝损伤模型,Sch B和热休克蛋白抑制剂槲皮素(Quercetin)灌胃给药.取4周龄雄性ICR小鼠72只,体质量18 ~22 g,按体质量分为6组:正常对照组、模型组(Con A)、Sch B+Con A组、Sch B+Con A+Quercetin组、Con A+Quercetin组和Quercetin组.Western blot法检测小鼠肝脏HSP27和HSP70蛋白的表达;实时荧光定量PCR法检测小鼠肝脏HSP27和HSP70 mRNA的表达;光镜下观察肝脏组织病理学改变;试剂盒检测小鼠血清转氨酶水平.结果 与ConA组相比,Sch B+Con A组小鼠肝脏HSP27和HSP70在蛋白水平和mRNA水平的表达均显著增加,同时肝细胞坏死程度明显减轻、小鼠血清转氨酶水平显著降低.与Sch B+Con A组相比,Sch B+Con A+Quercetin组小鼠肝脏HSP27和HSP70在蛋白水平和mRNA水平的表达均明显降低,同时小鼠肝细胞坏死程度明显增加、血清转氨酶水平显著增高.而与正常对照组相比,Quercetin组小鼠的上述各项指标未见改变.结论 SchB可通过诱导小鼠肝脏HSP27和HSP70的表达保护Con A所致的小鼠免疫性肝损伤.%Objective To determine the protective effect of Schisandrin B (Sch B) against Concanavalin A (Con A)-induced liver injury in mice and the roles of heat shock protein (HSP) 27/70 in the hepatic protection.Methods Seventy-two four-week-old male ICR mice were divided into 6 groups (n =12 for each group),that is,control group,model group,Sch B + Con A group,Sch B + Con A + Quercetin group,Con A + Quercetin group and Quercetin group.Sch B (200 mg/kg) and the heat shock protein inhibitor,Quercetin (200 mg/kg) were administered intragastrically,alone or at the same time for the corresponding mice.Mouse model of liver injury was induced by intravenous injection of 25 mg/kg Con A in 1 h after last intra gastrical

  5. PHARMACOLOGICAL IMPORTANCE OF CITRUS FRUITS

    OpenAIRE

    Amita Tomar *, Mridula Mall and Pragya Rai

    2013-01-01

    This paper reviews the pharmacological importance of citrus fruits. Citrus fruits are used for various pharmacological importance. According to literature the citrus fruit possess anti-cancer, antimicrobial, antioxidant, antiulcer, anti-inflammatory, and hypolipidemic and hepatoprotective properties.

  6. Molecular characterization of SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPL gene family from Citrus and the effect of fruit load on their expression

    Directory of Open Access Journals (Sweden)

    Liron eShalom

    2015-05-01

    Full Text Available We recently identified a Citrus gene encoding SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPL transcription factor that contained a sequence complementary to miR156. Genes of the SPL family are known to play a role in flowering regulation and phase transition. In Citrus, the mRNA levels of the gene were significantly altered by fruit load in buds; under heavy fruit load (ON-Crop trees, known to suppress next year flowering, the mRNA levels were down-regulated, while fruit removal (de-fruiting, inducing next-year flowering, resulted in its up-regulation. In the current work, we set on to study the function of the gene. We showed that the Citrus SPL was able promote flowering independently of photoperiod in Arabidopsis, while miR156 repressed its flowering-promoting activity. In order to find out if fruit load affected the expression of additional genes of the SPL family, we identified and classified all SPL members in the Citrus genome, and studied their seasonal expression patterns in buds and leaves, and in response to de-fruiting. Results showed that two additional SPL-like genes and miR172, known to be induced by SPLs in Arabidopsis, were altered by fruit load. The relationships between these factors in relation to the fruit-load effect on Citrus flowering are discussed.

  7. Molecular characterization of SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPL) gene family from Citrus and the effect of fruit load on their expression.

