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1.
Glucocorticoid hormone receptor exists in the cytoplasm of target cells in the form of dynamic multiprotein heterocomplexes with heat shock proteins Hsp90 and Hsp70, and additional components of the molecular chaperone machinery. Whole body hyperthermic stress was previously shown to induce alterations in protein composition of these complexes increasing the share of Hsp70, but participation of individual Hsp70 family members was not investigated. In the present study the association of glucocorticoid receptor with constitutive and inducible forms of Hsp70 in the liver cytosol of rats exposed to 41 degrees C whole body hyperthermic stress was examined. Immunoprecipitation of glucocorticoid receptor heterocomplexes by monoclonal anti-receptor antibody (BuGR2) followed by quantitative immunoblotting revealed the presence of both nucleocytoplasmic Hsp70 family members, constitutive--Hsc70 and inducible--Hsp72, within the complexes. Immediately after the stress only Hsc70 was found in association with glucocorticoid receptor. However, after the induction of Hsp72 by stress, its appearance within the glucocorticoid receptor heterocomplexes was also recorded and the presence of both Hsp70 forms within the heterocomplexes was evident by the end of examined 24h period after the stress. This study confirms that heat stress affects protein composition of rat liver glucocorticoid receptor heterocomplexes increasing the share of Hsp70 and shows that this increase could be equally ascribed to constitutive and inducible forms of Hsp70. 相似文献
2.
The influence of cadmium on the rat liver glucocorticoid receptor (GR) binding capacity, on the cytosolic level of 90 kDa
heat shock protein (Hsp90), and on the association of the two proteins was investigated. The results showed that the mode
of metal application led to diverse alterations in hormone binding to the GR. Reduction of the GR binding capacity observed
afterin vitro treatment was proportional to the applied metal concentrations. In animals administered different doses of cadmium, GR binding
capacity was not reduced, except in those that received the highest dose. A concomitant elevation of Hsp90 level was detected
both in the cytosol and within the GR untransformed heterocomplexes. The results suggest that cadmium-induced reduction of
the GR binding capacity seenin vitro was prevented in intact animals by the elevated level of Hsp90 within the GR heterocomplexes.
This revised version was published online in July 2006 with corrections to the Cover Date. 相似文献
3.
The level and phosphorylation of Hsp70 in the rat liver cytosol after adrenalectomy and hyperthermia
Hepatic heat shock protein Hsp70 synthesis and in vitro phosphorylation were studied in the liver cytosol of intact, adrenalectomized and dexamethasone-administered adrenalectomized rats after 41 degrees C whole body hyperthermic stress. Hsp70 was detected by immunoblotting with N27F3-4 monoclonal antibody recognizing both constitutive and inducible forms of the protein. A comparison between basal and heat stress-induced levels of the protein in the liver cytosol of the three groups of animals suggested that glucocorticoid hormones stimulate the basal synthesis of Hsp70 and inhibit its induction by stress. In both unstressed and hyperthermia-exposed animals, hepatic Hsp70 was detected as a phosphoprotein. The extent of its in vitro phosphorylation was found to be significantly reduced by heat stress or adrenalectomy, but dexamethasone failed to restore it to the original level. 相似文献
4.
Cell Stress & Chaperones journal has become a major outlet for papers and review articles about anti-heat shock protein (HSP) antibodies. In the last
decade, it became evident that apart from their intracellular localization, members of the heat shock protein 90 (Hsp90; HSPC)
and Hsp70 (HSPA) family are also found on the cell surface. In this review, we will focus on Hsp70 (HSPA1A), the major stress-inducible
member of the human Hsp70 family. Depending on the cell type, the membrane association of Hsp70 comes in two forms. In tumor
cells, Hsp70 appears to be integrated within the plasma membrane, whereas in non-malignantly transformed (herein termed normal)
cells, Hsp70 is associated with cell surface receptors. This observation raises the question whether or not these two surface
forms of Hsp70 in tumor and normal cells can be distinguished using Hsp70 specific antibodies. Presently a number of Hsp70
specific antibodies are commercially available. These antibodies were generated by immunizing mice either with recombinant
or HeLa-derived human Hsp70 protein, parts of the Hsp70 protein, or with synthetic peptides. This review aims to characterize
the binding of different anti-human Hsp70 antibodies and their capacity to distinguish between integrated and receptor-bound
Hsp70 in tumor and normal cells. 相似文献
5.