    Science.gov (United States)

    Shalom, Liron; Shlizerman, Lyudmila; Zur, Naftali; Doron-Faigenboim, Adi; Blumwald, Eduardo; Sadka, Avi

    2015-01-01

    We recently identified a Citrus gene encoding SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPL) transcription factor that contained a sequence complementary to miR156. Genes of the SPL family are known to play a role in flowering regulation and phase transition. In Citrus, the mRNA levels of the gene were significantly altered by fruit load in buds; under heavy fruit load (ON-Crop trees), known to suppress next year flowering, the mRNA levels were down-regulated, while fruit removal (de-fruiting), inducing next-year flowering, resulted in its up-regulation. In the current work, we set on to study the function of the gene. We showed that the Citrus SPL was able promote flowering independently of photoperiod in Arabidopsis, while miR156 repressed its flowering-promoting activity. In order to find out if fruit load affected the expression of additional genes of the SPL family, we identified and classified all SPL members in the Citrus genome, and studied their seasonal expression patterns in buds and leaves, and in response to de-fruiting. Results showed that two additional SPL-like genes and miR172, known to be induced by SPLs in Arabidopsis, were altered by fruit load. The relationships between these factors in relation to the fruit-load effect on Citrus flowering are discussed. PMID:26074947

  8. Citrus Waste Biomass Program

    Energy Technology Data Exchange (ETDEWEB)

    Karel Grohman; Scott Stevenson

    2007-01-30

    Renewable Spirits is developing an innovative pilot plant bio-refinery to establish the commercial viability of ehtanol production utilizing a processing waste from citrus juice production. A novel process based on enzymatic hydrolysis of citrus processing waste and fermentation of resulting sugars to ethanol by yeasts was successfully developed in collaboration with a CRADA partner, USDA/ARS Citrus and Subtropical Products Laboratory. The process was also successfully scaled up from laboratory scale to 10,000 gal fermentor level.

  9. Molecular characterization of SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPL) gene family from Citrus and the effect of fruit load on their expression

    OpenAIRE

    Shalom, Liron; Shlizerman, Lyudmila; Zur, Naftali; Doron-Faigenboim, Adi; Blumwald, Eduardo; Sadka, Avi

    2015-01-01

    We recently identified a Citrus gene encoding SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPL) transcription factor that contained a sequence complementary to miR156. Genes of the SPL family are known to play a role in flowering regulation and phase transition. In Citrus, the mRNA levels of the gene were significantly altered by fruit load in buds; under heavy fruit load (ON-Crop trees), known to suppress next year flowering, the mRNA levels were down-regulated, while fruit removal (de-fruiting),...

  10. Founder lines for improved citrus biotechnology

    Science.gov (United States)

    This article discusses the research needed to develop the RMCE strategy and molecular assays for site-specific recombinases as tools for genome manipulation. Explanation of genetic engineering used to generate transgenic citrus plants to exhibit a novel phenotype, but not to contain the recombinase...

  11. Cercosporoid diseases of Citrus.

    Science.gov (United States)

    Huang, Feng; Groenewald, J Z; Zhu, Li; Crous, P W; Li, Hongye

    2015-01-01

    Citrus leaves and fruits exhibiting disease symptoms ranging from greasy spot, yellow spot, small or large brown spot, black dot, and brown dot were sampled from Fujian, Guangdong, Guizhou, Hunan, Jiangxi, Yunnan, Zhejiang provinces and the Guangxi Zhuang Autonomous Region in China. In total 82 isolates representing various cercosporoid genera were isolated from these disease symptoms, which were supplemented with eight Citrus cercosporoid isolates collected from other countries. Based on a morphological and phylogenetic study using sequence data from the nuclear ribosomal DNA's ITS1-5.8S-ITS2 regions (ITS), and partial actin (act), β-tubulin (tub2), 28S nuclear ribosomal RNA (28S rDNA) and translation elongation factor 1-α (tef1) genes, these strains were placed in the following genera: Cercospora, Pallidocercospora, Passalora, Pseudocercospora, Verrucisporota and Zasmidium. All isolates tended to be sterile, except the Zasmidium isolates associated with citrus greasy spot-like symptoms, which subsequently were compared with phylogenetically similar isolates occurring on Citrus and other hosts elsewhere. From these results four Zasmidium species were recognized on Citrus, namely Z. indonesianum on Citrus in Indonesia, Z. fructicola and Z. fructigenum on Citrus in China and Z. citri-griseum, which appears to have a wide host range including Acacia, Citrus, Eucalyptus and Musa, as well as a global distribution.

  12. 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 [Corrigendum

    OpenAIRE

    Ibrahim, Mohamed Yousif

    2015-01-01

    Ibrahim MY, Hashim NM, Mohan S, et al. Drug Des Devel Ther. 2014;8:2193–2211. On page 2193, author affiliations, “Department of Molecular Medicine, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia” should be “Department of Biomedical Science, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia”. Introduction, first paragraph, the text should read: “Breast cancer has become a major cause of morb...

  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

    OpenAIRE

    Ibrahim, Mohamed Yousif

    2014-01-01

    Mohamed Yousif Ibrahim,1 Najihah Mohd Hashim,1 Syam Mohan,2 Mahmood Ameen Abdulla,3 Siddig Ibrahim Abdelwahab,2 Behnam Kamalidehghan,1 Mostafa Ghaderian,1,4 Firouzeh Dehghan,1,5 Landa Zeenelabdin Ali,1 Hamed Karimian,1 Maizatulakmal Yahayu,6 Gwendoline Cheng Lian Ee,7 Abdoreza Soleimani Farjam,8 Hapipah Mohd Ali9 1Department of Pharmacy, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia; 2Medical Research Centre, Jazan University, Jazan, Saudi Arabia; 3Department of Molecular ...

  14. Consumption of vitamin B6 reduces colonic damage and protein expression of HSP70 and HO-1, the anti-tumor targets, in rats exposed to 1,2-dimethylhydrazine

    OpenAIRE

    KAYASHIMA, TOMOKO; Tanaka, Kenta; OKAZAKI, YUKAKO; Matsubara, Kiminori; Yanaka, Noriyuki; Kato, Norihisa

    2011-01-01

    Mounting evidence indicates that vitamin B6 is a protective factor for colon cancer. Elevations in colonic damage, cell proliferation and heat shock proteins (HSPs, molecular chaperones) have been suggested to be associated with colon carcinogenesis. This study was performed to examine the effect of dietary levels of vitamin B6 (1, 7 or 35 mg pyridoxine HCl/kg diet) for 22 weeks on colon damage, epithelial cell proliferation and expression of HSPs in rats exposed to 1,2-dimethylhydrazine (DMH...

  15. 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

    OpenAIRE

    Ibrahim MY; Hashim NM; Mohan S; Abdulla MA; Kamalidehghan B; Ghaderian M; Dehghan F; Ali LZ; Arbab IA; Yahayu M; Lian GE; Ahmadipour F; Ali HM

    2014-01-01

    Mohamed Yousif Ibrahim,1 Najihah Mohd Hashim,1 Syam Mohan,2 Mahmood Ameen Abdulla,3 Behnam Kamalidehghan,1 Mostafa Ghaderian,1,4 Firouzeh Dehghan,1,5 Landa Zeenelabdin Ali,1 Ismail Adam Arbab,6 Maizatulakmal Yahayu,7 Gwendoline Ee Cheng Lian,8 Fatemeh Ahmadipour,1 Hapipah Mohd Ali9 1Department of Pharmacy, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia; 2Medical Research Centre, Jazan University, Jazan, Saudi Arabia; 3Department of Molecular Medicine, Faculty of Medicine Un...

  16. 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

    OpenAIRE

    Ibrahim, Mohamed Yousif

    2014-01-01

    Mohamed Yousif Ibrahim,1 Najihah Mohd Hashim,1 Syam Mohan,2 Mahmood Ameen Abdulla,3 Behnam Kamalidehghan,1 Mostafa Ghaderian,1,4 Firouzeh Dehghan,1,5 Landa Zeenelabdin Ali,1 Ismail Adam Arbab,6 Maizatulakmal Yahayu,7 Gwendoline Ee Cheng Lian,8 Fatemeh Ahmadipour,1 Hapipah Mohd Ali9 1Department of Pharmacy, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia; 2Medical Research Centre, Jazan University, Jazan, Saudi Arabia; 3Department of Molecular Medicine, Faculty of Medicine Un...