The co-chaperone CHIP regulates protein triage decisions mediated by heat-shock proteins 总被引:1,自引:0,他引:1
Connell P Ballinger CA Jiang J Wu Y Thompson LJ Höhfeld J Patterson C 《Nature cell biology》2001,3(1):93-96
To maintain quality control in cells, mechanisms distinguish among improperly folded peptides, mature and functional proteins, and proteins to be targeted for degradation. The molecular chaperones, including heat-shock protein Hsp90, have the ability to recognize misfolded proteins and assist in their conversion to a functional conformation. Disruption of Hsp90 heterocomplexes by the Hsp90 inhibitor geldanamycin leads to substrate degradation through the ubiquitin-proteasome pathway, implicating this system in protein triage decisions. We previously identified CHIP (carboxyl terminus of Hsc70-interacting protein) to be an interaction partner of Hsc70 (ref. 4). CHIP also interacts directly with a tetratricopeptide repeat acceptor site of Hsp90, incorporating into Hsp90 heterocomplexes and eliciting release of the regulatory cofactor p23. Here we show that CHIP abolishes the steroid-binding activity and transactivation potential of the glucocorticoid receptor, a well-characterized Hsp90 substrate, even though it has little effect on its synthesis. Instead, CHIP induces ubiquitylation of the glucocorticoid receptor and degradation through the proteasome. By remodelling Hsp90 heterocomplexes to favour substrate degradation, CHIP modulates protein triage decisions that regulate the balance between protein folding and degradation for chaperone substrates. 相似文献
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7.
Heike Rampelt Sebastian R Scholz Richard I Morimoto Bernd Bukau 《The EMBO journal》2012,31(21):4221-4235
Accumulation of aggregation‐prone misfolded proteins disrupts normal cellular function and promotes ageing and disease. Bacteria, fungi and plants counteract this by solubilizing and refolding aggregated proteins via a powerful cytosolic ATP‐dependent bichaperone system, comprising the AAA+ disaggregase Hsp100 and the Hsp70‐Hsp40 system. Metazoa, however, lack Hsp100 disaggregases. We show that instead the Hsp110 member of the Hsp70 superfamily remodels the human Hsp70‐Hsp40 system to efficiently disaggregate and refold aggregates of heat and chemically denatured proteins in vitro and in cell extracts. This Hsp110 effect relies on nucleotide exchange, not on ATPase activity, implying ATP‐driven chaperoning is not required. Knock‐down of nematode Caenorhabditis elegans Hsp110, but not an unrelated nucleotide exchange factor, compromises dissolution of heat‐induced protein aggregates and severely shortens lifespan after heat shock. We conclude that in metazoa, Hsp70‐Hsp40 powered by Hsp110 nucleotide exchange represents the crucial disaggregation machinery that reestablishes protein homeostasis to counteract protein unfolding stress. 相似文献
8.
Morishima Y Kanelakis KC Silverstein AM Dittmar KD Estrada L Pratt WB 《The Journal of biological chemistry》2000,275(10):6894-6900
A system consisting of five purified proteins: Hsp90, Hsp70, Hop, Hsp40, and p23, acts as a machinery for assembly of glucocorticoid receptor (GR).Hsp90 heterocomplexes. Hop binds independently to Hsp90 and to Hsp70 to form a Hsp90.Hop.Hsp70.Hsp40 complex that is sufficient to convert the GR to its steroid binding form, and this four-protein complex will form stable GR.Hsp90 heterocomplexes if p23 is added to the system (Dittmar, K. D., Banach, M., Galigniana, M. D., and Pratt, W. B. (1998) J. Biol. Chem. 273, 7358-7366). Hop has been considered essential for the formation of receptor.Hsp90 heterocomplexes and GR folding. Here we use Hsp90 and Hsp70 purified free of all traces of Hop and Hsp40 to show that Hop is not required for GR.Hsp90 heterocomplex assembly and activation of steroid binding activity. Rather, Hop enhances the rate of the process. We also show that Hsp40 is not essential for GR folding by the five-protein system but enhances a process that occurs less effectively when it is not present. By carrying out assembly in the presence of radiolabeled steroid to bind to the GR as soon as it is converted to the steroid binding state, we show that the folding change is brought about by only two essential components, Hsp90 and Hsp70, and that Hop, Hsp40, and p23 act as nonessential co-chaperones. 相似文献
9.