  17. Fuel from citrus waste

    Energy Technology Data Exchange (ETDEWEB)

    1979-08-01

    University of Florida researchers are looking for methods of using citrus peel to produce fuel for the citrus industry by using the methane gas given off in a special continuous digestion vessel. They are trying to discover ways to reduce the peel oil content of oranges as this oil inhibits the growth of organisms which produce the methane. The citrus industry currently utilizes orange peel and pulp to make cattle feed, but since the peel has to be dried this step consumes a lot of energy.

  18. 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 [Corrigendum

    Directory of Open Access Journals (Sweden)

    Ibrahim MY

    2015-06-01

    Full Text Available Ibrahim MY, Hashim NM, Mohan S, et al. Drug Des Devel Ther. 2014;8:2193–2211. On page 2193, author affiliations, “Department of Molecular Medicine, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia” should be “Department of Biomedical Science, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia”. Introduction, first paragraph, the text should read: “Breast cancer has become a major cause of morbidity and mortality in women globally. The American Cancer Society (ACS reported that breast cancer incidence has an estimation of 26% of all new cancer cases, which is the highest in ratio among all the cancers in American women.1 The National Cancer Registry (NCR in Malaysia has reported that one in twenty Malaysian women are at a risk of acquiring breast in their lifetime.2 The incidence rate in Malaysia is still considered low if compared to Europe and United States.3 Up to 70% of breast cancer development causes occur in women is reported to be of environmental factors and lifestyle.4,5”  Introduction, second paragraph, first sentence, the text should read: “Radiation therapy has become a valuable tool among cancer treatment strategies for the control of local and regional diseases after 1960 with the invention of the linear accelerator, but, like surgery, radiation therapy alone cannot enucleate metastatic cancer.”  Discussion, first paragraph, first sentence, the text should read: “Apoptosis has a vital role in many functions, ranging from fetal development to adult tissue homeostasis.32”   Read the original article

  19. Citrus diseases with global ramifications including citrus canker and huanglongbing

    Science.gov (United States)

    Although there are a number of diseases that plague citrus production worldwide, two bacterial diseases are particularly problematic. Both are of Asian origin and currently cause severe economic damage: Asiatic citrus canker (ACC) and citrus huanglongbing (HLB). Although ACC has been found in the ...

  20. Phylogenetic Analysis of Citrus tristeza virus Isolates of Wild Type Citrus in China

    Institute of Scientific and Technical Information of China (English)

    YI Long; ZHOU Chang-yong

    2014-01-01

    The genetic variation and phylogenetic relationships of Citrus tristeza virus (CTV) isolates collected from Chinese wild type citrus were analyzed by comparing the sequences of nine genomic regions (p23, p20, p13, p18, p25, p27, POL, HEL and k17) with the CTV isolates of cultivated citrus from different countries. The results showed that the divergence pattern of genomic RNA of the CTV isolates from wild type citrus was similar to that of other isolates from cultivated citrus, the 3´ proximal region was relatively conserved, and the 5´ proximal region had greater variability. The nine genomic regions of CTV isolates analyzed were found to have been under purifying selection in the evolution process. Phylogenetic analysis showed that the eleven Chinese wild CTV isolates were located at different clades and did not relfect their geographical origins, suggesting genetic diversity among the Chinese wild CTV populations. These results will aid in the understanding of molecular evolution of the Chinese CTV populations.

  1. PHARMACOLOGICAL IMPORTANCE OF CITRUS FRUITS

    Directory of Open Access Journals (Sweden)

    Amita Tomar *, Mridula Mall and Pragya Rai

    2013-01-01

    Full Text Available This paper reviews the pharmacological importance of citrus fruits. Citrus fruits are used for various pharmacological importance. According to literature the citrus fruit possess anti-cancer, antimicrobial, antioxidant, antiulcer, anti-inflammatory, and hypolipidemic and hepatoprotective properties.

  2. High molecular diversity of the fungus Guignardia citricarpa and Guignardia mangiferae and new primers for the diagnosis of the citrus black spot

    Directory of Open Access Journals (Sweden)

    Danyelle Stringari

    2009-10-01

    Full Text Available RAPD markers were used to investigate the d