Shelley A. Brunt Gary H. Perdew David O. Toft Julie C. Silver 《Cell stress & chaperones》1998,3(1):44-56
In the oomycete fungus Achlya ambisexualis, hyphae of the male strain undergo sexual differentiation in the presence of the steroid hormone antheridiol. Earlier studies demonstrated that antheridiol binds with high affinity to a 9S multiprotein complex from A. ambisexualis cytosols. Although these complexes were found to contain the heat shock protein Hsp90, the other components were not known. It was of interest to determine if any of the other protein components in the Achlya Hsp90-heterocomplexes would be homologous to those found in the steroid receptor-Hsp90-heterocomplexes of vertebrates. Cytosolic proteins of 110 kDa, 74 kDa, 64 kDa, 61 kDa, 56 kDa, 47 kDa, 27 kDa and 23 kDa, were found in repeated trials, to co-immunoprecipitate with Achlya Hsp90. The 74 kDa protein was identified as the heat shock protein Hsp70, the 23 kDa protein was found to be related to the vertebrate protein p23 and the 56 kDa protein was found to be related to immunophilin FKBP51. All three of these proteins are components of the vertebrate receptor heterocomplexes. The 110 kDa, 61 kDa and 27 kDa proteins appeared to be unique to the Achlya complexes. Unlike the seven other proteins co-immunoprecipitating with Hsp90, the 61 kDa protein was observed only in the co-immunoprecipitates produced from in vitro translates of RNA isolated from antheridiol-treated mycelia. 相似文献
10.
Hsp90 family represents a group of highly conserved and strongly expressed proteins present in almost all biological species.
Heat shock proteins in the range of 90 kDa have been detected in a range of plant species andhsp90 genes have been cloned and characterized in selected instances. However, the expression characteristics of plant Hsp90 are
poorly understood. Work on expression characteristics of rice Hsp90 is reviewed in this paper. Experimental evidence is provided
for indicating that while the rice 87 kDa protein is transiently synthesized within initial 2 h of heat shock, high steady-state
levels of this protein are retained even under prolonged high temperature stress conditions or recovery following 4 h heat
shock. It is further shown that fifteen different wild rices accumulate differential levels of these proteins in response
to heat shock treatment. 相似文献
11.
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. 相似文献
12.
Zheng Zhang Zhenyuan You Rick T. Dobrowsky Brian S.J. Blagg 《Bioorganic & medicinal chemistry letters》2018,28(16):2701-2704
KU-596 is a second-generation C-terminal heat shock protein 90 KDa (Hsp90) modulator based on the natural product, novobiocin. KU-596 has been shown to induce Hsp70 levels and manifest neuroprotective activity through induction of the heat shock response. A ring-constrained analog of KU-596 was designed and synthesized to probe its binding orientation and ability to induce Hsp70 levels. Compound 2 was found to exhibit comparable or increased activity compared to KU-596, which is under clinical investigation for the treatment of neuropathy. 相似文献
13.
Yan S Sun X Xiang B Cang H Kang X Chen Y Li H Shi G Yeh ET Wang B Wang X Yi J 《The EMBO journal》2010,29(22):3773-3786
The molecular chaperone heat shock protein 90 (Hsp90) and the co-chaperone/ubiquitin ligase carboxyl terminus of Hsc70-interacting protein (CHIP) control the turnover of client proteins. How this system decides to stabilize or degrade the client proteins under particular physiological or pathological conditions is unclear. We report here a novel client protein, the SUMO2/3 protease SENP3, that is sophisticatedly regulated by CHIP and Hsp90. SENP3 is maintained at a low basal level under non-stress condition due to Hsp90-independent CHIP-mediated ubiquitination. Upon mild oxidative stress, SENP3 undergoes thiol modification, which recruits Hsp90. Hsp90/SENP3 association protects SENP3 from CHIP-mediated ubiquitination and subsequent degradation, but this effect of Hsp90 requires the presence of CHIP. Our data demonstrate for the first time that CHIP and Hsp90 interplay with a client alternately under non-stress and stress conditions, and the choice between stabilization and degradation is made by the redox state of the client. In addition, enhanced SENP3/Hsp90 association is found in cancer. These findings provide new mechanistic insight into how cells regulate the SUMO protease in response to oxidative stress. 相似文献
14.
Szymon Żwirowski Krzysztof Liberek Szymon Żwirowski Agnieszka Kłosowska Igor Obuchowski Nadinath B Nillegoda Artur Piróg Szymon Ziętkiewicz Bernd Bukau Axel Mogk Krzysztof Liberek 《The EMBO journal》2017,36(6):783-796
Small heat shock proteins (sHsps) are an evolutionary conserved class of ATP-independent chaperones that protect cells against proteotoxic stress. sHsps form assemblies with aggregation-prone misfolded proteins, which facilitates subsequent substrate solubilization and refolding by ATP-dependent Hsp70 and Hsp100 chaperones. Substrate solubilization requires disruption of sHsp association with trapped misfolded proteins. Here, we unravel a specific interplay between Hsp70 and sHsps at the initial step of the solubilization process. We show that Hsp70 displaces surface-bound sHsps from sHsp–substrate assemblies. This Hsp70 activity is unique among chaperones and highly sensitive to alterations in Hsp70 concentrations. The Hsp70 activity is reflected in the organization of sHsp–substrate assemblies, including an outer dynamic sHsp shell that is removed by Hsp70 and a stable core comprised mainly of aggregated substrates. Binding of Hsp70 to the sHsp/substrate core protects the core from aggregation and directs sequestered substrates towards refolding pathway. The sHsp/Hsp70 interplay has major impact on protein homeostasis as it sensitizes substrate release towards cellular Hsp70 availability ensuring efficient refolding of damaged proteins under favourable folding conditions. 相似文献
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18.
Duncan RF 《Cell stress & chaperones》2008,13(2):143-155
Rapamycin inhibits the activity of the target of rapamycin (TOR)-dependent signaling pathway, which has been characterized
as one dedicated to translational regulation through modulating cap-dependent translation, involving eIF4E binding protein
(eIF4E-BP) or 4E-BP. Results show that rapamycin strongly inhibits global translation in Drosophila cells. However, Hsp70 mRNA translation is virtually unaffected by rapamycin treatment, whereas Hsp90 mRNA translation is
strongly inhibited, at normal growth temperature. Intriguingly, during heat shock Hsp90 mRNA becomes significantly less sensitive
to rapamycin-mediated inhibition, suggesting the pathway for Hsp90 mRNA translation is altered during heat shock. Reporter
mRNAs containing the Hsp90 or Hsp70 mRNAs’ 5′ untranslated region recapitulate these rapamycin-dependent translational characteristics,
indicating this region regulates rapamycin-dependent translational sensitivity as well as heat shock preferential translation.
Surprisingly, rapamycin-mediated inhibition of Hsp90 mRNA translation at normal growth temperature is not caused by 4E-BP-mediated
inhibition of cap-dependent translation. Indeed, no evidence for rapamycin-mediated impaired eIF4E function is observed. These
results support the proposal that preferential translation of different Hsp mRNA utilizes distinct translation mechanisms,
even within a single species. 相似文献
19.
Baldo B Weiss A Parker CN Bibel M Paganetti P Kaupmann K 《The Journal of biological chemistry》2012,287(2):1406-1414
Mechanisms to reduce the cellular levels of mutant huntingtin (mHtt) provide promising strategies for treating Huntington disease (HD). To identify compounds enhancing the degradation of mHtt, we performed a high throughput screen using a hippocampal HN10 cell line expressing a 573-amino acid mHtt fragment. Several hit structures were identified as heat shock protein 90 (Hsp90) inhibitors. Cell treatment with these compounds reduced levels of mHtt without overt toxic effects as measured by time-resolved Förster resonance energy transfer assays and Western blots. To characterize the mechanism of mHtt degradation, we used the potent and selective Hsp90 inhibitor NVP-AUY922. In HdhQ150 embryonic stem (ES) cells and in ES cell-derived neurons, NVP-AUY922 treatment substantially reduced soluble full-length mHtt levels. In HN10 cells, Hsp90 inhibition by NVP-AUY922 enhanced mHtt clearance in the absence of any detectable Hsp70 induction. Furthermore, inhibition of protein synthesis with cycloheximide or overexpression of dominant negative heat shock factor 1 (Hsf1) in HdhQ150 ES cells attenuated Hsp70 induction but did not affect NVP-AUY922-mediated mHtt clearance. Together, these data provided evidence that direct inhibition of Hsp90 chaperone function was crucial for mHtt degradation rather than heat shock response induction and Hsp70 up-regulation. Co-immunoprecipitation experiments revealed a physical interaction of mutant and wild-type Htt with the Hsp90 chaperone. Hsp90 inhibition disrupted the interaction and induced clearance of Htt through the ubiquitin-proteasome system. Our data suggest that Htt is an Hsp90 client protein and that Hsp90 inhibition may provide a means to reduce mHtt in HD. 相似文献